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Han X, Hu Z, Surya W, Ma Q, Zhou F, Nordenskiöld L, Torres J, Lu L, Miao Y. The intrinsically disordered region of coronins fine-tunes oligomerization and actin polymerization. Cell Rep 2023; 42:112594. [PMID: 37269287 DOI: 10.1016/j.celrep.2023.112594] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/18/2022] [Revised: 04/21/2023] [Accepted: 05/16/2023] [Indexed: 06/05/2023] Open
Abstract
Coronins play critical roles in actin network formation. The diverse functions of coronins are regulated by the structured N-terminal β propeller and the C-terminal coiled coil (CC). However, less is known about a middle "unique region" (UR), which is an intrinsically disordered region (IDR). The UR/IDR is an evolutionarily conserved signature in the coronin family. By integrating biochemical and cell biology experiments, coarse-grained simulations, and protein engineering, we find that the IDR optimizes the biochemical activities of coronins in vivo and in vitro. The budding yeast coronin IDR plays essential roles in regulating Crn1 activity by fine-tuning CC oligomerization and maintaining Crn1 as a tetramer. The IDR-guided optimization of Crn1 oligomerization is critical for F-actin cross-linking and regulation of Arp2/3-mediated actin polymerization. The final oligomerization status and homogeneity of Crn1 are contributed by three examined factors: helix packing, the energy landscape of the CC, and the length and molecular grammar of the IDR.
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Affiliation(s)
- Xiao Han
- School of Biological Sciences, Nanyang Technological University, Singapore 637551, Singapore
| | - Zixin Hu
- School of Biological Sciences, Nanyang Technological University, Singapore 637551, Singapore
| | - Wahyu Surya
- School of Biological Sciences, Nanyang Technological University, Singapore 637551, Singapore
| | - Qianqian Ma
- School of Biological Sciences, Nanyang Technological University, Singapore 637551, Singapore
| | - Feng Zhou
- School of Biological Sciences, Nanyang Technological University, Singapore 637551, Singapore
| | - Lars Nordenskiöld
- School of Biological Sciences, Nanyang Technological University, Singapore 637551, Singapore
| | - Jaume Torres
- School of Biological Sciences, Nanyang Technological University, Singapore 637551, Singapore
| | - Lanyuan Lu
- School of Biological Sciences, Nanyang Technological University, Singapore 637551, Singapore
| | - Yansong Miao
- School of Biological Sciences, Nanyang Technological University, Singapore 637551, Singapore; Institute for Digital Molecular Analytics and Science, Nanyang Technological University, Singapore 636921, Singapore.
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2
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Pooley HB, Panag G, Plain KM, de Silva K, Begg DJ, Whittington RJ, Purdie AC. IP10 is a predictor of successful vaccine protection against paratuberculosis infection in sheep. Vaccine 2023; 41:274-283. [PMID: 36456390 DOI: 10.1016/j.vaccine.2022.11.018] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/21/2022] [Revised: 11/10/2022] [Accepted: 11/13/2022] [Indexed: 11/29/2022]
Abstract
The cell mediated immune response and ability of immune cells to migrate to the site of infection are both key aspects of protection against many pathogens. Mycobacterium avium subsp. paratuberculosis (MAP) is an intracellular pathogen and the causative agent of paratuberculosis, a chronic wasting disease of ruminants. Current commercial vaccines for paratuberculosis reduce the occurrence of clinical disease but not all animals are protected from infection. Therefore, there is a need to understand the immune responses triggered by these vaccines at the site of infection, in circulating immune cells and their relationships to vaccine-mediated protection. The magnitude and location of gene expression related to the cell mediated immune response and cellular migration were studied in the ileum of sheep. In addition, longitudinal IP10 (also known as IP10) secretion by circulating immune cells was examined in the same sheep. Animals were grouped based on vaccination status (vaccinated vs non-vaccinated) and MAP exposure (experimentally exposed vs unexposed). Vaccination of unexposed sheep increased the expression of IP10, CCL5 and COR1c. Sheep that were successfully protected by vaccination (uninfected following experimental exposure) had significantly reduced expression of IP10 in the ileum at 12 months post exposure compared to vaccine non-responders (those that became infected) and non-vaccinated infected sheep. Successfully protected sheep also had significantly increased secretion of IP10 in in vitro stimulated immune cells from whole blood compared to vaccine non responders at 4 months post exposure. Therefore, the IP10 recall response has the potential to be used as marker for infection status in vaccinated sheep and could be a biomarker for a DIVA test in sheep.
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Affiliation(s)
- Hannah B Pooley
- Sydney School of Veterinary Science, Faculty of Science, The University of Sydney, Camden, NSW, Australia.
| | - Guneet Panag
- Sydney School of Veterinary Science, Faculty of Science, The University of Sydney, Camden, NSW, Australia
| | - Karren M Plain
- Sydney School of Veterinary Science, Faculty of Science, The University of Sydney, Camden, NSW, Australia
| | - Kumudika de Silva
- Sydney School of Veterinary Science, Faculty of Science, The University of Sydney, Camden, NSW, Australia
| | - Douglas J Begg
- Sydney School of Veterinary Science, Faculty of Science, The University of Sydney, Camden, NSW, Australia
| | - Richard J Whittington
- Sydney School of Veterinary Science, Faculty of Science, The University of Sydney, Camden, NSW, Australia
| | - Auriol C Purdie
- Sydney School of Veterinary Science, Faculty of Science, The University of Sydney, Camden, NSW, Australia
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3
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Mustafov D, Karteris E, Braoudaki M. Deciphering the Role of microRNA Mediated Regulation of Coronin 1C in Glioblastoma Development and Metastasis. Noncoding RNA 2023; 9:4. [PMID: 36649032 PMCID: PMC9844418 DOI: 10.3390/ncrna9010004] [Citation(s) in RCA: 4] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/02/2022] [Revised: 12/28/2022] [Accepted: 01/02/2023] [Indexed: 01/06/2023] Open
Abstract
Glioblastoma multiforme (GBM) is a highly heterogenic and malignant brain tumour with a median survival of 15 months. The initial identification of primary glioblastomas is often challenging. Coronin 1C (CORO1C) is a key player in actin rearrangement and cofilin dynamics, as well as enhancing the processes of neurite overgrowth and migration of brain tumour cells. Different bioinformatic databases were accessed to measure CORO1C expression at the mRNA and protein level in normal and malignant brains. CORO1C expression was observed in brain regions which have retained high synaptic plasticity and myelination properties. CORO1C was also expressed mainly within the hippocampus formation, including the Cornu Ammonis (CA) fields: CA1-CA4. Higher expression was also noticed in paediatric GBM in comparison to their adult counterparts. Pediatric cell populations were observed to have an increased log2 expression of CORO1C. Furthermore, 62 miRNAs were found to target the CORO1C gene. Of these, hsa-miR-34a-5p, hsa-miR-512-3p, hsa-miR-136-5p, hsa-miR-206, hsa-miR-128-3p, and hsa-miR-21-5p have shown to act as tumour suppressors or oncomiRs in different neoplasms, including GBM. The elevated expression of CORO1C in high grade metastatic brain malignancies, including GBM, suggests that this protein could have a clinical utility as a biomarker linked to an unfavorable outcome.
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Affiliation(s)
- Denis Mustafov
- School of Life and Medical Sciences, University of Hertfordshire, Hatfield AL10 9AB, UK
- College of Health, Medicine and Life Sciences, Brunel University London, Uxbridge UB8 3PH, UK
| | - Emmanouil Karteris
- College of Health, Medicine and Life Sciences, Brunel University London, Uxbridge UB8 3PH, UK
| | - Maria Braoudaki
- School of Life and Medical Sciences, University of Hertfordshire, Hatfield AL10 9AB, UK
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4
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Striepen JF, Voeltz GK. Coronin 1C restricts endosomal branched actin to organize ER contact and endosome fission. J Biophys Biochem Cytol 2022; 221:213342. [PMID: 35802042 PMCID: PMC9274145 DOI: 10.1083/jcb.202110089] [Citation(s) in RCA: 14] [Impact Index Per Article: 4.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/15/2021] [Revised: 05/03/2022] [Accepted: 06/14/2022] [Indexed: 12/15/2022] Open
Abstract
ER contact sites define the position of endosome bud fission during actin-dependent cargo sorting. Disrupting endosomal actin structures prevents retrograde cargo movement; however, how actin affects ER contact site formation and endosome fission is not known. Here we show that in contrast with the WASH complex, actin, its nucleator ARP2/3, and COR1C form a contained structure at the bud neck that defines the site of bud fission. We found that actin confinement is facilitated by type I coronins. Depletion of type I coronins allows actin to extend along the length of the bud in an ARP2/3-dependent manner. We demonstrate that extension of branched actin prevents ER recruitment and stalls buds before fission. Finally, our structure-function studies show that the COR1C’s coiled-coil domain is sufficient to restore actin confinement, ER recruitment, and endosome fission. Together, our data reveal how the dynamics of endosomal actin and activity of actin regulators organize ER-associated bud fission.
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Affiliation(s)
- Jonathan F Striepen
- Department of Molecular, Cellular, and Developmental Biology, University of Colorado, Boulder, CO.,Howard Hughes Medical Institute, Chevy Chase, MD
| | - Gia K Voeltz
- Department of Molecular, Cellular, and Developmental Biology, University of Colorado, Boulder, CO.,Howard Hughes Medical Institute, Chevy Chase, MD
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CORO1C, a novel PAK4 binding protein, recruits phospho-PAK4 at serine 99 to the leading edge and promotes the migration of gastric cancer cells. Acta Biochim Biophys Sin (Shanghai) 2022; 54:673-685. [PMID: 35593474 PMCID: PMC9827817 DOI: 10.3724/abbs.2022044] [Citation(s) in RCA: 4] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/29/2022] Open
Abstract
Gastric cancer is one of the malignant tumors in the world. PAK4 plays an important role in the occurrence and development of gastric cancer, especially in the process of invasion and metastasis. Here we discover that CORO1C, a member of coronin family that regulates microfilament and lamellipodia formation, recruits cytoplasmic PAK4 to the leading edge of gastric cancer cells by C-terminal extension (CE) domain of CORO1C (353-457 aa). The localization of PAK4 on the leading edge of the cell depends on two necessary conditions: the phosphorylation of PAK4 on serine 99 and the binding to the CE domain of CORO1C. Unphosphorylated PAK4 on serine 99 is closely associated with microtubules by PAK4/GEF-H1/Tctex-1 complex. Once phosphorylated, PAK4 is released from microtubule, and then is recruited by CORO1C to the leading edge and regulates the CORO1C/RCC2 (regulator of chromosome condensation 2) complex, leading to the migration of gastric cancer cells. Our results reveal a new mechanism by which PAK4 regulates the migration potential of gastric cancer cells through microtubule-microfilament cross talk.
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Wang L, Zeng C, Chen Z, Qi J, Huang S, Liang H, Huang S, Ou Z. Circ_0025039 acts an oncogenic role in the progression of non-small cell lung cancer through miR-636-dependent regulation of CORO1C. Mol Cell Biochem 2022; 477:743-757. [PMID: 35034254 DOI: 10.1007/s11010-021-04320-0] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/25/2021] [Accepted: 11/29/2021] [Indexed: 11/28/2022]
Abstract
Non-small cell lung cancer remains the leading cause of cancer-related death worldwide. Circular RNA plays vital roles in NSCLC progression. This study is designed to reveal the role of circ_0025039 in NSCLC cell malignancy. The RNA expression of circ_0025039, microRNA-636 (miR-636), and coronin 1C was detected by quantitative real-time polymerase chain reaction. Protein expression was checked by Western blot analysis or immunohistochemistry assay. Cell proliferation, migration, invasion, tube formation ability, sphere formation capacity, and apoptosis were investigated by cell counting kit-8, 5-Ethynyl-29-deoxyuridine, transwell assay, tube formation assay, sphere formation assay, and flow cytometry analysis, respectively. Mouse model assay was conducted to reveal the effect of circ_0025039 silencing on tumor formation in vivo. The interaction between miR-636 and circ_0025039 or CORO1C was identified through dual-luciferase reporter and RNA pull-down assays. The expression of circ_0025039 and CORO1C was significantly increased, while miR-636 was decreased in NSCLC tissues and cells compared with controls. Circ_0025039 depletion repressed NSCLC cell proliferation, migration, invasion, tube-forming capacity, and sphere formation ability, but induced cell apoptosis. The neoplasm formation was repressed after circ_0025039 silencing. Additionally, circ_0025039 acted as a sponge for miR-636, which was found to target CORO1C. Importantly, the contribution of circ_0025039 to NSCLC progression was mediated by miR-636/CORO1C axis. Circ_0025039 silencing repressed NSCLC malignant progression by reducing CORO1C expression through miR-636, showing the possibility of circ_0025039 as a therapeutic target for NSCLC.
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Affiliation(s)
- Lei Wang
- Department of Respiratory and Critical Care Medicine, Affiliated Haikou Hospital of Xiangya Medical College, Central South University, No. 43, Renmin Avenue, Meilan District, Haikou City, Hainan Province, 570208, PR China
| | - Cimei Zeng
- Department of Respiratory and Critical Care Medicine, Affiliated Haikou Hospital of Xiangya Medical College, Central South University, No. 43, Renmin Avenue, Meilan District, Haikou City, Hainan Province, 570208, PR China
| | - Zhongren Chen
- Department of Respiratory and Critical Care Medicine, Affiliated Haikou Hospital of Xiangya Medical College, Central South University, No. 43, Renmin Avenue, Meilan District, Haikou City, Hainan Province, 570208, PR China
| | - Jianxu Qi
- Department of Respiratory and Critical Care Medicine, Affiliated Haikou Hospital of Xiangya Medical College, Central South University, No. 43, Renmin Avenue, Meilan District, Haikou City, Hainan Province, 570208, PR China
| | - Sini Huang
- Department of Respiratory and Critical Care Medicine, Affiliated Haikou Hospital of Xiangya Medical College, Central South University, No. 43, Renmin Avenue, Meilan District, Haikou City, Hainan Province, 570208, PR China
| | - Haimei Liang
- Department of Respiratory and Critical Care Medicine, Affiliated Haikou Hospital of Xiangya Medical College, Central South University, No. 43, Renmin Avenue, Meilan District, Haikou City, Hainan Province, 570208, PR China
| | - Shiren Huang
- Department of Respiratory and Critical Care Medicine, Affiliated Haikou Hospital of Xiangya Medical College, Central South University, No. 43, Renmin Avenue, Meilan District, Haikou City, Hainan Province, 570208, PR China
| | - Zongxing Ou
- Department of Respiratory and Critical Care Medicine, Affiliated Haikou Hospital of Xiangya Medical College, Central South University, No. 43, Renmin Avenue, Meilan District, Haikou City, Hainan Province, 570208, PR China.
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de Lima JB, da Silva Fonseca LP, Xavier LP, de Matos Macchi B, Cassoli JS, da Silva EO, da Silva Valadares RB, do Nascimento JLM, Santos AV, de Sena CBC. Culture of Mycobacterium smegmatis in Different Carbon Sources to Induce In Vitro Cholesterol Consumption Leads to Alterations in the Host Cells after Infection: A Macrophage Proteomics Analysis. Pathogens 2021; 10:pathogens10060662. [PMID: 34071265 PMCID: PMC8230116 DOI: 10.3390/pathogens10060662] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/30/2021] [Revised: 05/18/2021] [Accepted: 05/26/2021] [Indexed: 11/24/2022] Open
Abstract
During tuberculosis, Mycobacterium uses host macrophage cholesterol as a carbon and energy source. To mimic these conditions, Mycobacterium smegmatis can be cultured in minimal medium (MM) to induce cholesterol consumption in vitro. During cultivation, M. smegmatis consumes MM cholesterol and changes the accumulation of cell wall compounds, such as PIMs, LM, and LAM, which plays an important role in its pathogenicity. These changes lead to cell surface hydrophobicity modifications and H2O2 susceptibility. Furthermore, when M. smegmatis infects J774A.1 macrophages, it induces granuloma-like structure formation. The present study aims to assess macrophage molecular disturbances caused by M. smegmatis after cholesterol consumption, using proteomics analyses. Proteins that showed changes in expression levels were analyzed in silico using OmicsBox and String analysis to investigate the canonical pathways and functional networks involved in infection. Our results demonstrate that, after cholesterol consumption, M. smegmatis can induce deregulation of protein expression in macrophages. Many of these proteins are related to cytoskeleton remodeling, immune response, the ubiquitination pathway, mRNA processing, and immunometabolism. The identification of these proteins sheds light on the biochemical pathways involved in the mechanisms of action of mycobacteria infection, and may suggest novel protein targets for the development of new and improved treatments.
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Affiliation(s)
- Jaqueline Batista de Lima
- Laboratory of Structural Biology, Institute of Biological Sciences, Federal University of Pará, Belém 66075-110, PA, Brazil; (J.B.d.L.); (E.O.d.S.)
- Laboratory of Biotechnology of Enzymes and Biotransformations, Institute of Biological Sciences, Federal University of Pará, Belém 66075-110, PA, Brazil; (L.P.X.); (A.V.S.)
| | | | - Luciana Pereira Xavier
- Laboratory of Biotechnology of Enzymes and Biotransformations, Institute of Biological Sciences, Federal University of Pará, Belém 66075-110, PA, Brazil; (L.P.X.); (A.V.S.)
| | - Barbarella de Matos Macchi
- Laboratory of Molecular and Cellular Neurochemistry, Institute of Biological Sciences, Federal University of Pará, Belém 66075-110, PA, Brazil; (B.d.M.M.); (J.L.M.d.N.)
- National Institute of Science and Technology in Neuroimmunomodulation (INCT-NIM), Rio de Janeiro 21040-900, RJ, Brazil
| | - Juliana Silva Cassoli
- Institute of Biological Sciences, Federal University of Pará, Belém 66075-110, PA, Brazil;
| | - Edilene Oliveira da Silva
- Laboratory of Structural Biology, Institute of Biological Sciences, Federal University of Pará, Belém 66075-110, PA, Brazil; (J.B.d.L.); (E.O.d.S.)
- National Institute of Science and Technology in Structural Biology and Bioimaging, Rio de Janeiro 21941-901, RJ, Brazil
| | | | - José Luiz Martins do Nascimento
- Laboratory of Molecular and Cellular Neurochemistry, Institute of Biological Sciences, Federal University of Pará, Belém 66075-110, PA, Brazil; (B.d.M.M.); (J.L.M.d.N.)
- National Institute of Science and Technology in Neuroimmunomodulation (INCT-NIM), Rio de Janeiro 21040-900, RJ, Brazil
| | - Agenor Valadares Santos
- Laboratory of Biotechnology of Enzymes and Biotransformations, Institute of Biological Sciences, Federal University of Pará, Belém 66075-110, PA, Brazil; (L.P.X.); (A.V.S.)
| | - Chubert Bernardo Castro de Sena
- Laboratory of Structural Biology, Institute of Biological Sciences, Federal University of Pará, Belém 66075-110, PA, Brazil; (J.B.d.L.); (E.O.d.S.)
- National Institute of Science and Technology in Neuroimmunomodulation (INCT-NIM), Rio de Janeiro 21040-900, RJ, Brazil
- Correspondence:
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8
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Han S, Ding X, Wang S, Xu L, Li W, Sun W. miR-133a-3p Regulates Hepatocellular Carcinoma Progression Through Targeting CORO1C. Cancer Manag Res 2020; 12:8685-8693. [PMID: 33061567 PMCID: PMC7519587 DOI: 10.2147/cmar.s254617] [Citation(s) in RCA: 10] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/19/2020] [Accepted: 08/05/2020] [Indexed: 12/14/2022] Open
Abstract
Introduction MicroRNAs (miRNAs) are key modulators for gene expression via inducing translational repression or target gene degradation. miR-133a-3p was reported to stimulate or inhibit cancer progression but its role in hepatocellular carcinoma (HCC) remains to be explored. Methods Quantitative real-time PCR (RT-qPCR) was utilized to explore miR-133a-3p expression level in HCC cells. Dual-luciferase activity reporter assay was used to validate the direct interaction between miR-133a-3p and coronin-like actin-binding protein 1C (CORO1C). In addition, we analyzed the expression levels of miR-133a-3p and CORO1C in HCC tissues and normal tissues on the UCALAN website. Functional assays including cell counting kit-8 assay, colony formation assay, flow cytometry analysis and transwell invasion assay were conducted to explore the biological functions of miR-133a-3p in HCC. Results miR-133a-3p was found to have downregulated expression in HCC tissues and cells. Meanwhile, we showed that low miR-133a-3p levels were correlated with poorer overall survival of HCC patients. Overexpression of miR-133a-3p suppressed HCC cell growth and invasion but promoted cell apoptosis via targeting CORO1C. Discussion Our results revealed a novel mechanism of miR-133a-3p in regulating HCC progression and provided evidence that miR-133a-3p functions as a tumor suppressor in HCC.
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Affiliation(s)
- Shuangxi Han
- Department of Hepatobiliary Surgery, Beijing Chao-Yang Hospital, Capital Medical University, Beijing 100043, People's Republic of China.,Department of Hepatobiliary Surgery, Binzhou Central Hospital, Binzhou 251700, People's Republic of China
| | - Xuemei Ding
- Department of Hepatobiliary Surgery, Beijing Chao-Yang Hospital, Capital Medical University, Beijing 100043, People's Republic of China
| | - Shaohong Wang
- Department of Hepatobiliary Surgery, Beijing Chao-Yang Hospital, Capital Medical University, Beijing 100043, People's Republic of China
| | - Li Xu
- Department of Hepatobiliary Surgery, Beijing Chao-Yang Hospital, Capital Medical University, Beijing 100043, People's Republic of China
| | - Wenxiao Li
- Department of Hepatobiliary Surgery, Beijing Chao-Yang Hospital, Capital Medical University, Beijing 100043, People's Republic of China
| | - Wenbing Sun
- Department of Hepatobiliary Surgery, Beijing Chao-Yang Hospital, Capital Medical University, Beijing 100043, People's Republic of China
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Wang M, Li Q, Yu S, Zhang Z, Qiu P, Zhang Y, Yang W, Xu G, Xu T. Coronin 3 Promotes the Development of Oncogenic Properties in Glioma Through the Wnt/β-Catenin Signaling Pathway. Onco Targets Ther 2020; 13:6661-6673. [PMID: 32764958 PMCID: PMC7371924 DOI: 10.2147/ott.s257001] [Citation(s) in RCA: 6] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/16/2020] [Accepted: 06/25/2020] [Indexed: 11/26/2022] Open
Abstract
PURPOSE Evidence indicates that the actin-binding protein Coronin 3, which is aberrantly expressed in various cancers, is associated with cancer development and progression. However, little is known about the role of Coronin 3 in glioma tumorigenesis. Here, we aimed to explore the biological function and regulatory mechanism of Coronin 3 in glioblastoma (GBM). MATERIALS AND METHODS Coronin 3 level in human GBM clinical samples and cell lines was investigated. The shRNA knockdown strategy was used to assess the tumor characteristics of GBM cell lines. The role of β-catenin in Coronin 3-mediated oncogenic phenotypes was evaluated. RESULTS Coronin 3 was found to be highly upregulated in glioma cell lines. Furthermore, knockdown of Coronin 3 significantly inhibited the growth of glioma cells both in vivo and in vitro and suppressed the expression of Wnt/β-catenin pathway genes, including β-catenin, Cyclin D1, and c-Myc. Moreover, we demonstrated that Coronin 3 regulates the expression of β-catenin in glioma. Our results revealed that Coronin 3-stimulated tumor growth was β-catenin-dependent. CONCLUSION Our study reveals a new molecular mechanism of Coronin 3 in promoting glioma growth and development through regulating the Wnt/β-catenin signaling pathway.
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Affiliation(s)
- Min Wang
- Department of Neurosurgery, Shandong Provincial Hospital Affiliated to Shandong First Medical University, Jinan, Shandong250014, People’s Republic of China
| | - Qi Li
- Department of Neurosurgery, Shandong Provincial Hospital Affiliated to Shandong First Medical University, Jinan, Shandong250014, People’s Republic of China
| | - Shengyuan Yu
- Department of Neurosurgery, Shandong Provincial Hospital Affiliated to Shandong First Medical University, Jinan, Shandong250014, People’s Republic of China
| | - Zexiang Zhang
- Department of Gastrointestinal Surgery, Shandong Provincial Hospital Affiliated to Shandong First Medical University, Jinan, Shandong250014, People’s Republic of China
| | - Peng Qiu
- Department of Neurosurgery, Shandong Provincial Hospital Affiliated to Shandong First Medical University, Jinan, Shandong250014, People’s Republic of China
| | - Yubao Zhang
- Department of Neurosurgery, Shandong Provincial Hospital Affiliated to Shandong First Medical University, Jinan, Shandong250014, People’s Republic of China
| | - Wei Yang
- Department of Neurosurgery, Shandong Provincial Hospital Affiliated to Shandong First Medical University, Jinan, Shandong250014, People’s Republic of China
| | - Guangming Xu
- Department of Neurosurgery, Shandong Provincial Hospital Affiliated to Shandong First Medical University, Jinan, Shandong250014, People’s Republic of China
| | - Tongjiang Xu
- Department of Neurosurgery, Shandong Provincial Hospital Affiliated to Shandong First Medical University, Jinan, Shandong250014, People’s Republic of China
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10
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Fiedler T, Fabrice TN, Studer V, Vinet A, Faltova L, Kammerer RA, Steinmetz MO, Sharpe T, Pieters J. Homodimerization of coronin A through the C-terminal coiled-coil domain is essential for multicellular differentiation of Dictyostelium discoideum. FEBS Lett 2020; 594:2116-2127. [PMID: 32298460 DOI: 10.1002/1873-3468.13787] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/22/2019] [Revised: 03/25/2020] [Accepted: 03/26/2020] [Indexed: 11/09/2022]
Abstract
Coronin proteins are widely expressed among eukaryotic organisms. Most coronins consist of a WD-repeat domain followed by a C-terminal coiled coil. Dictyostelium discoideum expresses a single short coronin coronin A, which has been implicated in both actin modulation and multicellular differentiation. Whether coronin A's coiled coil is important for functionality, as well as the oligomeric state of coronin A is not known. Here, we show that the coiled-coil domain in Dictyostelium coronin A functions in homodimerization, is dispensable for coronin A stability and localization but essential for multicellular differentiation. These results allow a better understanding of the role for the coiled-coil domain of coronin A in oligomerization and demonstrate that its presence is essential for multicellular differentiation.
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Affiliation(s)
| | | | - Vera Studer
- Biozentrum, University of Basel, Switzerland
| | | | - Lenka Faltova
- Laboratory of Biomolecular Research, Division of Biology and Chemistry, Paul Scherrer Institut, Villigen, Switzerland
| | - Richard A Kammerer
- Laboratory of Biomolecular Research, Division of Biology and Chemistry, Paul Scherrer Institut, Villigen, Switzerland
| | - Michel O Steinmetz
- Biozentrum, University of Basel, Switzerland
- Laboratory of Biomolecular Research, Division of Biology and Chemistry, Paul Scherrer Institut, Villigen, Switzerland
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Riley DRJ, Khalil JS, Naseem KM, Rivero F. Biochemical and immunocytochemical characterization of coronins in platelets. Platelets 2019; 31:913-924. [PMID: 31801396 PMCID: PMC7497283 DOI: 10.1080/09537104.2019.1696457] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/01/2022]
Abstract
Rapid reorganization of the actin cytoskeleton in response to receptor-mediated signaling cascades allows platelets to transition from a discoid shape to a flat spread shape upon adhesion to damaged vessel walls. Coronins are conserved regulators of the actin cytoskeleton turnover but they also participate in signaling events. To gain a better picture of their functions in platelets we have undertaken a biochemical and immunocytochemical investigation with a focus on Coro1. We found that class I coronins Coro1, 2 and 3 are abundant in human and mouse platelets whereas little Coro7 can be detected. Coro1 is mainly cytosolic, but a significant amount associates with membranes in an actin-independent manner and does not translocate from or to the membrane fraction upon exposure to thrombin, collagen or prostacyclin. Coro1 rapidly translocates to the Triton insoluble cytoskeleton upon platelet stimulation with thrombin or collagen. Coro1, 2 and 3 show a diffuse cytoplasmic localization with discontinuous accumulation at the cell cortex and actin nodules of human platelets, where all three coronins colocalize. Our data are consistent with a role of coronins as integrators of extracellular signals with actin remodeling and suggests a high extent of functional overlap among class I coronins in platelets.
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Affiliation(s)
- David R J Riley
- Centre for Atherothrombosis and Metabolic Disease, Hull York Medical School, Faculty of Health Sciences, University of Hull , Hull, UK
| | - Jawad S Khalil
- Centre for Atherothrombosis and Metabolic Disease, Hull York Medical School, Faculty of Health Sciences, University of Hull , Hull, UK.,School of Physiology, Pharmacology and Neuroscience, Faculty of Life Sciences, University of Bristol , Bristol, UK
| | - Khalid M Naseem
- Leeds Institute for Cardiovascular and Metabolic Medicine, University of Leeds , Leeds, UK
| | - Francisco Rivero
- Centre for Atherothrombosis and Metabolic Disease, Hull York Medical School, Faculty of Health Sciences, University of Hull , Hull, UK
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12
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Solga R, Behrens J, Ziemann A, Riou A, Berwanger C, Becker L, Garrett L, de Angelis MH, Fischer L, Coras R, Barkovits K, Marcus K, Mahabir E, Eichinger L, Schröder R, Noegel AA, Clemen CS. CRN2 binds to TIMP4 and MMP14 and promotes perivascular invasion of glioblastoma cells. Eur J Cell Biol 2019; 98:151046. [PMID: 31677819 DOI: 10.1016/j.ejcb.2019.151046] [Citation(s) in RCA: 6] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/04/2019] [Revised: 09/13/2019] [Accepted: 09/30/2019] [Indexed: 12/21/2022] Open
Abstract
CRN2 is an actin filament binding protein involved in the regulation of various cellular processes including cell migration and invasion. CRN2 has been implicated in the malignant progression of different types of human cancer. We used CRN2 knock-out mice for analyses as well as for crossbreeding with a Tp53/Pten knock-out glioblastoma mouse model. CRN2 knock-out mice were subjected to a phenotyping screen at the German Mouse Clinic. Murine glioblastoma tissue specimens as well as cultured murine brain slices and glioblastoma cell lines were investigated by immunohistochemistry, immunofluorescence, and cell biological experiments. Protein interactions were studied by immunoprecipitation, pull-down, and enzyme activity assays. CRN2 knock-out mice displayed neurological and behavioural alterations, e.g. reduced hearing sensitivity, reduced acoustic startle response, hypoactivity, and less frequent urination. While glioblastoma mice with or without the additional CRN2 knock-out allele exhibited no significant difference in their survival rates, the increased levels of CRN2 in transplanted glioblastoma cells caused a higher tumour cell encasement of murine brain slice capillaries. We identified two important factors of the tumour microenvironment, the tissue inhibitor of matrix metalloproteinase 4 (TIMP4) and the matrix metalloproteinase 14 (MMP14, synonym: MT1-MMP), as novel binding partners of CRN2. All three proteins mutually interacted and co-localised at the front of lamellipodia, and CRN2 was newly detected in exosomes. On the functional level, we demonstrate that CRN2 increased the secretion of TIMP4 as well as the catalytic activity of MMP14. Our results imply that CRN2 represents a pro-invasive effector within the tumour cell microenvironment of glioblastoma multiforme.
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Affiliation(s)
- Roxana Solga
- Centre for Biochemistry, Institute of Biochemistry I, Medical Faculty, University of Cologne, 50931, Cologne, Germany
| | - Juliane Behrens
- Centre for Biochemistry, Institute of Biochemistry I, Medical Faculty, University of Cologne, 50931, Cologne, Germany
| | - Anja Ziemann
- Centre for Biochemistry, Institute of Biochemistry I, Medical Faculty, University of Cologne, 50931, Cologne, Germany
| | - Adrien Riou
- In-vivo NMR, Max Planck Institute for Metabolism Research, 50931, Cologne, Germany
| | - Carolin Berwanger
- Centre for Biochemistry, Institute of Biochemistry I, Medical Faculty, University of Cologne, 50931, Cologne, Germany; Institute of Aerospace Medicine, German Aerospace Center (DLR), 51147, Cologne, Germany
| | - Lore Becker
- German Mouse Clinic, Institute of Experimental Genetics, Helmholtz Centre Munich, German Research Centre for Environmental Health, 85764, Neuherberg, Germany
| | - Lillian Garrett
- German Mouse Clinic, Institute of Experimental Genetics, Helmholtz Centre Munich, German Research Centre for Environmental Health, 85764, Neuherberg, Germany; Institute of Developmental Genetics, Helmholtz Zentrum München, German Research Center for Environmental Health, 85764, Neuherberg, Germany
| | - Martin Hrabe de Angelis
- German Mouse Clinic, Institute of Experimental Genetics, Helmholtz Centre Munich, German Research Centre for Environmental Health, 85764, Neuherberg, Germany; Chair of Experimental Genetics, School of Life Science Weihenstephan, Technische Universität München, 85354, Freising, Germany; German Center for Diabetes Research (DZD), 85764, Neuherberg, Germany
| | - Lisa Fischer
- Comparative Medicine, Center for Molecular Medicine Cologne, University of Cologne, 50931, Cologne, Germany
| | - Roland Coras
- Institute of Neuropathology, University Hospital Erlangen, Friedrich-Alexander University Erlangen-Nürnberg, 91054, Erlangen, Germany
| | - Katalin Barkovits
- Medizinisches Proteom‑Center, Medical Faculty, Ruhr-University Bochum, 44801, Bochum, Germany
| | - Katrin Marcus
- Medizinisches Proteom‑Center, Medical Faculty, Ruhr-University Bochum, 44801, Bochum, Germany
| | - Esther Mahabir
- Comparative Medicine, Center for Molecular Medicine Cologne, University of Cologne, 50931, Cologne, Germany
| | - Ludwig Eichinger
- Centre for Biochemistry, Institute of Biochemistry I, Medical Faculty, University of Cologne, 50931, Cologne, Germany
| | - Rolf Schröder
- Institute of Neuropathology, University Hospital Erlangen, Friedrich-Alexander University Erlangen-Nürnberg, 91054, Erlangen, Germany
| | - Angelika A Noegel
- Centre for Biochemistry, Institute of Biochemistry I, Medical Faculty, University of Cologne, 50931, Cologne, Germany.
| | - Christoph S Clemen
- Centre for Biochemistry, Institute of Biochemistry I, Medical Faculty, University of Cologne, 50931, Cologne, Germany; Institute of Neuropathology, University Hospital Erlangen, Friedrich-Alexander University Erlangen-Nürnberg, 91054, Erlangen, Germany; Center for Physiology and Pathophysiology, Institute of Vegetative Physiology, Medical Faculty, University of Cologne, 50931, Cologne, Germany.
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13
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Fan L, Wei Y, Ding X, Li B. Coronin3 Promotes Nasopharyngeal Carcinoma Migration And Invasion By Induction Of Epithelial-To-Mesenchymal Transition. Onco Targets Ther 2019; 12:9585-9598. [PMID: 32009795 PMCID: PMC6859123 DOI: 10.2147/ott.s215674] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/15/2019] [Accepted: 10/04/2019] [Indexed: 12/17/2022] Open
Abstract
Purpose Coronin3 is a cytoskeletal protein that has been implicated in metastasis in many cancer types. Here, we demonstrate its effect in nasopharyngeal carcinoma (NPC) and propose a new probable mechanism of CORO1C-mediated cell migration and invasion by regulation of epithelial-to-mesenchymal transition (EMT) and CDH11. Patients and methods First, we measured the differential expression of CORO1C between NPC and non-NPC cells in both cell lines and clinical specimens, using public datasets. Then, we investigated its relationship with clinicopathological factors and its potential as a biomarker to predict the prognosis of NPC patients. We also explored its influence on the cell behaviors of migration and invasion by upregulating and downregulating the expression of CORO1C and attempted to determine the underlying mechanism. Results The results verified our original hypothesis. CORO1C was overexpressed in both NPC cell lines and clinical specimens, in both public datasets and our own samples. NPC patients with lower CORO1C expression levels in primary cancer tissues had longer OS (hazard ratio [HR] 1.814, 95% CI 0.831–3.960, p=0.0341) and PFS (HR 1.798, 95% CI 0.907–3.564, p=0.0155), indicating that it could be used as a prognostic biomarker. It was also confirmed that CORO1C enhanced cells’ migration and invasion abilities, by inducing morphological and marker changes typical of EMT. Finally, we found that expression was correlated with and regulated CDH11 expression in NPC cell lines. Conclusion Our study provided evidence for the contribution of CORO1C to NPC metastasis, and indicated that it could be used as a new therapeutic target and prognostic biomarker.
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Affiliation(s)
- Liyuan Fan
- Department of Radiation Oncology, Shandong Cancer Hospital and Institute, Shandong First Medical University and Shandong Academy of Medical Sciences, Jinan, Shandong 250117, People's Republic of China.,Cheeloo College of Medicine, Shandong University, Jinan, Shandong 250112, People's Republic of China
| | - Yumei Wei
- Department of Head and Neck Radiotherapy, Shandong Provincial ENT Hospital, Shandong Provincial ENT Hospital Affiliated to Shandong University, Jinan, Shandong 250022, People's Republic of China.,Key Laboratory of Otorhinolaryngology, National Health Commission, Shandong University, Jinan, Shandong 250022, People's Republic of China
| | - Xiuping Ding
- Department of Radiation Oncology, Shandong Cancer Hospital and Institute, Shandong First Medical University and Shandong Academy of Medical Sciences, Jinan, Shandong 250117, People's Republic of China
| | - Baosheng Li
- Department of Radiation Oncology, Shandong Cancer Hospital and Institute, Shandong First Medical University and Shandong Academy of Medical Sciences, Jinan, Shandong 250117, People's Republic of China.,Cheeloo College of Medicine, Shandong University, Jinan, Shandong 250112, People's Republic of China
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14
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Cheng X, Wang X, Wu Z, Tan S, Zhu T, Ding K. CORO1C expression is associated with poor survival rates in gastric cancer and promotes metastasis in vitro. FEBS Open Bio 2019; 9:1097-1108. [PMID: 30974047 PMCID: PMC6551501 DOI: 10.1002/2211-5463.12639] [Citation(s) in RCA: 27] [Impact Index Per Article: 4.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/14/2019] [Revised: 03/13/2019] [Accepted: 04/08/2019] [Indexed: 12/21/2022] Open
Abstract
Coronin-like actin-binding protein 1C (CORO1C) is a member of the WD repeat protein family that regulates actin-dependent processes by assembling F-actin. CORO1C was previously reported to promote metastasis in breast cancer and lung squamous cell carcinoma. Here, we investigated the role of CORO1C in gastric cancer. Higher expression levels of CORO1C were detected in gastric cancer tissues as compared with normal gastric tissues. In addition, CORO1C levels were found to be positively correlated with lymph node metastasis in gastric cancer patients. The expression levels of CORO1C were higher in stage III-IV gastric cancer patients (80.8%) than in stage I-II gastric cancer patients(57.1%). Gastric cancer patients positive for CORO1C expression showed lower relapse-free survival and overall survival rates. Knockdown of CORO1C dramatically suppressed total cell number, cell viability, cell colony formation, cell mitosis and cell metastasis, and promoted apoptosis of gastric cancer cells. Furthermore, cyclin D1 and vimentin were found to be positively regulated by CORO1C. As cyclin D1 and vimentin play an oncogenic role in gastric cancer, CORO1C may exert its tumor-promoting activity through these proteins.
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Affiliation(s)
- Xiao Cheng
- Department of Pathology, Anhui Medical University, Hefei, China
| | - Xiaonan Wang
- Laboratory of Pathogenic Microbiology and Immunology, Anhui Medical University, Hefei, China
| | - Zhengsheng Wu
- Department of Pathology, Anhui Medical University, Hefei, China
| | - Sheng Tan
- Hefei National Laboratory for Physical Sciences at Microscale, University of Science and Technology of China, Hefei, China
| | - Tao Zhu
- Hefei National Laboratory for Physical Sciences at Microscale, University of Science and Technology of China, Hefei, China
| | - Keshuo Ding
- Department of Pathology, Anhui Medical University, Hefei, China
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15
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Coronin 1C promotes triple-negative breast cancer invasiveness through regulation of MT1-MMP traffic and invadopodia function. Oncogene 2018; 37:6425-6441. [PMID: 30065298 DOI: 10.1038/s41388-018-0422-x] [Citation(s) in RCA: 32] [Impact Index Per Article: 4.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/02/2018] [Revised: 06/07/2018] [Accepted: 06/22/2018] [Indexed: 01/11/2023]
Abstract
Membrane type 1-matrix metalloproteinase (MT1-MMP), a membrane-tethered protease, is key for matrix breakdown during cancer invasion and metastasis. Assembly of branched actin networks by the Arp2/3 complex is required for MT1-MMP traffic and formation of matrix-degradative invadopodia. Contrasting with the well-established role of actin filament branching factor cortactin in invadopodia function during cancer cell invasion, the contribution of coronin-family debranching factors to invadopodia-based matrix remodeling is not known. Here, we investigated the contribution of coronin 1C to the invasive potential of breast cancer cells. We report that expression of coronin 1C is elevated in invasive human breast cancers, correlates positively with MT1-MMP expression in relation with increased metastatic risk and is a new independent prognostic factor in breast cancer. We provide evidence that, akin to cortactin, coronin 1C is required for invadopodia formation and matrix degradation by breast cancer cells lines and for 3D collagen invasion by multicellular spheroids. Using intravital imaging of orthotopic human breast tumor xenografts, we find that coronin 1C accumulates in structures forming in association with collagen fibrils in the tumor microenvironment. Moreover, we establish the role of coronin 1C in the regulation of positioning and trafficking of MT1-MMP-positive endolysosomes. These results identify coronin 1C as a novel player of the multi-faceted mechanism responsible for invadopodia formation, MT1-MMP surface exposure and invasiveness in breast cancer cells.
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16
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Knüppel L, Heinzelmann K, Lindner M, Hatz R, Behr J, Eickelberg O, Staab-Weijnitz CA. FK506-binding protein 10 (FKBP10) regulates lung fibroblast migration via collagen VI synthesis. Respir Res 2018; 19:67. [PMID: 29673351 PMCID: PMC5909279 DOI: 10.1186/s12931-018-0768-1] [Citation(s) in RCA: 21] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/08/2018] [Accepted: 04/02/2018] [Indexed: 02/07/2023] Open
Abstract
Background In idiopathic pulmonary fibrosis (IPF), fibroblasts gain a more migratory phenotype and excessively secrete extracellular matrix (ECM), ultimately leading to alveolar scarring and progressive dyspnea. Here, we analyzed the effects of deficiency of FK506-binding protein 10 (FKBP10), a potential IPF drug target, on primary human lung fibroblast (phLF) adhesion and migration. Methods Using siRNA, FKBP10 expression was inhibited in phLF in absence or presence of 2ng/ml transforming growth factor-β1 (TGF-β1) and 0.1mM 2-phosphoascorbate. Effects on cell adhesion and migration were monitored by an immunofluorescence (IF)-based attachment assay, a conventional scratch assay, and single cell tracking by time-lapse microscopy. Effects on expression of key players in adhesion dynamics and migration were analyzed by qPCR and Western Blot. Colocalization was evaluated by IF microscopy and by proximity ligation assays. Results FKBP10 knockdown significantly attenuated adhesion and migration of phLF. Expression of collagen VI was decreased, while expression of key components of the focal adhesion complex was mostly upregulated. The effects on migration were 2-phosphoascorbate-dependent, suggesting collagen synthesis as the underlying mechanism. FKBP10 colocalized with collagen VI and coating culture dishes with collagen VI, and to a lesser extent with collagen I, abolished the effect of FKBP10 deficiency on migration. Conclusions These findings show, to our knowledge for the first time, that FKBP10 interacts with collagen VI and that deficiency of FKBP10 reduces phLF migration mainly by downregulation of collagen VI synthesis. The results strengthen FKBP10 as an important intracellular regulator of ECM remodeling and support the concept of FKBP10 as drug target in IPF.
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Affiliation(s)
- Larissa Knüppel
- Comprehensive Pneumology Center, Ludwig-Maximilians-Universität and Helmholtz Zentrum Munich, Max-Lebsche-Platz 31, 81377, Munich, Germany.,Member of the German Center of Lung Research (DZL), Munich, Germany
| | - Katharina Heinzelmann
- Comprehensive Pneumology Center, Ludwig-Maximilians-Universität and Helmholtz Zentrum Munich, Max-Lebsche-Platz 31, 81377, Munich, Germany.,Member of the German Center of Lung Research (DZL), Munich, Germany
| | | | - Rudolf Hatz
- Asklepios Fachkliniken Munich-Gauting, Munich, Germany.,Thoraxchirurgisches Zentrum, Klinik für Allgemeine-, Viszeral-, Transplantations-, Gefäß- und Thoraxchirurgie, Klinikum Großhadern, Ludwig-Maximilians-Universität, Munich, Germany
| | - Jürgen Behr
- Asklepios Fachkliniken Munich-Gauting, Munich, Germany.,Medizinische Klinik und Poliklinik V, Klinikum der Ludwig-Maximilians-Universität, Munich, Germany
| | - Oliver Eickelberg
- Comprehensive Pneumology Center, Ludwig-Maximilians-Universität and Helmholtz Zentrum Munich, Max-Lebsche-Platz 31, 81377, Munich, Germany.,Member of the German Center of Lung Research (DZL), Munich, Germany.,Colorado Anschutz Medical Campus, Pulmonary and Critical Care Medicine University, Denver, Colorado, USA
| | - Claudia A Staab-Weijnitz
- Comprehensive Pneumology Center, Ludwig-Maximilians-Universität and Helmholtz Zentrum Munich, Max-Lebsche-Platz 31, 81377, Munich, Germany. .,Member of the German Center of Lung Research (DZL), Munich, Germany.
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17
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Yamaoka M, Ishizaki T, Kimura T. GTP- and GDP-Dependent Rab27a Effectors in Pancreatic Beta-Cells. Biol Pharm Bull 2016; 38:663-8. [PMID: 25947911 DOI: 10.1248/bpb.b14-00886] [Citation(s) in RCA: 8] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/22/2022]
Abstract
Small guanosine triphosphatases (GTPases) participate in a wide variety of cellular functions including proliferation, differentiation, adhesion, and intracellular transport. Conventionally, only the guanosine 5'-triphosphate (GTP)-bound small GTPase interacts with effector proteins, and the resulting downstream signals control specific cellular functions. Therefore, the GTP-bound form is regarded as active, and the focus has been on searching for proteins that bind the GTP form to look for their effectors. The Rab family small GTPase Rab27a is highly expressed in some secretory cells and is involved in the control of membrane traffic. The present study reviews recent progress in our understanding of the roles of Rab27a and its effectors in pancreatic beta-cells. In the basal state, GTP-bound Rab27a controls insulin secretion at pre-exocytic stages via its GTP-dependent effectors. We previously identified novel guanosine 5'-diphosphate (GDP)-bound Rab27-interacting proteins. Interestingly, GDP-bound Rab27a controls endocytosis of the secretory membrane via its interaction with these proteins. We also demonstrated that the insulin secretagogue glucose converts Rab27a from its GTP- to GDP-bound forms. Thus, GTP- and GDP-bound Rab27a regulate pre-exocytic and endocytic stages in membrane traffic, respectively. Since the physiological importance of GDP-bound GTPases has been largely overlooked, we consider that the investigation of GDP-dependent effectors for other GTPases is necessary for further understanding of cellular function.
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Affiliation(s)
- Mami Yamaoka
- Department of Pharmacology, Oita University Faculty of Medicine
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18
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Behrens J, Solga R, Ziemann A, Rastetter RH, Berwanger C, Herrmann H, Noegel AA, Clemen CS. Coronin 1C-free primary mouse fibroblasts exhibit robust rearrangements in the orientation of actin filaments, microtubules and intermediate filaments. Eur J Cell Biol 2016; 95:239-51. [PMID: 27178841 DOI: 10.1016/j.ejcb.2016.04.004] [Citation(s) in RCA: 11] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/15/2016] [Revised: 04/25/2016] [Accepted: 04/25/2016] [Indexed: 01/01/2023] Open
Abstract
Coronin 1C is an established modulator of actin cytoskeleton dynamics. It has been shown to be involved in protrusion formation, cell migration and invasion. Here, we report the generation of primary fibroblasts from coronin 1C knock-out mice in order to investigate the impact of the loss of coronin 1C on cellular structural organisation. We demonstrate that the lack of coronin 1C not only affects the actin system, but also the microtubule and the vimentin intermediate filament networks. In particular, we show that the knock-out cells exhibit a reduced proliferation rate, impaired cell migration and protrusion formation as well as an aberrant subcellular localisation and function of mitochondria. Moreover, we demonstrate that coronin 1C specifically interacts with the non-α-helical amino-terminal domain ("head") of vimentin. Our data suggest that coronin 1C acts as a cytoskeletal integrator of actin filaments, microtubules and intermediate filaments.
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Affiliation(s)
- Juliane Behrens
- Center for Biochemistry, Institute of Biochemistry I, Medical Faculty, University of Cologne, 50931 Cologne, Germany
| | - Roxana Solga
- Center for Biochemistry, Institute of Biochemistry I, Medical Faculty, University of Cologne, 50931 Cologne, Germany
| | - Anja Ziemann
- Center for Biochemistry, Institute of Biochemistry I, Medical Faculty, University of Cologne, 50931 Cologne, Germany
| | - Raphael H Rastetter
- Center for Biochemistry, Institute of Biochemistry I, Medical Faculty, University of Cologne, 50931 Cologne, Germany; Center for Molecular Medicine Cologne (CMMC), University of Cologne, 50931 Cologne, Germany; Cologne Excellence Cluster on Cellular Stress Responses in Aging-Associated Diseases (CECAD), University of Cologne, 50931Cologne, Germany
| | - Carolin Berwanger
- Center for Biochemistry, Institute of Biochemistry I, Medical Faculty, University of Cologne, 50931 Cologne, Germany
| | - Harald Herrmann
- Institute of Neuropathology, University Hospital Erlangen, 91054 Erlangen, Germany
| | - Angelika A Noegel
- Center for Biochemistry, Institute of Biochemistry I, Medical Faculty, University of Cologne, 50931 Cologne, Germany; Center for Molecular Medicine Cologne (CMMC), University of Cologne, 50931 Cologne, Germany; Cologne Excellence Cluster on Cellular Stress Responses in Aging-Associated Diseases (CECAD), University of Cologne, 50931Cologne, Germany
| | - Christoph S Clemen
- Center for Biochemistry, Institute of Biochemistry I, Medical Faculty, University of Cologne, 50931 Cologne, Germany.
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19
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Caster DJ, Korte EA, Merchant ML, Klein JB, Wilkey DW, Rovin BH, Birmingham DJ, Harley JB, Cobb BL, Namjou B, McLeish KR, Powell DW. Autoantibodies targeting glomerular annexin A2 identify patients with proliferative lupus nephritis. Proteomics Clin Appl 2015; 9:1012-20. [PMID: 25824007 DOI: 10.1002/prca.201400175] [Citation(s) in RCA: 32] [Impact Index Per Article: 3.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/03/2014] [Revised: 02/10/2015] [Accepted: 03/26/2015] [Indexed: 11/09/2022]
Abstract
PURPOSE Patients with systemic lupus erythematosus (SLE) frequently develop lupus nephritis (LN), a complication frequently leading to end stage kidney disease. Immune complex deposition in the glomerulus is central to the development of LN. Using a targeted proteomic approach, we tested the hypothesis that autoantibodies targeting glomerular antigens contribute to the development of LN. EXPERIMENTAL DESIGN Human podocyte and glomerular proteins were separated by SDS-PAGE and immunoblotted with sera from SLE patients with and without LN. The regions of those gels corresponding to reactive bands observed with sera from LN patients were analyzed using LC-MS/MS. RESULTS LN reactive bands were seen at approximately 50 kDa in podocyte extracts and between 36 and 50 kDa in glomerular extracts. Those bands were analyzed by LC-MS/MS and 102 overlapping proteins were identified. Bioinformatic analysis determined that 36 of those proteins were membrane associated, including a protein previously suggested to contribute to glomerulonephritis and LN, annexin A2. By ELISA, patients with proliferative LN demonstrated significantly increased antibodies against annexin A2. CONCLUSION AND CLINICAL RELEVANCE Proteomic approaches identified multiple candidate antigens for autoantibodies in patients with LN. Serum antibodies against annexin A2 were significantly elevated in subjects with proliferative LN, validating those antibodies as potential biomarkers.
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Affiliation(s)
- Dawn J Caster
- Department of Medicine, University of Louisville School of Medicine, Louisville, KY, USA.,Robley Rex Veterans Affairs Medical Center, Louisville, KY, USA
| | - Erik A Korte
- Department of Biochemistry and Molecular Biology, University of Louisville School of Medicine, Louisville, KY, USA
| | - Michael L Merchant
- Department of Medicine, University of Louisville School of Medicine, Louisville, KY, USA
| | - Jon B Klein
- Department of Medicine, University of Louisville School of Medicine, Louisville, KY, USA.,Robley Rex Veterans Affairs Medical Center, Louisville, KY, USA
| | - Daniel W Wilkey
- Department of Medicine, University of Louisville School of Medicine, Louisville, KY, USA
| | - Brad H Rovin
- Department of Medicine, the Ohio State University, Columbus, OH, USA
| | - Dan J Birmingham
- Department of Medicine, the Ohio State University, Columbus, OH, USA
| | - John B Harley
- Center for Autoimmune Genomics and Etiology, Department of Pediatrics, Cincinnati Children's Hospital Medical Center and the University of Cincinnati, Cincinnati, OH, USA.,US Department of Veterans Affairs Medical Center, Cincinnati, OH, USA
| | - Beth L Cobb
- Center for Autoimmune Genomics and Etiology, Department of Pediatrics, Cincinnati Children's Hospital Medical Center and the University of Cincinnati, Cincinnati, OH, USA
| | - Bahram Namjou
- Center for Autoimmune Genomics and Etiology, Department of Pediatrics, Cincinnati Children's Hospital Medical Center and the University of Cincinnati, Cincinnati, OH, USA
| | - Kenneth R McLeish
- Department of Medicine, University of Louisville School of Medicine, Louisville, KY, USA.,Robley Rex Veterans Affairs Medical Center, Louisville, KY, USA
| | - David W Powell
- Department of Medicine, University of Louisville School of Medicine, Louisville, KY, USA.,Department of Biochemistry and Molecular Biology, University of Louisville School of Medicine, Louisville, KY, USA
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20
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Williamson RC, Cowell CAM, Reville T, Roper JA, Rendall TCS, Bass MD. Coronin-1C Protein and Caveolin Protein Provide Constitutive and Inducible Mechanisms of Rac1 Protein Trafficking. J Biol Chem 2015; 290:15437-15449. [PMID: 25925950 PMCID: PMC4505459 DOI: 10.1074/jbc.m115.640367] [Citation(s) in RCA: 10] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/25/2015] [Indexed: 02/05/2023] Open
Abstract
Sustained directional fibroblast migration requires both polarized activation of the protrusive signal, Rac1, and redistribution of inactive Rac1 from the rear of the cell so that it can be redistributed or degraded. In this work, we determine how alternative endocytic mechanisms dictate the fate of Rac1 in response to the extracellular matrix environment. We discover that both coronin-1C and caveolin retrieve Rac1 from similar locations at the rear and sides of the cell. We find that coronin-1C-mediated extraction, which is responsible for Rac1 recycling, is a constitutive process that maintains Rac1 protein levels within the cell. In the absence of coronin-1C, the effect of caveolin-mediated endocytosis, which targets Rac1 for proteasomal degradation, becomes apparent. Unlike constitutive coronin-1C-mediated trafficking, caveolin-mediated Rac1 endocytosis is induced by engagement of the fibronectin receptor syndecan-4. Such an inducible endocytic/degradation mechanism would predict that, in the presence of fibronectin, caveolin defines regions of the cell that are resistant to Rac1 activation but, in the absence of fibronectin leaves more of the membrane susceptible to Rac1 activation and protrusion. Indeed, we demonstrate that fibronectin-stimulated activation of Rac1 is accelerated in the absence of caveolin and that, when caveolin is knocked down, polarization of active Rac1 is lost in FRET experiments and culminates in shunting migration in a fibrous fibronectin matrix. Although the concept of polarized Rac1 activity in response to chemoattractants has always been apparent, our understanding of the balance between recycling and degradation explains how polarity can be maintained when the chemotactic gradient has faded.
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Affiliation(s)
- Rosalind C Williamson
- School of Biochemistry and University of Bristol, University Walk, Bristol BS8 1TD, United Kingdom
| | - Christopher A M Cowell
- School of Biochemistry and University of Bristol, University Walk, Bristol BS8 1TD, United Kingdom
| | - Thomas Reville
- School of Biochemistry and University of Bristol, University Walk, Bristol BS8 1TD, United Kingdom
| | - James A Roper
- School of Biochemistry and University of Bristol, University Walk, Bristol BS8 1TD, United Kingdom
| | - Thomas C S Rendall
- Department of Engineering, University of Bristol, University Walk, Bristol BS8 1TD, United Kingdom
| | - Mark D Bass
- School of Biochemistry and University of Bristol, University Walk, Bristol BS8 1TD, United Kingdom; Centre for Membrane Interactions and Dynamics, Department of Biomedical Science, University of Sheffield, Western Bank, Sheffield S10 2TN, United Kingdom.
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21
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Yamaoka M, Ishizaki T, Kimura T. Interplay between Rab27a effectors in pancreatic β-cells. World J Diabetes 2015; 6:508-516. [PMID: 25897360 PMCID: PMC4398906 DOI: 10.4239/wjd.v6.i3.508] [Citation(s) in RCA: 13] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 08/25/2014] [Revised: 12/24/2014] [Accepted: 02/09/2015] [Indexed: 02/05/2023] Open
Abstract
The small GTPase Rab27a is a member of the Rab family that is involved in membrane trafficking in various kinds of cells. Rab27a has GTP- and GDP-bound forms, and their interconversion regulates intracellular signaling pathways. Typically, only a GTP-bound GTPase binds its specific effectors with the resulting downstream signals controlling specific cellular functions. We previously identified novel Rab27a-interacting proteins. Surprisingly, some of these proteins interacted with GDP-bound Rab27a. The present study reviews recent progress in our understanding of the roles of Rab27a and its effectors in the secretory process. In pancreatic β-cells, GTP-bound Rab27a regulates insulin secretion at the pre-exocytotic stages via its GTP-specific effectors such as Exophilin8/Slac2-c/MyRIP and Slp4/Granuphilin. Glucose stimulation causes insulin exocytosis. Glucose stimulation also converts Rab27a from its GTP- to its GDP-bound form. GDP-bound Rab27a interacts with GDP-specific effectors and controls endocytosis of the secretory membrane. Thus, Rab27a cycling between GTP- and GDP-bound forms synchronizes with the recycling of secretory membrane to re-use the membrane and keep the β-cell volume constant.
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Downregulation of the microRNA-1/133a cluster enhances cancer cell migration and invasion in lung-squamous cell carcinoma via regulation of Coronin1C. J Hum Genet 2014; 60:53-61. [DOI: 10.1038/jhg.2014.111] [Citation(s) in RCA: 55] [Impact Index Per Article: 5.0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/06/2014] [Revised: 11/13/2014] [Accepted: 11/13/2014] [Indexed: 11/08/2022]
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miR-206 inhibits cell migration through direct targeting of the actin-binding protein coronin 1C in triple-negative breast cancer. Mol Oncol 2014; 8:1690-702. [PMID: 25074552 DOI: 10.1016/j.molonc.2014.07.006] [Citation(s) in RCA: 63] [Impact Index Per Article: 5.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/22/2014] [Revised: 07/07/2014] [Accepted: 07/07/2014] [Indexed: 12/31/2022] Open
Abstract
Patients with triple-negative breast cancer (TNBC) have an overall poor prognosis, which is primarily due to a high metastatic capacity of these tumors. Novel therapeutic approaches to target the signaling pathways that promote metastasis are desirable, in order to improve the outcome for these patients. A loss of function of a microRNA, miR-206, is related to increased metastasis potential in breast cancers but the mechanism is not known. In this study, we show that miR-206 was decreased in TNBC clinical tumor samples and cell lines whereas one of its predicted targets, actin-binding protein CORO1C, was increased. Expression of miR-206 significantly reduced proliferation and migration while repressing CORO1C mRNA and protein levels. We demonstrate that miR-206 interacts with the 3'-untranslated region (3'-UTR) of CORO1C and regulates this gene post-transcriptionally. This post-transcriptional regulation was dependent on two miR-206-binding sites within the 3'-UTR of CORO1C and was relieved by mutations of corresponding sites. Further, silencing of CORO1C reduced tumor cell migration and affected the actin skeleton and cell morphology, similar to miR-206 expression, but did not reduce proliferation. In accordance with this, overexpression of CORO1C rescued the inhibitory effect of miR-206 on cell migration. Our findings suggest that miR-206 represses tumor cell migration through direct targeting of CORO1C in TNBC cells which modulates the actin filaments. This pathway is a novel mechanism that offers a mechanistic basis through which the metastatic potential of TNBC tumors could be targeted.
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Sun Y, Shang Y, Ren G, Zhou L, Feng B, Li K, Deng L, Liang J, Lu Y, Wang X. Coronin3 regulates gastric cancer invasion and metastasis by interacting with Arp2. Cancer Biol Ther 2014; 15:1163-73. [PMID: 24918434 DOI: 10.4161/cbt.29501] [Citation(s) in RCA: 12] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/07/2023] Open
Abstract
Coronin3 expression is increased in gastric cancer (GC) tissues and can promote GC invasion and metastasis. However, the mechanisms underlying Coronin3 function in GC remain unclear. In this study, we aimed to explore the interacting molecules essential for the tumor-promoting effects of Coronin3 in GC. Using mass spectrometric analysis, functional studies, and immunohistochemistry, we found that Arp2 interacted with Coronin3, and ectopic expression of Arp2 promoted GC cell migration and invasion, while Arp2 knockdown suppressed whole-cell motility and attenuated the Coronin3-mediated upregulation of cell migration and invasion. In addition, both proteins correlated with the metastatic status of GC patients. Furthermore, survival analyses demonstrated that both Coronin3 and Arp2 correlated with overall GC patient survival, and the combination of Coronin3 and Arp2 most accurately predicted GC patient prognosis. Combined, these data demonstrate that Coronin3 can regulate GC invasion and metastasis through Arp2, and the combination of Coronin3 and Arp2 provides a potential marker for predicting GC prognosis.
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Affiliation(s)
- Yi Sun
- State Key Laboratory of Cancer Biology & Xijing Hospital of Digestive Diseases; Xijing Hospital; Fourth Military Medical University; Xi'an, PR China; Department of Ultrasound Diagnostics; Tangdu Hospital; Fourth Military Medical University; Xi'an, PR China
| | - Yulong Shang
- State Key Laboratory of Cancer Biology & Xijing Hospital of Digestive Diseases; Xijing Hospital; Fourth Military Medical University; Xi'an, PR China
| | - Gui Ren
- State Key Laboratory of Cancer Biology & Xijing Hospital of Digestive Diseases; Xijing Hospital; Fourth Military Medical University; Xi'an, PR China
| | - Lin Zhou
- The 88th Hospital of PLA; Tai'an, PR China
| | - Bin Feng
- State Key Laboratory of Cancer Biology & Xijing Hospital of Digestive Diseases; Xijing Hospital; Fourth Military Medical University; Xi'an, PR China
| | - Kai Li
- State Key Laboratory of Cancer Biology & Xijing Hospital of Digestive Diseases; Xijing Hospital; Fourth Military Medical University; Xi'an, PR China
| | - Lin Deng
- State Key Laboratory of Cancer Biology & Xijing Hospital of Digestive Diseases; Xijing Hospital; Fourth Military Medical University; Xi'an, PR China
| | - Jie Liang
- State Key Laboratory of Cancer Biology & Xijing Hospital of Digestive Diseases; Xijing Hospital; Fourth Military Medical University; Xi'an, PR China
| | - Yuanyuan Lu
- State Key Laboratory of Cancer Biology & Xijing Hospital of Digestive Diseases; Xijing Hospital; Fourth Military Medical University; Xi'an, PR China
| | - Xin Wang
- State Key Laboratory of Cancer Biology & Xijing Hospital of Digestive Diseases; Xijing Hospital; Fourth Military Medical University; Xi'an, PR China
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Kleppe L, Karlsen O, Edvardsen RB, Norberg B, Andersson E, Taranger GL, Wargelius A. Cortisol treatment of prespawning female cod affects cytogenesis related factors in eggs and embryos. Gen Comp Endocrinol 2013; 189:84-95. [PMID: 23660444 DOI: 10.1016/j.ygcen.2013.04.028] [Citation(s) in RCA: 18] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 11/20/2012] [Revised: 04/20/2013] [Accepted: 04/22/2013] [Indexed: 10/26/2022]
Abstract
A stable supply of viable eggs and embryos is crucial for successful farming of Atlantic cod. Stress during broodstock rearing can have negative effects on offspring, but little is known about the molecular mechanisms that cause abnormal development. Maternally transferred mRNAs have been shown to be essential for normal development, and stress may therefore influence their expression and the subsequent embryonic development. We investigated if mimicked stress in cod females affects mRNA concentrations in eggs/embryos, and if this can be linked to viability of embryos. Three weeks before peak spawning, 20 fish were intraperitoneally implanted with either cortisol-containing or cortisol-free (sham) osmotic pumps. At peak spawning all individuals were stripped and eggs were fertilized and incubated until hatching. Samples were collected from unfertilized eggs and embryos for analysis of gene expression (microarray), viability, steroids and vitellogenin. Plasma concentration of cortisol (ng/ml) in treated females was significantly higher at spawning (127.1±20.9) than that of sham control (11.3±6.7). This difference was also reflected in eggs and embryos. Percent fertilization, asymmetric cell division and hatching were not affected. However, numerous genes were differentially expressed in eggs and embryos in response to elevated cortisol, especially in maternal (oocyte and blastula) stages. Among these differentially expressed genes, some were found to be linked to cytogenesis (stxbp6, fbxw2, capn12, thbs4, sytl2, coro1c, sel1l3), induction of mesodermal fate (fgfrl1) and import of the glucocorticoid receptor to the cell nucleus (ipo7). Gene ontology overrepresentation analysis on the whole set of differentially expressed genes at maternal stages (539 genes) revealed enriched activity in membrane associated regions, which largely corresponds to cytogenesis related processes. These results suggest that despite no visible phenotypic effects in early embryos, broodstock stress affects the egg/embryonic transcriptome, especially in relation to cytogenesis. Furthermore, effects related to egg/embryo phenotypes are difficult to measure at early stages of development, and instead might become apparent at later life stages.
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Affiliation(s)
- Lene Kleppe
- Institute of Marine Research, P. O. Box 1870, 5817 Bergen, Norway.
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Ziemann A, Hess S, Bhuwania R, Linder S, Kloppenburg P, Noegel AA, Clemen CS. CRN2 enhances the invasiveness of glioblastoma cells. Neuro Oncol 2013; 15:548-61. [PMID: 23410663 PMCID: PMC3635520 DOI: 10.1093/neuonc/nos388] [Citation(s) in RCA: 11] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/04/2012] [Accepted: 12/17/2012] [Indexed: 12/11/2022] Open
Abstract
BACKGROUND Movement of tumor cells involves dynamic remodeling of the actin cytoskeleton, which is regulated by actin binding proteins, such as CRN2 (synonyms: coronin 1C, coronin 3). In vitro, CRN2 participates in secretion, matrix degradation, protrusion formation, and cell migration. Furthermore, expression of CRN2 correlates with the malignant phenotype of human diffuse gliomas. CRN2's effects on actin polymerization and F-actin bundling are abolished by protein kinase 2 (CK2) dependent phosphorylation at serine 463. METHODS We generated human U373 glioblastoma cell lines with knock-down of CRN2 or over-expression of CRN2 variants and studied their behavior in vitro and ex vivo in organotypic brain slice cultures. RESULTS CRN2 over-expression and expression of the S463A phospho-resistant CRN2 variant increase proliferation, matrix degradation, and invasion but decrease adhesion and formation of invadopodia-like extensions in vitro. Knock-down of CRN2 and expression of S463D phospho-mimetic CRN2 generally have opposite effects. Analysis of invadopodia-like cell extensions shows a diffuse relocalization of F-actin in CRN2 knockdown cells, whereas expression of S463A and S463D mutant CRN2 causes enrichments of F-actin structures at the center and rime zone, respectively. Fluorescence recovery after photobleaching studies of CRN2 and F-actin in lamellipodia show that both CRN2 variants decrease the turnover of actin filaments. Glioblastoma cells over-expressing wild-type or S463A CRN2, which were transplanted onto brain slices, characteristically developed into tumors with an invasive phenotype. CONCLUSIONS Overall, our data indicate that CRN2 participates in cancer progression via modulation of the actin cytoskeleton.
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Affiliation(s)
- Anja Ziemann
- Center for Biochemistry, Institute of Biochemistry I, Medical Faculty (A.Z., A.A.N., C.S.C.), Institute of Zoology (S.H., P.K.), Center for Molecular Medicine Cologne (S.H., P.K., A.A.N.), and Cologne Excellence Cluster on Cellular Stress Responses in Aging-Associated Diseases, University of Cologne, Cologne (S.H., P.K., A.A.N.); and Institute of Medical Microbiology, Virology and Hygiene, University Medical Center Hamburg-Eppendorf, Hamburg, Germany (R.B., S.L.)
| | - Simon Hess
- Center for Biochemistry, Institute of Biochemistry I, Medical Faculty (A.Z., A.A.N., C.S.C.), Institute of Zoology (S.H., P.K.), Center for Molecular Medicine Cologne (S.H., P.K., A.A.N.), and Cologne Excellence Cluster on Cellular Stress Responses in Aging-Associated Diseases, University of Cologne, Cologne (S.H., P.K., A.A.N.); and Institute of Medical Microbiology, Virology and Hygiene, University Medical Center Hamburg-Eppendorf, Hamburg, Germany (R.B., S.L.)
| | - Ridhirama Bhuwania
- Center for Biochemistry, Institute of Biochemistry I, Medical Faculty (A.Z., A.A.N., C.S.C.), Institute of Zoology (S.H., P.K.), Center for Molecular Medicine Cologne (S.H., P.K., A.A.N.), and Cologne Excellence Cluster on Cellular Stress Responses in Aging-Associated Diseases, University of Cologne, Cologne (S.H., P.K., A.A.N.); and Institute of Medical Microbiology, Virology and Hygiene, University Medical Center Hamburg-Eppendorf, Hamburg, Germany (R.B., S.L.)
| | - Stefan Linder
- Center for Biochemistry, Institute of Biochemistry I, Medical Faculty (A.Z., A.A.N., C.S.C.), Institute of Zoology (S.H., P.K.), Center for Molecular Medicine Cologne (S.H., P.K., A.A.N.), and Cologne Excellence Cluster on Cellular Stress Responses in Aging-Associated Diseases, University of Cologne, Cologne (S.H., P.K., A.A.N.); and Institute of Medical Microbiology, Virology and Hygiene, University Medical Center Hamburg-Eppendorf, Hamburg, Germany (R.B., S.L.)
| | - Peter Kloppenburg
- Center for Biochemistry, Institute of Biochemistry I, Medical Faculty (A.Z., A.A.N., C.S.C.), Institute of Zoology (S.H., P.K.), Center for Molecular Medicine Cologne (S.H., P.K., A.A.N.), and Cologne Excellence Cluster on Cellular Stress Responses in Aging-Associated Diseases, University of Cologne, Cologne (S.H., P.K., A.A.N.); and Institute of Medical Microbiology, Virology and Hygiene, University Medical Center Hamburg-Eppendorf, Hamburg, Germany (R.B., S.L.)
| | - Angelika A. Noegel
- Center for Biochemistry, Institute of Biochemistry I, Medical Faculty (A.Z., A.A.N., C.S.C.), Institute of Zoology (S.H., P.K.), Center for Molecular Medicine Cologne (S.H., P.K., A.A.N.), and Cologne Excellence Cluster on Cellular Stress Responses in Aging-Associated Diseases, University of Cologne, Cologne (S.H., P.K., A.A.N.); and Institute of Medical Microbiology, Virology and Hygiene, University Medical Center Hamburg-Eppendorf, Hamburg, Germany (R.B., S.L.)
| | - Christoph S. Clemen
- Center for Biochemistry, Institute of Biochemistry I, Medical Faculty (A.Z., A.A.N., C.S.C.), Institute of Zoology (S.H., P.K.), Center for Molecular Medicine Cologne (S.H., P.K., A.A.N.), and Cologne Excellence Cluster on Cellular Stress Responses in Aging-Associated Diseases, University of Cologne, Cologne (S.H., P.K., A.A.N.); and Institute of Medical Microbiology, Virology and Hygiene, University Medical Center Hamburg-Eppendorf, Hamburg, Germany (R.B., S.L.)
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The Wilms' tumor suppressor Wt1 regulates Coronin 1B expression in the epicardium. Exp Cell Res 2013; 319:1365-81. [PMID: 23562652 DOI: 10.1016/j.yexcr.2013.03.027] [Citation(s) in RCA: 8] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/18/2012] [Revised: 03/06/2013] [Accepted: 03/11/2013] [Indexed: 01/17/2023]
Abstract
Coronin 1B has been shown to be critical for cell motility and various actin-dependent processes. To understand its role more extensively, the expression and transcriptional regulation of Coro1b gene during mouse development were explored. Coronin 1B is ubiquitously expressed in the whole embryo but nevertheless shows distinct expression pattern in developing heart. In addition to the localization in endocardium, Coronin 1B is specifically expressed in the endocardial cushion and epicardium where cardiac EMT processes take place as the heart develops. Promoter deletion analysis identified the positions between -1038 and -681 is important for Coro1b basal promoter activity. In addition to a correlation of Coronin 1B localization with Wt1 expression in the epicardium, we also identified putative Wt1 binding sequences within Coro1b promoter. Direct binding of Wt1 to GC-rich sequences within the Coro1b promoter is required for the regulation of Coro1b gene expression. In accordance with the motility defect found in Coronin 1B-knockdown cells, a modest decrease in expression of Coronin 1B in the remaining epicardium of Wt1(EGFPCre/EGFPCre) mutant embryos was observed. These findings seem to shed light on the role of Wt1 during cell migration and suggest that, at least in part, this involves transcriptional control of Coro1b gene expression.
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Wang ZG, Jia MK, Cao H, Bian P, Fang XD. Knockdown of Coronin-1C disrupts Rac1 activation and impairs tumorigenic potential in hepatocellular carcinoma cells. Oncol Rep 2012; 29:1066-72. [PMID: 23292607 DOI: 10.3892/or.2012.2216] [Citation(s) in RCA: 18] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/06/2012] [Accepted: 10/10/2012] [Indexed: 11/05/2022] Open
Abstract
Coronin-1C is an important F-actin binding protein which is critical for cell motility. Furthermore, the expression of this protein was found to be increased in diffuse tumors and was correlated with the degree of tumor malignancy. However, the mechanism(s) through which this protein enhances malignancy in hepatocellular carcinoma (HCC) is poorly understood. In this study, we found that Coronin-1C was overexpressed in human HCC tissues compared with the adjacent non-tumor tissues. Overexpression of Coronin-1C enhanced the cell migration in the human HCC cell line BEL-7402, whereas suppressed cell migration and proliferation were observed in Coronin-1C-knockdown BEL-7402 cells together with impaired cell polarity, disrupted cytoskeleton and decreased Rac-1 activation. Moreover, the Coronin-1C knockdown cells displayed a lower degree of malignancy by inducing smaller tumors in nude mice. Thus, we demonstrated a relationship between Coronin-1C overexpression and human HCC growth through enhancement of tumor cell proliferation and migration, which are correlated with Rac-1 activation.
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Affiliation(s)
- Zhi-Gang Wang
- The Second Affiliated Hospital of Jilin University, Changchun, Jilin 130041, PR China
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Ren G, Tian Q, An Y, Feng B, Lu Y, Liang J, Li K, Shang Y, Nie Y, Wang X, Fan D. Coronin 3 promotes gastric cancer metastasis via the up-regulation of MMP-9 and cathepsin K. Mol Cancer 2012; 11:67. [PMID: 22974233 PMCID: PMC3522055 DOI: 10.1186/1476-4598-11-67] [Citation(s) in RCA: 48] [Impact Index Per Article: 3.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/20/2012] [Accepted: 09/10/2012] [Indexed: 12/18/2022] Open
Abstract
Background Coronins are a family of highly evolutionary conserved proteins reportedly involved in the regulation of actin cytoskeletal dynamics, although only coronin 3 has been shown to be related to cancer cell migration. In glioblastoma cells, the knockdown of coronin 3 inhibits cell proliferation and invasion. Coronin 3 is also associated with the aggression and metastasis of hepatocellular carcinoma. In this paper, we analyze the migration, invasion and metastasis abilities of gastric cancer cells after up- or down-regulation of coronin 3, and explore the mechanism of coronin 3 in the process of gastric cancer metastasis. Results The expression of coronin 3 was higher in the highly metastatic sub-cell line MKN28-M, which we established in our laboratory. We also demonstrated that the expression of coronin 3 was remarkably higher in lymph lode metastases than in primary gastric cancer tissues, and over-expression of coronin 3 was correlated with the increased clinical stage and lymph lode metastasis. Recombinant lentiviral vectors encoding shRNAs were designed to down-regulate coronin 3 expression in gastric cancer cell lines. Stable knockdown of coronin 3 by this lentiviral vector could efficiently inhibit the migration and invasion of MKN45 gastric cancer cells. In contrast, up-regulation of coronin 3 significantly enhanced migration and invasion of MKN28-NM cells. In addition, knockdown of coronin 3 significantly reduced liver metastasis in mice after tail vein injection of gastric cancer cells. The Human Tumor Metastasis PCR Array was used to screen the metastasis-associated genes identified by the down-regulation of coronin 3, and the results suggested that, following the knockdown of coronin 3, the tumor cell migration and invasion were inhibited by the reduced expression of MMP-9 and cathepsin K. Conclusion Coronin 3 is highly expressed in gastric cancer metastases and can promote the metastatic behaviors of gastric cancer cells, including their migration and invasion.
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Affiliation(s)
- Gui Ren
- State Key Laboratory of Cancer Biology and Xijing Hospital of Digestive Diseases, Xijing Hospital, Fourth Military Medical University, Xi'an, 710032, China
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Abstract
Dynamic rearrangement of actin filament networks is critical for cell motility, phagocytosis and endocytosis. Coronins facilitate these processes, in part, by their ability to bind F-actin (filamentous actin). We previously identified a conserved surface-exposed arginine (Arg30) in the β-propeller of Coronin 1B required for F-actin binding in vitro and in vivo. However, whether this finding translates to other coronins has not been well defined. Using quantitative actin-binding assays, we show that mutating the equivalent residue abolishes F-actin binding in Coronin 1A, but not Coronin 1C. By mutagenesis and biochemical competition, we have identified a second actin-binding site in the unique region of Coronin 1C. Interestingly, leading-edge localization of Coronin 1C in fibroblasts requires the conserved site in the β-propeller, but not the site in the unique region. Furthermore, in contrast with Coronin 1A and Coronin 1B, Coronin 1C displays highly co-operative binding to actin filaments. In the present study, we highlight a novel mode of coronin regulation, which has implications for how coronins orchestrate cytoskeletal dynamics.
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Xavier CP, Rastetter RH, Blömacher M, Stumpf M, Himmel M, Morgan RO, Fernandez MP, Wang C, Osman A, Miyata Y, Gjerset RA, Eichinger L, Hofmann A, Linder S, Noegel AA, Clemen CS. Phosphorylation of CRN2 by CK2 regulates F-actin and Arp2/3 interaction and inhibits cell migration. Sci Rep 2012; 2:241. [PMID: 22355754 PMCID: PMC3268813 DOI: 10.1038/srep00241] [Citation(s) in RCA: 31] [Impact Index Per Article: 2.4] [Reference Citation Analysis] [Abstract] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/20/2011] [Accepted: 12/20/2011] [Indexed: 01/27/2023] Open
Abstract
CRN2 (synonyms: coronin 1C, coronin 3) functions in the re-organization of the actin network and is implicated in cellular processes like protrusion formation, secretion, migration and invasion. We demonstrate that CRN2 is a binding partner and substrate of protein kinase CK2, which phosphorylates CRN2 at S463 in its C-terminal coiled coil domain. Phosphomimetic S463D CRN2 loses the wild-type CRN2 ability to inhibit actin polymerization, to bundle F-actin, and to bind to the Arp2/3 complex. As a consequence, S463D mutant CRN2 changes the morphology of the F-actin network in the front of lamellipodia. Our data imply that CK2-dependent phosphorylation of CRN2 is involved in the modulation of the local morphology of complex actin structures and thereby inhibits cell migration.
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Affiliation(s)
- Charles-Peter Xavier
- Center for Biochemistry, Institute of Biochemistry I, Medical Faculty, University of Cologne, 50931, Cologne, Germany
- Both authors contributed equally to this work
- Present address: Laboratory of Cellular and Molecular Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892-4256, USA
| | - Raphael H. Rastetter
- Center for Biochemistry, Institute of Biochemistry I, Medical Faculty, University of Cologne, 50931, Cologne, Germany
- Both authors contributed equally to this work
| | - Margit Blömacher
- Center for Biochemistry, Institute of Biochemistry I, Medical Faculty, University of Cologne, 50931, Cologne, Germany
| | - Maria Stumpf
- Center for Biochemistry, Institute of Biochemistry I, Medical Faculty, University of Cologne, 50931, Cologne, Germany
| | - Mirko Himmel
- Institute of Medical Microbiology, Virology and Hygiene, University Medical Center Hamburg-Eppendorf, 20246, Hamburg, Germany
| | - Reginald O. Morgan
- Department of Biochemistry and Molecular Biology, University of Oviedo and University Institute of Biotechnology of Asturias, Oviedo, 33006, Spain
| | - Maria-Pilar Fernandez
- Department of Biochemistry and Molecular Biology, University of Oviedo and University Institute of Biotechnology of Asturias, Oviedo, 33006, Spain
| | - Conan Wang
- Structural Chemistry, Eskitis Institute for Cell and Molecular Therapies, Griffith University, Brisbane, Qld 4111, Australia
| | - Asiah Osman
- Structural Chemistry, Eskitis Institute for Cell and Molecular Therapies, Griffith University, Brisbane, Qld 4111, Australia
| | - Yoshihiko Miyata
- Department of Cell and Developmental Biology, Graduate School of Biostudies, Kyoto University, Kyoto, 606-8502, Japan
| | - Ruth A. Gjerset
- Torrey Pines Institute for Molecular Studies, San Diego, California, 92121, USA
| | - Ludwig Eichinger
- Center for Biochemistry, Institute of Biochemistry I, Medical Faculty, University of Cologne, 50931, Cologne, Germany
| | - Andreas Hofmann
- Structural Chemistry, Eskitis Institute for Cell and Molecular Therapies, Griffith University, Brisbane, Qld 4111, Australia
| | - Stefan Linder
- Institute of Medical Microbiology, Virology and Hygiene, University Medical Center Hamburg-Eppendorf, 20246, Hamburg, Germany
| | - Angelika A. Noegel
- Center for Biochemistry, Institute of Biochemistry I, Medical Faculty, University of Cologne, 50931, Cologne, Germany
- Center for Molecular Medicine Cologne (CMMC), University of Cologne, 50931, Cologne, Germany
- Cologne Excellence Cluster on Cellular Stress Responses in Aging-Associated Diseases (CECAD), University of Cologne, 50931, Cologne, Germany
| | - Christoph S. Clemen
- Center for Biochemistry, Institute of Biochemistry I, Medical Faculty, University of Cologne, 50931, Cologne, Germany
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Sampson HW, Chaput CD, Brannen J, Probe RA, Guleria RS, Pan J, Baker KM, VanBuren V. Alcohol induced epigenetic perturbations during the inflammatory stage of fracture healing. Exp Biol Med (Maywood) 2011; 236:1389-401. [PMID: 22087020 DOI: 10.1258/ebm.2011.011207] [Citation(s) in RCA: 19] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/04/2023] Open
Abstract
It is well recognized by orthopedic surgeons that fractures of alcoholics are more difficult to heal successfully and have a higher incidence of non-union, but the mechanism of alcohol's effect on fracture healing is unknown. In order to give direction for the study of the effects of alcohol on fracture healing, we propose to identify gene expression and microRNA changes during the early stages of fracture healing that might be attributable to alcohol consumption. As the inflammatory stage appears to be the most critical for successful fracture healing, this paper focuses on the events at day three following fracture or the stage of inflammation. Sprague-Dawley rats were placed on an ethanol-containing or pair-fed Lieber and DeCarli diet for four weeks prior to surgical fracture. Following insertion of a medullary pin, a closed mid-diaphyseal fracture was induced using a Bonnarens and Einhorn fracture device. At three days' post-fracture, the region of the fracture calluses was harvested from the right hind-limb. RNA was extracted and microarray analysis was conducted against the entire rat genome. There were 35 genes that demonstrated significant increased expression due to alcohol consumption and 20 that decreased due to alcohol. In addition, the expression of 20 microRNAs was increased and six decreased. In summary, while it is recognized that mRNA levels may or may not represent protein levels successfully produced by the cell, these studies reveal changes in gene expression that support the hypothesis that alcohol consumption affects events involved with inflammation. MicroRNAs are known to modulate mRNA and these findings were consistent with much of what was seen with mRNA microarray analysis, especially the involvement of smad4 which was demonstrated by mRNA microarray, microRNA and polymerase chain reaction.
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Affiliation(s)
- H Wayne Sampson
- Department of Systems Biology and Translational Medicine, Texas A&M Health Science Center, College of Medicine, USA.
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Landkocz Y, Poupin P, Atienzar F, Vasseur P. Transcriptomic effects of di-(2-ethylhexyl)-phthalate in Syrian hamster embryo cells: an important role of early cytoskeleton disturbances in carcinogenesis? BMC Genomics 2011; 12:524. [PMID: 22026506 PMCID: PMC3218109 DOI: 10.1186/1471-2164-12-524] [Citation(s) in RCA: 13] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/11/2011] [Accepted: 10/25/2011] [Indexed: 01/18/2023] Open
Abstract
Background Di-(2-ethylhexyl)-phthalate (DEHP) is a commonly used plasticizer in polyvinylchloride (PVC) formulations and a potentially non-genotoxic carcinogen. The aim of this study was to identify genes whose level of expression is altered by DEHP by using a global wide-genome approach in Syrian hamster embryo (SHE) cells, a model similar to human cells regarding their responses to this type of carcinogen. With mRNA Differential Display (DD), we analysed the transcriptional regulation of SHE cells exposed to 0, 12.5, 25 and 50 μM of DEHP for 24 hrs, conditions which induced neoplastic transformation of these cells. A real-time quantitative polymerase chain reaction (qPCR) was used to confirm differential expression of genes identified by DD. Results Gene expression profiling showed 178 differentially-expressed fragments corresponding to 122 genes after tblastx comparisons, 79 up-regulated and 43 down-regulated. The genes of interest were involved in many biological pathways, including signal transduction, regulation of the cytoskeleton, xenobiotic metabolism, apoptosis, lipidogenesis, protein conformation, transport and cell cycle. We then focused particularly on genes involved in the regulation of the cytoskeleton, one of the processes occurring during carcinogenesis and in the early steps of neoplastic transformation. Twenty one cytoskeleton-related genes were studied by qPCR. The down-regulated genes were involved in focal adhesion or cell junction. The up-regulated genes were involved in the regulation of the actin cytoskeleton and this would suggest a role of cellular plasticity in the mechanism of chemical carcinogenesis. The gene expression changes identified in the present study were PPAR-independent. Conclusion This study identified a set of genes whose expression is altered by DEHP exposure in mammalian embryo cells. This is the first study that elucidates the genomic changes of DEHP involved in the organization of the cytoskeleton. The latter genes may be candidates as biomarkers predictive of early events in the multistep carcinogenic process.
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Affiliation(s)
- Yann Landkocz
- CNRS UMR7146, Laboratoire I.E.B.E., Rue General Delestraint, 57070 Metz, France.
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Kimura T, Niki I. Rab27a in pancreatic beta-cells, a busy protein in membrane trafficking. PROGRESS IN BIOPHYSICS AND MOLECULAR BIOLOGY 2011; 107:219-23. [PMID: 21762718 DOI: 10.1016/j.pbiomolbio.2011.06.016] [Citation(s) in RCA: 15] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Subscribe] [Scholar Register] [Received: 01/25/2011] [Revised: 06/15/2011] [Accepted: 06/20/2011] [Indexed: 12/14/2022]
Abstract
The small GTPases have the 'active' GTP-bound and 'inactive' GDP-bound states, and thereby act as a molecular switch in cells. Rab27a is a member of this family and exists in T-lymphocytes, melanocytes and pancreatic beta-cells. Rab27a regulates secretion of cytolytic granules from cytotoxic T-lymphocytes and intracellular transport of melanosomes in melanocytes. In pancreatic beta-cells, Rab27a controls pre-exocytotic stages of insulin secretion. A few GTP-dependent Rab27a effectors are known to mediate these cellular functions. We recently found that Rab27a also possesses the GDP-dependent effector coronin 3. Coronin 3 regulates endocytosis in pancreatic beta-cells through its interaction with GDP-Rab27a. These results imply that GTP- and GDP-Rab27a actively regulate distinct stages in the insulin secretory pathway. In this review, we provide an overview of the roles of both GTP- and GDP-Rab27a in pancreatic beta-cells.
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Affiliation(s)
- Toshihide Kimura
- Department of Pharmacology, Oita University Faculty of Medicine, 1-1 Idaigaoka, Hasama, Yufu, Oita 8795593, Japan
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Chan KT, Creed SJ, Bear JE. Unraveling the enigma: progress towards understanding the coronin family of actin regulators. Trends Cell Biol 2011; 21:481-8. [PMID: 21632254 DOI: 10.1016/j.tcb.2011.04.004] [Citation(s) in RCA: 136] [Impact Index Per Article: 9.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/18/2011] [Revised: 04/15/2011] [Accepted: 04/18/2011] [Indexed: 11/25/2022]
Abstract
Coronins are a conserved family of actin cytoskeleton regulators that promote cell motility and modulate other actin-dependent processes. Although these proteins have been known for 20 years, substantial progress has been made in the past 5 years towards their understanding. In this review, we examine this progress, place it into the context of what was already known, and pose several questions that remain to be addressed. In particular, we cover the emerging consensus about the role of Type I coronins in coordinating the function of Arp2/3 complex and ADF/cofilin proteins. This coordination plays an important role in leading-edge actin dynamics and overall cell motility. Finally, we discuss the roles played by the more exotic coronins of the Type II and III classes in cellular processes away from the leading edge.
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Affiliation(s)
- Keefe T Chan
- Lineberger Comprehensive Cancer Center and Department of Cell & Developmental Biology, Howard Hughes Medical Institute. University of North Carolina at Chapel Hill, USA
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36
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Donowitz M, Singh S, Singh P, Chakraborty M, Chen Y, Murtazina R, Gucek M, Cole RN, Zachos NC, Salahuddin FF, Kovbasnjuk O, Broere N, Smalley-Freed WG, Reynolds AB, Hubbard AL, Seidler U, Weinman E, de Jonge HR, Hogema BM, Li X. Alterations in the proteome of the NHERF2 knockout mouse jejunal brush border membrane vesicles. Physiol Genomics 2011; 43:674-84. [PMID: 21427361 DOI: 10.1152/physiolgenomics.00258.2010] [Citation(s) in RCA: 9] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/22/2022] Open
Abstract
To identify additional potential functions for the multi-PDZ domain containing protein Na+/H+ exchanger regulatory factor 2 (NHERF2), which is present in the apical domain of intestinal epithelial cells, proteomic studies of mouse jejunal villus epithelial cell brush border membrane vesicles compared wild-type to homozygous NHERF2 knockout FVB mice by a two-dimensional liquid chromatography-tandem mass spectrometry (LC-MS/MS)-iTRAQ approach. Jejunal architecture appeared normal in NHERF2 null in terms of villus length and crypt depth, Paneth cell number, and microvillus structure by electron microscopy. There was also no change in proliferative activity based on BrdU labeling. Four brush border membrane vesicles (BBMV) preparations from wild-type mouse jejunum were compared with four preparations from NHERF2 knockout mice. LC-MS/MS identified 450 proteins in both matched wild-type and NHERF2 null BBMV; 13 proteins were changed in two or more separate BBMV preparations (9 increased and 4 decreased in NHERF2 null mice), while an additional 92 proteins were changed in a single BBMV preparation (68 increased and 24 decreased in NHERF2 null mice). These proteins were categorized as 1) transport proteins (one increased and two decreased in NHERF2 null); 2) signaling molecules (2 increased in NHERF2 null); 3) cytoskeleton/junctional proteins (4 upregulated and 1 downregulated in NHERF2 null); and 4) metabolic proteins/intrinsic BB proteins) (2 upregulated and 1 downregulated in NHERF2 null). Immunoblotting of BBMV was used to validate or extend the findings, demonstrating increase in BBMV of NHERF2 null of MCT1, coronin 3, and ezrin. The proteome of the NHERF2 null mouse small intestinal BB demonstrates up- and downregulation of multiple transport proteins, signaling molecules, cytoskeletal proteins, tight junctional and adherens junction proteins, and proteins involved in metabolism, suggesting involvement of NHERF2 in multiple apical regulatory processes and interactions with luminal contents.
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Affiliation(s)
- M Donowitz
- Department of Medicine, Johns Hopkins University School of Medicine, Baltimore, MD 21205-2195, USA.
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Rehberg K, Bergado-Acosta JR, Koch JC, Stork O. Disruption of fear memory consolidation and reconsolidation by actin filament arrest in the basolateral amygdala. Neurobiol Learn Mem 2010; 94:117-26. [PMID: 20416387 DOI: 10.1016/j.nlm.2010.04.007] [Citation(s) in RCA: 30] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/17/2009] [Revised: 03/19/2010] [Accepted: 04/16/2010] [Indexed: 10/19/2022]
Abstract
The dynamic re-arrangement of actin filaments is an essential process in the plasticity of synaptic connections during memory formation. In this study, we determined in mice effects of actin filament arrest in the basolateral complex of the amygdala (BLA) at different time points after memory acquisition and re-activation, using the fungal cytotoxin phalloidin. Our data show a selective disruption of auditory cued but not contextual fear memory, when phalloidin was injected 6h after conditioning. In contrast, no effect was observed when phalloidin was applied after 24h, ruling out an interference with the retrieval or expression of conditioned fear. A comparable result was obtained after memory re-activation, hence suggesting similar actin-dependent mechanisms to be active during consolidation and reconsolidation of auditory fear memory. Biochemical analysis showed that phalloidin-mediated filament arrest leads to a transient increase of highly cross-linked actin filaments in the BLA, evident 2h after injection. Together, these observations indicate that dynamic re-arrangements of actin filaments in the BLA during a late phase of fear memory consolidation and reconsolidation are critical for fear memory storage.
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Affiliation(s)
- Kati Rehberg
- Department of Genetics & Molecular Neurobiology, Institute of Biology, Otto-von-Guericke University Magdeburg, 39120 Magdeburg, Germany
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Kimura T, Taniguchi S, Niki I. Actin assembly controlled by GDP-Rab27a is essential for endocytosis of the insulin secretory membrane. Arch Biochem Biophys 2010; 496:33-7. [PMID: 20138020 DOI: 10.1016/j.abb.2010.01.017] [Citation(s) in RCA: 26] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/18/2009] [Revised: 01/27/2010] [Accepted: 01/29/2010] [Indexed: 12/16/2022]
Abstract
We have recently reported that GDP-bound Rab27a regulates endocytosis of the insulin secretory membrane via its binding to coronin 3, an actin-binding protein. The aim of this study was to examine the participation of actin assembly in the Rab27a-dependent regulation of endocytosis using a pancreatic beta cell line, MIN6. Coronin 3 promoted F-actin bundling only in the presence of GDP-Rab27a. This effect was independent of coronin-3-binding to the actin-related proteins 2 and 3 (Arp2/3). Uptake of anti-phogrin-lumen antibody into MIN6 was inhibited by anti-coronin-3-C antibody which recognizes the actin-binding site. This inhibition was also observed with coronin-3-R28D, which lacks in actin binding. These results suggest that coronin 3 is a genuine GDP-Rab27a effector, and that controls endocytosis of the secretory membrane via modulating actin assembly in pancreatic beta-cells.
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Affiliation(s)
- Toshihide Kimura
- Department of Pharmacology, Oita University Faculty of Medicine at Oita, Japan
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Samarin SN, Koch S, Ivanov AI, Parkos CA, Nusrat A. Coronin 1C negatively regulates cell-matrix adhesion and motility of intestinal epithelial cells. Biochem Biophys Res Commun 2009; 391:394-400. [PMID: 19913511 DOI: 10.1016/j.bbrc.2009.11.069] [Citation(s) in RCA: 16] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/31/2009] [Accepted: 11/07/2009] [Indexed: 11/24/2022]
Abstract
Coronins, WD-repeat actin-binding proteins, are known to regulate cell motility by coordinating actin filament turnover in lamellipodia of migrating cell. Here we report a novel mechanism of Coronin 1C-mediated cell motility that involves regulation of cell-matrix adhesion. RNAi silencing of Coronin 1C in intestinal epithelial cells enhanced cell migration and modulated lamellipodia dynamics by increasing the persistence of lamellipodial protrusion. Coronin 1C-depleted cells showed increased cell-matrix adhesions and enhanced cell spreading compared to control cells, while over-expression of Coronin 1C antagonized cell adhesion and spreading. Enhanced cell-matrix adhesion of coronin-deficient cells correlated with hyperphosphorylation of focal adhesion kinase (FAK) and paxillin, and an increase in number of focal adhesions and their redistribution at the cell periphery. siRNA depletion of FAK in coronin-deficient cells rescued the effects of Coronin 1C depletion on motility, cell-matrix adhesion, and spreading. Thus, our findings provide the first evidence that Coronin 1C negatively regulates epithelial cell migration via FAK-mediated inhibition of cell-matrix adhesion.
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Affiliation(s)
- Stanislav N Samarin
- Epithelial Pathobiology Research Unit, Department of Pathology and Laboratory Medicine, Emory University, Atlanta, GA 30322, USA.
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Xavier CP, Rastetter RH, Stumpf M, Rosentreter A, Müller R, Reimann J, Cornfine S, Linder S, van Vliet V, Hofmann A, Morgan RO, Fernandez MP, Schröder R, Noegel AA, Clemen CS. Structural and Functional Diversity of Novel Coronin 1C (CRN2) Isoforms in Muscle. J Mol Biol 2009; 393:287-99. [DOI: 10.1016/j.jmb.2009.07.079] [Citation(s) in RCA: 8] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/29/2008] [Revised: 07/27/2009] [Accepted: 07/28/2009] [Indexed: 01/07/2023]
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Marshall TW, Aloor HL, Bear JE. Coronin 2A regulates a subset of focal-adhesion-turnover events through the cofilin pathway. J Cell Sci 2009; 122:3061-9. [PMID: 19654210 DOI: 10.1242/jcs.051482] [Citation(s) in RCA: 34] [Impact Index Per Article: 2.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/01/2023] Open
Abstract
Coronins are conserved F-actin-binding proteins that are important for motility and actin dynamics. Unlike type I coronins, coronin 2A localizes to stress fibers and some focal adhesions, and is excluded from the leading edge. Depletion of coronin 2A in MTLn3 cells decreases cell motility and turnover of focal adhesions. Surprisingly, none of the pathways known to regulate focal-adhesion turnover are affected by depletion of coronin 2A. Depletion of coronin 2A does, however, increase phospho-cofilin, suggesting that misregulation of cofilin might affect adhesion dynamics. Slingshot-1L, a cofilin-activating phosphatase, localizes to focal adhesions and interacts with coronin 2A. Depletion of coronin 2A reduces cofilin activity at focal adhesions, as measured by barbed-end density and actin FRAP. In both fixed cells and live cells, cofilin localizes to the proximal end of some focal adhesions. Although expression of wild-type cofilin in coronin-2A-depleted cells has no major effect on focal-adhesion dynamics, expression of an active mutant of cofilin bypasses the defects in cell motility and focal-adhesion disassembly. These results implicate both coronin 2A and cofilin as factors that can regulate a subset of focal-adhesion-turnover events.
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Affiliation(s)
- Thomas W Marshall
- Lineberger Comprehensive Cancer Center and Department of Cell and Developmental Biology, University of North Carolina-Chapel Hill, Chapel Hill, NC 27599, USA
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Kimura T, Kaneko Y, Yamada S, Ishihara H, Senda T, Iwamatsu A, Niki I. The GDP-dependent Rab27a effector coronin 3 controls endocytosis of secretory membrane in insulin-secreting cell lines. J Cell Sci 2008; 121:3092-8. [PMID: 18768935 DOI: 10.1242/jcs.030544] [Citation(s) in RCA: 59] [Impact Index Per Article: 3.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/11/2022] Open
Abstract
Rab27a is involved in the control of membrane traffic, a crucial step in the regulated secretion. Typically, the guanosine triphosphate (GTP)-bound form has been considered to be active and, therefore, searching for proteins binding to the GTP-form has been attempted to look for their effectors. Here, we have identified the actin-bundling protein coronin 3 as a novel Rab27a effector that paradoxically bound guanosine diphosphate (GDP)-Rab27a in the pancreatic beta-cell line MIN6. Coronin 3 directly bound GDP-Rab27a through its beta-propeller structure. The most important insulin secretagogue glucose promptly shifted Rab27a from the GTP- to GDP-bound form. Knockdown of coronin 3 by RNAi resulted in the inhibition of phogrin (an insulin-granule-associated protein) internalization and the uptake of FM4-64 (a marker of endocytosis). Similar results were reproduced by disruption of the coronin-3-GDP-Rab27a interaction with the dominant-negative coronin 3, and coexpression of the GDP-Rab27a mutant rescued these changes. Taken together, our results indicate that interaction of GDP-Rab27a and coronin 3 is important in stimulus-endocytosis coupling, and that GTP- and GDP-Rab27a regulates insulin membrane recycling at the distinct stages.
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Affiliation(s)
- Toshihide Kimura
- Department of Pharmacology, Oita University Faculty of Medicine, Hasama, Yufu, Oita 879-5593, Japan
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Abstract
Until recently, structural information about coronins was scarce and the earlier identification of five WD40 repeats gave rise to a structural prediction of a five-bladed beta propeller for the N-terminal domain of these proteins. More detailed analyses revealed the presence of seven WD40 repeats and the hypothesis of a seven-bladed beta propeller structure. This model has recently been validated due to structural information from crystal structures of C-terminally truncated coronin 1 (1A), as well as its C-terminal coiled coil domain. Further structural information is available only indirectly from binding and functional studies.Phosphorylation at distinct serine and tyrosine residues seems to be a common theme for various coronins. There are indications that this modification regulates the quaternary structure of coronin 3 (1C) and thus has implications for the cellular localisation and the general link between signalling and cytoskeletal remodelling. Similarly, phosphorylation-dependent sorting sequences recently discovered on coronin 7 might prove important for the molecular mechanisms of the longer coronins.A matter that will require further clarification is the localisation of protein binding sites on coronins. While earlier reports presented a rather diverse map of actin binding sites, more recent studies, including the crystal structure of the coronin 1 N-terminal domain, deliver more detailed information in this respect. Interaction sites for other target proteins, such as Arp2/3, remain to be identified. Also, while membrane binding is a known feature of coronins, further details as to the binding sites and molecular level events remain to be elucidated. The N-terminal WD40 repeat domain seems to be the membrane-interacting domain, but other domains might provide regulatory effects, most likely by posttranslational modification, in a fashion that is specific for each coronin.In this chapter, we provide a structural overview of coronins 1 (1A), 2 (1B), 3 (1C) and 7 and also present results of our recent efforts to obtain structural models of coronins 3 and 7. Possible implications of these models on the function of these proteins are discussed.
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Abstract
The coronin gene family comprises seven vertebrate paralogs and at least five unclassified subfamilies in nonvertebrate metazoa, fungi and protozoa, but no representatives in plants or distant protists. All known members exhibit elevated structural conservation in two unique domains of unknown function (DUF1899 and DUF1900) interspaced by three canonical WD40 domains (plus additional pseudo domains) that form part of a 7-bladed beta-propeller scaffold, plus a C-terminal variable "coiled coil domain" responsible for oligomerization. Phylogenetic analysis of the N-terminal conserved region in known members (i.e.420 aa in 250 taxa) established the origin of the founding monomeric unit and a dimeric paralog in unicellular eukaryotes. The monomeric ancestor duplicated to two distinct lineages in basal metazoa and later propagated during the whole genome duplications in primitive chordates 450-550 million years ago to form six vertebrate-specific genes. The delineation of 12 subfamily clades in distinct phyla provided a rational basis for proposing a simplified, universal nomenclature for the coronin family in accordance with evolutionary history, structural relationships and functional divergence.Comparative genomic analysis of coronin subfamily locus maps and gene organization provided corroboratory evidence for their chromosomal dispersal and structural relatedness. Statistical analysis of evolutionary sequence conservation by profile hidden Markov models (pHMM) and the prediction of Specificity Determining Positions (SDPpred) helped to characterize coronin domains by highlighting structurally conserved sites relevant to coronin function and subfamily divergence. The incorporation of such evolutionary information into 3D models facilitated the distinction between candidate sites with a structural role versus those implicated in dynamic, actin-related cytoskeletal interactions. A highly conserved "KGD" motif identified in the coronin DUF1900 domain has been observed in other actin-binding proteins such as annexins and is a potential ligand for integrins and C2 domains known to be associated with structural and signalling roles in the membrane cytoskeleton. Molecular evolution studies provide a comprehensive overview of the structural history of the coronin gene family and a systematic methodology to gain deeper insight into the function(s) of individual members.
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Abstract
What I'd like to do in this chapter is to share with you my recollections from the earliest days of coronin research and then to provide an overview of the still-developing story of this fascinating family of proteins.
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Coronin-1A links cytoskeleton dynamics to TCR alpha beta-induced cell signaling. PLoS One 2008; 3:e3467. [PMID: 18941544 PMCID: PMC2568942 DOI: 10.1371/journal.pone.0003467] [Citation(s) in RCA: 58] [Impact Index Per Article: 3.4] [Reference Citation Analysis] [Abstract] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/17/2008] [Accepted: 09/20/2008] [Indexed: 11/19/2022] Open
Abstract
Actin polymerization plays a critical role in activated T lymphocytes both in regulating T cell receptor (TCR)-induced immunological synapse (IS) formation and signaling. Using gene targeting, we demonstrate that the hematopoietic specific, actin- and Arp2/3 complex-binding protein coronin-1A contributes to both processes. Coronin-1A-deficient mice specifically showed alterations in terminal development and the survival of αβT cells, together with defects in cell activation and cytokine production following TCR triggering. The mutant T cells further displayed excessive accumulation yet reduced dynamics of F-actin and the WASP-Arp2/3 machinery at the IS, correlating with extended cell-cell contact. Cell signaling was also affected with the basal activation of the stress kinases sAPK/JNK1/2; and deficits in TCR-induced Ca2+ influx and phosphorylation and degradation of the inhibitor of NF-κB (IκB). Coronin-1A therefore links cytoskeleton plasticity with the functioning of discrete TCR signaling components. This function may be required to adjust TCR responses to selecting ligands accounting in part for the homeostasis defect that impacts αβT cells in coronin-1A deficient mice, with the exclusion of other lympho/hematopoietic lineages.
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Abstract
Coronin is a conserved actin binding protein that promotes cellular processes that rely on rapid remodeling of the actin cytoskeleton, including endocytosis and cell motility. However, the exact mechanism by which coronin contributes to actin dynamics has remained elusive for many years. Here, we integrate observations from many groups and propose a unified model to explain how coronin controls actin dynamics through coordinated effects on Arp2/3 complex and cofilin. At the front end of actin networks, coronin protects new (ATP-rich) filaments from premature disassembly by cofilin and recruits Arp2/3 complex to filament sides, leading to nucleation, branching and network expansion. At the rear of networks, coronin has strikingly different activities, synergizing with cofilin to dismantle old (ADP-rich) filaments. Thus, coronin spatially targets Arp2/3 complex and cofilin to opposite ends of actin networks. The net effect of coronin's activities is acceleration of polarized actin subunit flux through filamentous arrays. This increases actin network plasticity and replenishes the actin monomer pool required for new filament growth.
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Affiliation(s)
- Meghal Gandhi
- Department of Biology, Rosenstiel Basic Medical Science Research Center, Brandeis University, 415 South Street, Waltham, MA 02454, USA
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Roadcap DW, Clemen CS, Bear JE. The role of mammalian coronins in development and disease. Subcell Biochem 2008; 48:124-35. [PMID: 18925377 DOI: 10.1007/978-0-387-09595-0_12] [Citation(s) in RCA: 29] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/21/2022]
Abstract
Coronins have maintained a high degree of conservation over the roughly 800 million years of eukaryotic evolution.1,2 From its origins as a single gene in simpler eukaryotes, the mammalian Coronin gene family has expanded to include at least six members (see Chapter 4). Increasing evidence indicates that Coronins play critical roles as regulators of actin dependent processes such as cell motility and vesicle trafficking3,4 (see Chapters 6-9). Considering the importance of these processes, it is not surprising that recent findings have implicated the involvement of Coronins in multiple diseases. This review primarily focuses on Coronin 1C (HGNC symbol: CORO1C, also known as Coronin 3) which is a transcriptionally dynamic gene that is up-regulated in multiple types of clinically aggressive cancer. In addition to reviewing the molecular signals and events that lead to Coronin 1C transcription, we summarize the results of several studies describing the possible functional roles of Coronin 1C in development as well as disease progression. Here, the main focus is on brain development and on the progression of melanoma and glioma. Finally, we will also review the role of other mammalian Coronin genes in clinically relevant processes such as neural regeneration and pathogenic bacterial infections (see Chapter 10).
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Affiliation(s)
- David W Roadcap
- Lineberger Comprehensive Cancer Center and Department of Cell and Developmental Biology, UNC-Chapel Hill, Chapel Hill, NC 27599, USA
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Thal D, Xavier CP, Rosentreter A, Linder S, Friedrichs B, Waha A, Pietsch T, Stumpf M, Noegel A, Clemen C. Expression of coronin-3 (coronin-1C) in diffuse gliomas is related to malignancy. J Pathol 2008; 214:415-24. [PMID: 18189330 DOI: 10.1002/path.2308] [Citation(s) in RCA: 46] [Impact Index Per Article: 2.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/07/2022]
Abstract
Coronin-3 (coronin-1C), a homotrimeric F-actin binding protein, has been shown to be important for cell migration and brain morphogenesis. Here, we present for the first time a detailed analysis of the expression pattern of coronin-3 in human brain tumours and demonstrate that coronin-3 expression correlates with malignant phenotype in diffuse gliomas. In general, the expression of coronin-3 varies in different brain tumour entities. However, in diffuse gliomas, the number of coronin-3 expressing tumour cells correlates with the degree of malignancy. High-grade gliomas, such as anaplastic astrocytomas, anaplastic oligodendrogliomas, anaplastic oligoastrocytomas and glioblastomas, show high numbers of tumour cells positive for coronin-3, while diffuse low-grade gliomas, such as diffuse astrocytomas, oligodendrogliomas and oligoastrocytomas, exhibit low numbers of coronin-3-positive tumour cells. In order to explore and verify a contribution of coronin-3 to the malignant phenotype of diffuse gliomas, we employed an efficient shRNA-mediated coronin-3 knockdown in U373 and A172 human glioblastoma cells. Coronin-3 knockdown glioblastoma cells exhibited reduced levels of cell proliferation, cell motility and invasion into extracellular matrix compared to control cells. Together, our findings demonstrate evidence for a contribution of coronin-3 expression in the malignant progression of diffuse gliomas.
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Affiliation(s)
- Dr Thal
- Department of Neuropathology, University of Bonn Medical Centre, Sigmund-Freud-Strasse 25, 53105 Bonn, Germany
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