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Owczarek C, Ortiz-Zapater E, Kim J, Papaevangelou E, Santis G, Parsons M. CAR Co-Operates With Integrins to Promote Lung Cancer Cell Adhesion and Invasion. Front Oncol 2022; 12:829313. [PMID: 35252000 PMCID: PMC8889575 DOI: 10.3389/fonc.2022.829313] [Citation(s) in RCA: 6] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/05/2021] [Accepted: 01/19/2022] [Indexed: 12/14/2022] Open
Abstract
The coxsackie and adenovirus receptor (CAR) is a member of the junctional adhesion molecule (JAM) family of adhesion receptors and is localised to epithelial cell tight and adherens junctions. CAR has been shown to be highly expressed in lung cancer where it is proposed to promote tumor growth and regulate epithelial mesenchymal transition (EMT), however the potential role of CAR in lung cancer metastasis remains poorly understood. To better understand the role of this receptor in tumor progression, we manipulated CAR expression in both epithelial-like and mesenchymal-like lung cancer cells. In both cases, CAR overexpression promoted tumor growth in vivo in immunocompetent mice and increased cell adhesion in the lung after intravenous injection without altering the EMT properties of each cell line. Overexpression of WTCAR resulted in increased invasion in 3D models and enhanced β1 integrin activity in both cell lines, and this was dependent on phosphorylation of the CAR cytoplasmic tail. Furthermore, phosphorylation of CAR was enhanced by substrate stiffness in vitro, and CAR expression increased at the boundary of solid tumors in vivo. Moreover, CAR formed a complex with the focal adhesion proteins Src, Focal Adhesion Kinase (FAK) and paxillin and promoted activation of the Guanine Triphosphate (GTP)-ase Ras-related Protein 1 (Rap1), which in turn mediated enhanced integrin activation. Taken together, our data demonstrate that CAR contributes to lung cancer metastasis via promotion of cell-matrix adhesion, providing new insight into co-operation between cell-cell and cell-matrix proteins that regulate different steps of tumorigenesis.
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Affiliation(s)
- Claudia Owczarek
- Randall Centre for Cell and Molecular Biophysics, King’s College London, London, United Kingdom
| | - Elena Ortiz-Zapater
- Randall Centre for Cell and Molecular Biophysics, King’s College London, London, United Kingdom
- School of Biomedical Engineering and Imaging Sciences, King’s College London, St Thomas Hospital, London, United Kingdom
| | - Jana Kim
- School of Biomedical Engineering and Imaging Sciences, King’s College London, St Thomas Hospital, London, United Kingdom
| | - Efthymia Papaevangelou
- Peter Gorer Department of Immunobiology, School of Immunology and Microbial Science, King’s College London, London, United Kingdom
| | - George Santis
- Peter Gorer Department of Immunobiology, School of Immunology and Microbial Science, King’s College London, London, United Kingdom
| | - Maddy Parsons
- Randall Centre for Cell and Molecular Biophysics, King’s College London, London, United Kingdom
- *Correspondence: Maddy Parsons,
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White SH, Sturgeon RM, Gu Y, Nensi A, Magoski NS. Tyrosine Phosphorylation Determines Afterdischarge Initiation by Regulating an Ionotropic Cholinergic Receptor. Neuroscience 2018; 372:273-288. [PMID: 29306054 DOI: 10.1016/j.neuroscience.2017.12.049] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/21/2017] [Revised: 11/30/2017] [Accepted: 12/26/2017] [Indexed: 12/12/2022]
Abstract
Changes to neuronal activity often involve a rapid and precise transition from low to high excitability. In the marine snail, Aplysia, the bag cell neurons control reproduction by undergoing an afterdischarge, which begins with synaptic input releasing acetylcholine to open an ionotropic cholinergic receptor. Gating of this receptor causes depolarization and a shift from silence to continuous action potential firing, leading to the neuroendocrine secretion of egg-laying hormone and ovulation. At the onset of the afterdischarge, there is a rise in intracellular Ca2+, followed by both protein kinase C (PKC) activation and tyrosine dephosphorylation. To determine whether these signals influence the acetylcholine ionotropic receptor, we examined the bag cell neuron cholinergic response both in culture and isolated clusters using whole-cell and/or sharp-electrode electrophysiology. The acetylcholine-induced current was not altered by increasing intracellular Ca2+ via voltage-gated Ca2+ channels, clamping intracellular Ca2+ with exogenous Ca2+ buffers, or activating PKC with phorbol esters. However, lowering phosphotyrosine levels by inhibiting tyrosine kinases both reduced the cholinergic current and prevented acetylcholine from triggering action potentials or afterdischarge-like bursts. In other systems, acetylcholine receptors are often modulated by multiple signals, but bag cell neurons appear to be more restrictive in this regard. Prior work finds that, as the afterdischarge proceeds, tyrosine dephosphorylation leads to biophysical alterations that promote persistent firing. Because this firing is subsequent to the cholinergic input, inhibiting the acetylcholine receptor may represent a means of properly orchestrating synaptically induced changes in excitability.
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Affiliation(s)
- Sean H White
- Department of Biomedical and Molecular Sciences, Physiology and Neuroscience Graduate Programs, Centre for Neuroscience Studies, Queen's University, Kingston, ON K7L 3N6, Canada
| | - Raymond M Sturgeon
- Department of Biomedical and Molecular Sciences, Physiology and Neuroscience Graduate Programs, Centre for Neuroscience Studies, Queen's University, Kingston, ON K7L 3N6, Canada
| | - Yueling Gu
- Department of Biomedical and Molecular Sciences, Physiology and Neuroscience Graduate Programs, Centre for Neuroscience Studies, Queen's University, Kingston, ON K7L 3N6, Canada
| | - Alysha Nensi
- Department of Biomedical and Molecular Sciences, Physiology and Neuroscience Graduate Programs, Centre for Neuroscience Studies, Queen's University, Kingston, ON K7L 3N6, Canada
| | - Neil S Magoski
- Department of Biomedical and Molecular Sciences, Physiology and Neuroscience Graduate Programs, Centre for Neuroscience Studies, Queen's University, Kingston, ON K7L 3N6, Canada.
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Chen H, Zeng Q, Yao C, Cai Z, Wei T, Huang Z, Su J. Src family tyrosine kinase inhibitors suppress Nav1.1 expression in cultured rat spiral ganglion neurons. J Comp Physiol A Neuroethol Sens Neural Behav Physiol 2016; 202:185-93. [PMID: 26790420 DOI: 10.1007/s00359-016-1066-3] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/24/2015] [Revised: 12/22/2015] [Accepted: 01/01/2016] [Indexed: 11/25/2022]
Abstract
Src family kinases regulate neuronal voltage-gated Na(+) channels, which generate action potentials. The mechanisms of action, however, remain poorly understood. The aim of the present study was to further elucidate the effects of Src family kinases on Nav1.1 mRNA and protein expression in spiral ganglion neurons. Immunofluorescence staining techniques detected Nav1.1 expression in the spiral ganglion neurons. Additionally, quantitative PCR and western blot techniques were used to analyze Nav1.1 mRNA and protein expression, respectively, in spiral ganglion neurons following exposure to Src family kinase inhibitors PP2 (1 and 10 μM) and SU6656 (0.1 and 1 μM) for different lengths of time (6 and 24 h). In the spiral ganglion neurons, Nav1.1 protein expression was detected in the somas and axons. The Src family kinase inhibitors PP2 and SU6665 significantly decreased Nav1.1 mRNA and protein expression (p < 0.05), respectively, in the spiral ganglion neurons, and changes in expression were not dependent on time or dose (p > 0.05).
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Affiliation(s)
- Huiying Chen
- Department of Otolaryngology-Head and Neck Surgery, First Affiliated Hospital of Guangxi Medical University, Nanning, 530021, Guangxi, China
| | - Qingjiao Zeng
- Department of Otolaryngology-Head and Neck Surgery, First Affiliated Hospital of Guangxi Medical University, Nanning, 530021, Guangxi, China
| | - Chen Yao
- Department of Otolaryngology-Head and Neck Surgery, First Affiliated Hospital of Guangxi Medical University, Nanning, 530021, Guangxi, China
| | - Zheng Cai
- Department of Otolaryngology-Head and Neck Surgery, First Affiliated Hospital of Guangxi Medical University, Nanning, 530021, Guangxi, China
| | - Tingjia Wei
- Department of Otolaryngology-Head and Neck Surgery, First Affiliated Hospital of Guangxi Medical University, Nanning, 530021, Guangxi, China
| | - Zhihui Huang
- Department of Otolaryngology-Head and Neck Surgery, First Affiliated Hospital of Guangxi Medical University, Nanning, 530021, Guangxi, China
| | - Jiping Su
- Department of Otolaryngology-Head and Neck Surgery, First Affiliated Hospital of Guangxi Medical University, Nanning, 530021, Guangxi, China.
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Kabbani N, Nordman JC, Corgiat BA, Veltri DP, Shehu A, Seymour VA, Adams DJ. Are nicotinic acetylcholine receptors coupled to G proteins? Bioessays 2014; 35:1025-34. [PMID: 24185813 DOI: 10.1002/bies.201300082] [Citation(s) in RCA: 69] [Impact Index Per Article: 6.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/12/2022]
Abstract
It was, until recently, accepted that the two classes of acetylcholine (ACh) receptors are distinct in an important sense: muscarinic ACh receptors signal via heterotrimeric GTP binding proteins (G proteins), whereas nicotinic ACh receptors (nAChRs) open to allow flux of Na+, Ca2+, and K+ ions into the cell after activation. Here we present evidence of direct coupling between G proteins and nAChRs in neurons. Based on proteomic, biophysical, and functional evidence, we hypothesize that binding to G proteins modulates the activity and signaling of nAChRs in cells. It is important to note that while this hypothesis is new for the nAChR, it is consistent with known interactions between G proteins and structurally related ligand-gated ion channels. Therefore, it underscores an evolutionarily conserved metabotropic mechanism of G protein signaling via nAChR channels.
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Li SC, Yin HZ, Loudon WG, Weiss JH. Cultivating stem cells for treating amyotrophic lateral sclerosis. World J Stem Cells 2012; 4:117-119. [PMID: 23516096 PMCID: PMC3600561 DOI: 10.4252/wjsc.v4.i12.117] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 06/04/2012] [Revised: 09/17/2012] [Accepted: 12/20/2012] [Indexed: 02/06/2023] Open
Abstract
This editorial addresses the current challenges and future directions in the use of stem cells as an approach for treating amyotrophic lateral sclerosis. A wide variety of literature has been reviewed to enlighten the reader on the many facets of stem cell research that are important to consider before using them for a cell based therapy.
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Affiliation(s)
- Shengwen Calvin Li
- Shengwen Calvin Li, William G Loudon, CHOC Children's Hospital, University of California Irvine, Orange, CA 92868, United States
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