|
1
|
Zeng Z, Chen W, Moshensky A, Shakir Z, Khan R, Crotty Alexander LE, Ware LB, Aldaz CM, Jacobson JR, Dudek SM, Natarajan V, Machado RF, Singla S. Cigarette Smoke and Nicotine-Containing Electronic-Cigarette Vapor Downregulate Lung WWOX Expression, Which Is Associated with Increased Severity of Murine Acute Respiratory Distress Syndrome. Am J Respir Cell Mol Biol 2021; 64:89-99. [PMID: 33058734 PMCID: PMC7780991 DOI: 10.1165/rcmb.2020-0145oc] [Citation(s) in RCA: 4] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/18/2020] [Accepted: 09/22/2020] [Indexed: 12/21/2022] Open
Abstract
A history of chronic cigarette smoking is known to increase risk for acute respiratory distress syndrome (ARDS), but the corresponding risks associated with chronic e-cigarette use are largely unknown. The chromosomal fragile site gene, WWOX, is highly susceptible to genotoxic stress from environmental exposures and thus an interesting candidate gene for the study of exposure-related lung disease. Lungs harvested from current versus former/never-smokers exhibited a 47% decrease in WWOX mRNA levels. Exposure to nicotine-containing e-cigarette vapor resulted in an average 57% decrease in WWOX mRNA levels relative to vehicle-treated controls. In separate studies, endothelial (EC)-specific WWOX knockout (KO) versus WWOX flox control mice were examined under ARDS-producing conditions. EC WWOX KO mice exhibited significantly greater levels of vascular leak and histologic lung injury. ECs were isolated from digested lungs of untreated EC WWOX KO mice using sorting by flow cytometry for CD31+ CD45-cells. These were grown in culture, confirmed to be WWOX deficient by RT-PCR and Western blotting, and analyzed by electric cell impedance sensing as well as an FITC dextran transwell assay for their barrier properties during methicillin-resistant Staphylococcus aureus or LPS exposure. WWOX KO ECs demonstrated significantly greater declines in barrier function relative to cells from WWOX flox controls during either methicillin-resistant S. aureus or LPS treatment as measured by both electric cell impedance sensing and the transwell assay. The increased risk for ARDS observed in chronic smokers may be mechanistically linked, at least in part, to lung WWOX downregulation, and this phenomenon may also manifest in the near future in chronic users of e-cigarettes.
Collapse
Affiliation(s)
- Zhenguo Zeng
- Department of Critical Care Medicine, The First Affiliated Hospital of Nanchang University, Nanchang, Jiangxi, People’s Republic of China
| | - Weiguo Chen
- Division of Pulmonary, Critical Care, Sleep and Allergy Medicine, University of Illinois, Chicago, Illinois
| | | | - Zaid Shakir
- Division of Pulmonary, Critical Care, Sleep and Allergy Medicine, University of Illinois, Chicago, Illinois
| | - Raheel Khan
- Division of Pulmonary, Critical Care, Sleep and Allergy Medicine, University of Illinois, Chicago, Illinois
| | | | | | - C. M. Aldaz
- MD Anderson Cancer Center, University of Texas, Houston, Texas; and
| | - Jeffrey R. Jacobson
- Division of Pulmonary, Critical Care, Sleep and Allergy Medicine, University of Illinois, Chicago, Illinois
| | - Steven M. Dudek
- Division of Pulmonary, Critical Care, Sleep and Allergy Medicine, University of Illinois, Chicago, Illinois
| | - Viswanathan Natarajan
- Division of Pulmonary, Critical Care, Sleep and Allergy Medicine, University of Illinois, Chicago, Illinois
| | - Roberto F. Machado
- Division of Pulmonary, Critical Care, Sleep, and Occupational Medicine, Indiana University School of Medicine, Indianapolis, Indiana
| | - Sunit Singla
- Division of Pulmonary, Critical Care, Sleep and Allergy Medicine, University of Illinois, Chicago, Illinois
| |
Collapse
|
|
2
|
Palumbo E, Russo A. Common fragile site instability in normal cells: Lessons and perspectives. Genes Chromosomes Cancer 2018; 58:260-269. [PMID: 30387295 DOI: 10.1002/gcc.22705] [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: 08/07/2018] [Revised: 09/25/2018] [Accepted: 10/01/2018] [Indexed: 12/26/2022] Open
Abstract
Mechanisms and events related to common fragile site (CFS) instability are well known in cancer cells. Here, we argue that normal cells remain an important experimental model to address questions related to CFS instability in the absence of alterations in cell cycle and DNA damage repair pathways, which are common features acquired in cancer. Furthermore, a major gap of knowledge concerns the stability of CFSs during gametogenesis. CFS instability in meiotic or postmeiotic stages of the germ cell line could generate chromosome deletions or large rearrangements. This in turn can lead to the functional loss of the several CFS-associated genes with tumor suppressor function. Our hypothesis is that such mutations can potentially result in genetic predisposition to develop cancer. Indirect evidence for CFS instability in human germ cells has been provided by genomic investigations in family pedigrees associated with genetic disease. The issue of CFS instability in the germ cell line should represent one of the future efforts, and may take advantage of the existence of sequence and functional conservation of CFSs between rodents and humans.
Collapse
Affiliation(s)
- Elisa Palumbo
- Department of Molecular Medicine, University of Padova, Padova, Italy
| | - Antonella Russo
- Department of Molecular Medicine, University of Padova, Padova, Italy
| |
Collapse
|
|
3
|
Pospiech K, Płuciennik E, Bednarek AK. WWOX Tumor Suppressor Gene in Breast Cancer, a Historical Perspective and Future Directions. Front Oncol 2018; 8:345. [PMID: 30211123 PMCID: PMC6121138 DOI: 10.3389/fonc.2018.00345] [Citation(s) in RCA: 23] [Impact Index Per Article: 3.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/14/2018] [Accepted: 08/06/2018] [Indexed: 11/18/2022] Open
Abstract
The WWOX tumor suppressor gene is located at 16q23. 1–23.2, which covers the region of FRA16D—a common fragile sites. Deletions within the WWOX coding sequence are observed in up to 80% of breast cancer cases, which makes it one of the most common genetic alterations in this tumor type. The WWOX gene is known to play a role in breast cancer: increased expression of WWOX inhibits cell proliferation in suspension, reduces tumor growth rates in xenographic transplants, but also enhances cell migration through the basal membrane and contributes to morphological changes in 3D matrix-based cell cultures. The WWOX protein may act in several ways, as it has three functional domains—two WW domains, responsible for protein-protein interactions and an SDR domain (short dehydrogenase/reductase domain) which catalyzes conversions of low molecular weight ligands, most likely steroids. In epithelial cells, WWOX modulates gene transcription through interaction with p73, AP-2γ, and ERBB4 proteins. In steroid hormone-regulated tissues like mammary gland epithelium, the WWOX SDR domain acts as a steroid dehydrogenase. The relationship between WWOX and hormone receptors was shown in an animal model, where WWOX(C3H)+/–mice exhibited loss of both ER and PR receptors. Moreover, in breast cancer specimens, a positive correlation was observed between WWOX expression and ER status. On the other hand, decreased WWOX expression was associated with worse prognosis, namely higher relapse and mortality rates in BC patients. Recently, it was shown that genomic instability might be driven by the loss of WWOX expression. It was reported that WWOX plays role in DNA damage response (DDR) and DNA repair by regulating ATM activation through physical interaction. A genome caretaker function has also been proposed for WWOX, as it was found that WWOX sufficiency decreases homology directed repair (HDR) and supports non-homologous end-joining (NHEJ) repair as the dominant DSB repair pathway by Brca1-Wwox interaction. In breast cancer cells, WWOX was also found to modulate the expression of glycolysis pathway genes, through hypoxia-inducible transcription factor 1α (HIF1α) regulation. The paper presents the current state of knowledge regarding the WWOX tumor suppressor gene in breast cancer, as well as future research perspectives.
Collapse
Affiliation(s)
- Karolina Pospiech
- Department of Molecular Carcinogenesis, Medical University of Lodz, Lodz, Poland
| | - Elzbieta Płuciennik
- Department of Molecular Carcinogenesis, Medical University of Lodz, Lodz, Poland
| | - Andrzej K Bednarek
- Department of Molecular Carcinogenesis, Medical University of Lodz, Lodz, Poland
| |
Collapse
|
|
4
|
Menconi G, Bedini A, Barale R, Sbrana I. Global mapping of DNA conformational flexibility on Saccharomyces cerevisiae. PLoS Comput Biol 2015; 11:e1004136. [PMID: 25860149 PMCID: PMC4393290 DOI: 10.1371/journal.pcbi.1004136] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/30/2014] [Accepted: 01/16/2015] [Indexed: 11/25/2022] Open
Abstract
In this study we provide the first comprehensive map of DNA conformational flexibility in Saccharomyces cerevisiae complete genome. Flexibility plays a key role in DNA supercoiling and DNA/protein binding, regulating DNA transcription, replication or repair. Specific interest in flexibility analysis concerns its relationship with human genome instability. Enrichment in flexible sequences has been detected in unstable regions of human genome defined fragile sites, where genes map and carry frequent deletions and rearrangements in cancer. Flexible sequences have been suggested to be the determinants of fragile gene proneness to breakage; however, their actual role and properties remain elusive. Our in silico analysis carried out genome-wide via the StabFlex algorithm, shows the conserved presence of highly flexible regions in budding yeast genome as well as in genomes of other Saccharomyces sensu stricto species. Flexibile peaks in S. cerevisiae identify 175 ORFs mapping on their 3'UTR, a region affecting mRNA translation, localization and stability. (TA)n repeats of different extension shape the central structure of peaks and co-localize with polyadenylation efficiency element (EE) signals. ORFs with flexible peaks share common features. Transcripts are characterized by decreased half-life: this is considered peculiar of genes involved in regulatory systems with high turnover; consistently, their function affects biological processes such as cell cycle regulation or stress response. Our findings support the functional importance of flexibility peaks, suggesting that the flexible sequence may be derived by an expansion of canonical TAYRTA polyadenylation efficiency element. The flexible (TA)n repeat amplification could be the outcome of an evolutionary neofunctionalization leading to a differential 3'-end processing and expression regulation in genes with peculiar function. Our study provides a new support to the functional role of flexibility in genomes and a strategy for its characterization inside human fragile sites.
Collapse
Affiliation(s)
- Giulia Menconi
- Dip. Informatica, Università di Pisa, Largo Pontecorvo, Pisa, Italy
- Istituto Nazionale di Alta Matematica “Francesco Severi”, Piazzale Aldo Moro, Città Universitaria, Roma, Italy
| | - Andrea Bedini
- Dept. Mathematics and Statistics, The University of Melbourne Victoria, Australia
| | - Roberto Barale
- Dip. Biologia, Università di Pisa, Via Derna, Pisa, Italy
| | | |
Collapse
|
|
5
|
Abstract
WWOX is a gene that spans an extremely large chromosomal region. It is derived from within chromosomal band 16q23.2 which is a region with frequent deletions and other alterations in a variety of different cancers. This chromosomal band also contains the FRA16D common fragile site (CFS). CFSs are chromosomal regions found in all individuals which are highly unstable. WWOX has also been demonstrated to function as a tumor suppressor that is involved in the development of many cancers. Two other highly unstable CFSs, FRA3B (3p14.2) and FRA6E (6q26), also span extremely large genes, FHIT and PARK2, respectively, and these two genes are also found to be important tumor suppressors. There are a number of interesting similarities between these three large CFS genes. In spite of the fact that they are derived from some of the most unstable chromosomal regions in the genome, they are found to be highly evolutionarily conserved and the chromosomal region spanning the mouse homologs of both WWOX and FHIT are also CFSs in mice. Many of the other CFSs also span extremely large genes and many of these are very attractive tumor suppressor candidates. WWOX is therefore a member of a very interesting family of very large CFS genes.
Collapse
Affiliation(s)
- Ge Gao
- Division of Experimental Pathology, Department of Laboratory Medicine and Pathology, Mayo Clinic, Rochester, MN 55905, USA
| | - David I Smith
- Division of Experimental Pathology, Department of Laboratory Medicine and Pathology, Mayo Clinic, Rochester, MN 55905, USA
| |
Collapse
|
|
6
|
Gao G, Smith DI. Very large common fragile site genes and their potential role in cancer development. Cell Mol Life Sci 2014; 71:4601-15. [PMID: 25300511 PMCID: PMC11113612 DOI: 10.1007/s00018-014-1753-6] [Citation(s) in RCA: 31] [Impact Index Per Article: 2.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/29/2014] [Accepted: 09/30/2014] [Indexed: 10/24/2022]
Abstract
Common fragile sites (CFSs) are large chromosomal regions that are hot-spots for alterations especially within cancer cells. The three most frequently expressed CFS regions (FRA3B, FRA16D and FRA6E) contain genes that span extremely large genomic regions (FHIT, WWOX and PARK2, respectively), and these genes were found to function as important tumor suppressors. Many other CFS regions contain extremely large genes that are also targets of alterations in multiple cancers, but none have yet been demonstrated to function as tumor suppressors. The loss of expression of just FHIT or WWOX has been found to be associated with a worse overall clinical outcome. Studies in different cancers have revealed that some cancers have decreased expression of multiple large CFS genes. This loss of expression could have a profound phenotypic effect on these cells. In this review, we will summarize the known large common fragile site genes and discuss their potential relationship to cancer development.
Collapse
Affiliation(s)
- Ge Gao
- Division of Experimental Pathology, Department of Laboratory Medicine and Pathology, Mayo Clinic, Rochester, MN 55905 USA
| | - David I. Smith
- Division of Experimental Pathology, Department of Laboratory Medicine and Pathology, Mayo Clinic, Rochester, MN 55905 USA
| |
Collapse
|
|
7
|
AlFaisal AHM, AL-Ramahi IJK, Abdul-Hassan IAR. Micronucleus frequency among Iraqi thyroid disorder patients. COMPARATIVE CLINICAL PATHOLOGY 2012; 23:683-688. [PMID: 24829552 PMCID: PMC4016807 DOI: 10.1007/s00580-012-1671-7] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 04/13/2012] [Accepted: 12/12/2012] [Indexed: 12/05/2022]
Abstract
Micronucleus (MN) assay has been extensively used in detection of DNA damage, instability in cancer, and genetic disorders. In the current study, MN, binucleated cells, and nuclear division index (NDI) were investigated in Iraqi patients with thyroid disorders. The results indicated significantly (p < 0.05) increased binucleated cells with micronucleus (BNMN) frequencies in thyroid cancer group (37.58 ± 3.07) versus other thyroid disorder groups (6.60 ± 1.29, 14.90 ± 1.69, 15.56 ± 1.76). On the other hand, the frequency of micronucleus per 1,000 and the NDI were significantly (p < 0.05) decreased in hypothyroidism (MN 1.55 ± 0.36) (NDI 0.009 ± 0.001) versus other thyroid disorder groups (MN: 6.05 ± 0.97, 6.09 ± 0.53, 5.34 ± 0.56) (NDI: 0.049 ± 0.003, 0.032 ± 0.002, 0.025 ± 0.002), with no difference versus healthy group (0.0 ± 0.0). The number of BNMN and MN are parallel to the severity of thyroid disorders which were 6.60 ± 1.29, 14.90 ± 1.69, 15.56 ± 1.76, and 37.58 ± 3.07 for hypothyroidism, thyroid toxic goiter, thyroid nontoxic goiter, and thyroid cancer, respectively. The number of BNMN and MN are parallel to the severity of thyroid disorders which were 6.60 ± 1.29, 14.90 ± 1.69, 15.56 ± 1.76, and 37.58 ± 3.07 for hypothyroidism, thyroid toxic goiter, thyroid nontoxic goiter, and thyroid cancer, respectively. The results also indicate that there were no significant differences among age and sex groups as related with BNMN formation within each thyroid disorder groups and healthy control group.
Collapse
|
|
8
|
Hu BS, Tan JW, Zhu GH, Wang DF, Zhou X, Sun ZQ. WWOX induces apoptosis and inhibits proliferation of human hepatoma cell line SMMC-7721. World J Gastroenterol 2012; 18:3020-6. [PMID: 22736928 PMCID: PMC3380332 DOI: 10.3748/wjg.v18.i23.3020] [Citation(s) in RCA: 27] [Impact Index Per Article: 2.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 07/11/2011] [Revised: 09/28/2011] [Accepted: 01/07/2012] [Indexed: 02/06/2023] Open
Abstract
AIM: To investigate the effects of the WWOX gene on the human hepatic carcinoma cell line SMMC-7721.
METHODS: Full-length WWOX cDNA was amplified from normal human liver tissues. Full-length cDNA was subcloned into pEGFP-N1, a eukaryotic expression vector. After introduction of the WWOX gene into cancer cells using liposomes, the WWOX protein level in the cells was detected through Western blotting. Cell growth rates were assessed by methyl thiazolyl tetrazolium (MTT) and colony formation assays. Cell cycle progression and cell apoptosis were measured by flow cytometry. The phosphorylated protein kinase B (AKT) and activated fragments of caspase-9 and caspase-3 were examined by Western blotting analysis.
RESULTS: WWOX significantly inhibited cell proliferation, as evaluated by the MTT and colony formation assays. Cells transfected with WWOX showed significantly higher apoptosis ratios when compared with cells transfected with a mock plasmid, and overexpression of WWOX delayed cell cycle progression from G1 to S phase, as measured by flow cytometry. An increase in apoptosis was also indicated by a remarkable activation of caspase-9 and caspase-3 and a dephosphorylation of AKT (Thr308 and Ser473) measured with Western blotting analysis.
CONCLUSION: Overexpression of WWOX induces apoptosis and inhibits proliferation of the human hepatic carcinoma cell line SMMC-7721.
Collapse
|
|
9
|
Ekizoglu S, Muslumanoglu M, Dalay N, Buyru N. Genetic alterations of the WWOX gene in breast cancer. Med Oncol 2011; 29:1529-35. [PMID: 21983861 DOI: 10.1007/s12032-011-0080-0] [Citation(s) in RCA: 20] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/08/2011] [Accepted: 09/26/2011] [Indexed: 11/26/2022]
Abstract
FRA3B and FRA16D are the most sensitive common chromosomal fragile site loci in the human genome and two tumor suppressor genes FHIT (Fragile Histidine Triad) and WWOX (WW domain-containing oxidoreductase gene) map to this sites. The WWOX gene is composed of 9 exons and encodes a 46-kD protein that contains 414 amino acids. Loss of heterozygosity, homozygous deletions, and chromosomal translocations affecting WWOX has been reported in several types of cancer, including ovarian, esophageal, lung and stomach carcinoma, and multiple myeloma. The aim of this study was to determine the role of WWOX as a tumor suppressor gene in patients with breast cancer. Tumor and adjacent non-cancerous tissue samples were obtained from 81 patients with breast cancer. DNA was isolated from all tissue samples, and all exons and flanking intronic sequences of the WWOX gene were analyzed by PCR amplification and direct sequencing. We detected 14 different alterations in the coding sequence and one base substitution at the intron 6 splice site (+1 G-A). In addition to exonic and splice-site alterations, we detected 23 different alterations in the non-coding region of the gene. All coding region mutations identified in this study were in the exons between 4 and 9. We did not observe any alterations in exons 1-3. We conclude that mutations in critical region of the WWOX gene are frequent and may have an important role in breast carcinogenesis.
Collapse
Affiliation(s)
- Seda Ekizoglu
- Cerrahpasa Medical Faculty, Department of Medical Biology, Istanbul University, Kocamustafapasa, 34098 Istanbul, Turkey
| | | | | | | |
Collapse
|
|
10
|
Evidences that the polymorphism Pro-282-Ala within the tumor suppressor gene WWOX is a new risk factor for differentiated thyroid carcinoma. Int J Cancer 2011; 129:2816-24. [DOI: 10.1002/ijc.25937] [Citation(s) in RCA: 27] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/26/2010] [Accepted: 12/20/2010] [Indexed: 11/07/2022]
|
|
11
|
Molecular analysis of WWOX expression correlation with proliferation and apoptosis in glioblastoma multiforme. J Neurooncol 2010; 101:207-13. [PMID: 20535528 PMCID: PMC2996532 DOI: 10.1007/s11060-010-0254-1] [Citation(s) in RCA: 24] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/21/2009] [Accepted: 05/24/2010] [Indexed: 12/27/2022]
Abstract
Glioblastoma multiforme is the most common type of primary brain tumor in adults. WWOX is a tumor suppressor gene involved in carcinogenesis and cancer progression in many different neoplasms. Reduced WWOX expression is associated with more aggressive phenotype and poor patient outcome in several cancers. We investigated alternations of WWOX expression and its correlation with proliferation, apoptosis and signal trafficking in 67 glioblastoma multiforme specimens. Moreover, we examined the level of WWOX LOH and methylation status in WWOX promoter region. Our results suggest that loss of heterozygosity (relatively frequent in glioblastoma multiforme) along with promoter methylation may decrease the expression of this tumor suppressor gene. Our experiment revealed positive correlations between WWOX and Bcl2 and between WWOX and Ki67. We also confirmed that WWOX is positively correlated with ErbB4 signaling pathway in glioblastoma multiforme.
Collapse
|
|
12
|
Recurrent Chromosomal Alterations in Molecularly Classified AIDS-Related Lymphomas: An Integrated Analysis of DNA Copy Number and Gene Expression. J Acquir Immune Defic Syndr 2010; 54:18-26. [DOI: 10.1097/qai.0b013e3181d3d9eb] [Citation(s) in RCA: 19] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/26/2022]
|
|
13
|
Scarpato R, Tusa I, Antonelli A, Fallhai P, Sbrana I. Spontaneous and bleomycin-induced chromosome damage in non cancer thyroid patients. Eur J Clin Invest 2009; 39:1091-7. [PMID: 19807783 DOI: 10.1111/j.1365-2362.2009.02214.x] [Citation(s) in RCA: 9] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 12/22/2022]
Abstract
BACKGROUND Presence of chromosome damage in lymphocytes of patients affected by several diseases, including cancer, was detected by the micronucleus (MN) assay. Individual susceptibility to DNA damage, considered as a risk factor for cancer, can be also evaluated using the bleomycin (BLM) sensitivity test. MATERIALS AND METHODS We aimed to evaluate spontaneous or BLM-induced MN frequencies in autoimmune (AI, n = 19) and non autoimmune (NAI, n = 11) thyroid patients, not receiving (131)I radiometabolic therapy with respect to a control group of 18 healthy subjects. According to thyroid function, patients were also divided into hypothyroid (n = 10), euthyroid (n = 13) or hyperthyroid (n = 7) subjects. RESULTS Spontaneous MN frequencies of AI and NAI patients did not differ from those of controls. Hypothyroid patients had more elevated MN basal levels (9.00 + or - 1.71 per thousand) than hyperthyroid (3.75 + or - 1.17 per thousand, P < 0.05) and euthyroid (5.38 + or - 0.97 per thousand, P < 0.01) patients or healthy subjects (4.17 + or - 0.63 per thousand, P < 0.01). In particular, the hypothyroid AI group showed the highest value (9.79 + or - 2.26 per thousand, P < 0.01). All thyroid patients responded differently to BLM than controls (39.90 + or - 2.48 per thousand vs. 31.08 + or - 2.51 per thousand, P = 0.0377). The NAI group had BLM-induced MN levels (45.00 + or - 2.56 per thousand) significantly higher (P = 0.0215) than AI patients (36.95 + or - 3.49 per thousand) or healthy subjects (31.08 + or - 2.51 per thousand). No significant difference was seen when patients were stratified according to autoimmunity. CONCLUSIONS We report that hypothyroid patients exhibit a moderate increase in the level of spontaneous genome damage, and that AI thyroid patients resulted to be less sensitive than NAI patients to the mutagen sensitivity test. In prospective, it may be of interest to reinvestigate hypothyroid patients when correction of their dysfunction is achieved.
Collapse
Affiliation(s)
- R Scarpato
- Dipartimento di Biologia, University of Pisa, Via Derna 1, 56100 Pisa, Italy.
| | | | | | | | | |
Collapse
|
|
14
|
Giarnieri E, Zanesi N, Bottoni A, Alderisio M, Lukic A, Vecchione A, Ziparo V, Croce CM, Mancini R. Oncosuppressor proteins of fragile sites are reduced in cervical cancer. Cancer Lett 2009; 289:40-5. [PMID: 19700237 DOI: 10.1016/j.canlet.2009.07.017] [Citation(s) in RCA: 15] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/23/2009] [Revised: 07/15/2009] [Accepted: 07/20/2009] [Indexed: 11/16/2022]
Abstract
FHIT and WWOX are tumor suppressor genes that span the common fragile sites FRA3B and FRA16D, respectively. To analyze possible synergisms among these genes in cervical cancer progression, we considered 159 cervical intraepithelial neoplasias, and 58 invasive squamous cell carcinomas of the uterine cervix. All cases were previously selected as high risk HPV. FHIT and WWOX proteins were examined by immunohistochemistry and their expression was inversely correlated with precancerous vs. invasive lesions. Statistics among biological markers indicated an association between FHIT and WWOX. Protein expression of these two genes was also absent or reduced in cancer cell lines. Thus, WWOX may be considered as a novel important genetic marker in cervical cancer and the association between the altered expression of FHIT and WWOX may be a critical event in the progression of this neoplasia.
Collapse
Affiliation(s)
- Enrico Giarnieri
- Cytopathology, II Faculty of Medicine, University of Rome La Sapienza, Sant'Andrea Hospital, Rome, Italy
| | | | | | | | | | | | | | | | | |
Collapse
|
|
15
|
Guler G, Huebner K, Himmetoglu C, Jimenez RE, Costinean S, Volinia S, Pilarski RT, Hayran M, Shapiro CL. Fragile histidine triad protein, WW domain-containing oxidoreductase protein Wwox, and activator protein 2gamma expression levels correlate with basal phenotype in breast cancer. Cancer 2009; 115:899-908. [PMID: 19130459 DOI: 10.1002/cncr.24103] [Citation(s) in RCA: 36] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/09/2022]
Abstract
BACKGROUND The expression of fragile histidine triad protein (Fhit) and WW domain-containing oxidoreductase protein (Wwox), tumor suppressors that are encoded by fragile (FRA) loci FRA3B and FRA16D, are lost concordantly in breast cancers. In the current study, the authors examined correlations among Fhit, Wwox, the activator protein 2 transcription factors AP2alpha and AP2gamma, cytokeratins 5 and 6 (CK5/6), epidermal growth factor receptor (EGFR), estrogen receptor (ER), progesterone receptor (PR), and human epidermal growth factor receptor 2 (HER-2) and their associations with breast cancer phenotypes. METHODS Tissue microarrays constructed from 837 breast cancer blocks were immunostained. Expression in >10% of tumor cells was considered positive for cytoplasmic CK5/6, membranous EGFR, and nuclear AP2alpha and AP2gamma. Cytoplasmic Fhit and Wwox staining was scored according to staining intensity. ER, PR, and HER-2 status of tumors was derived from records. Correlations among immunohistochemical markers and tumor subtypes were assessed by univariate and multivariate statistical methods. RESULTS Triple-negative tumors had more frequent expression of EGFR, CK5/6 (P < .001), and AP2gamma (P = .003) and more frequent loss of Fhit and Wwox (P < .001), and an inverse correlation was observed between Fhit, Wwox expression and EGFR, ER, and PR expression (P < .001). Reduced Fhit expression was more common in HER-2-positive and AP2gamma-positive cases (P < .001 and P = .002, respectively). There was a direct correlation noted between Fhit and Wwox (P < .001) and a borderline positive relation between AP2alpha and AP2gamma (P = .054). CONCLUSIONS The results from this investigation suggested that reduced expression levels of Fhit, Wwox, and nuclear AP2gamma have roles in the pathogenesis of basal-like differentiation in breast cancer. Alteration in the expression of fragile site genes occurs in most of these cancers and may contribute to defects in DNA repair, as observed in breast cancer 1 (BRCA1)-deficient cancers. Thus, DNA damage response checkpoint proteins may be targets for treatment.
Collapse
Affiliation(s)
- Gulnur Guler
- Department of Pathology, Hacettepe University, Ankara, Turkey
| | | | | | | | | | | | | | | | | |
Collapse
|
|
16
|
Lo CP, Hsu LJ, Li MY, Hsu SY, Chuang JI, Tsai MS, Lin SR, Chang NS, Chen ST. MPP+-induced neuronal death in rats involves tyrosine 33 phosphorylation of WW domain-containing oxidoreductase WOX1. Eur J Neurosci 2008; 27:1634-46. [PMID: 18371080 DOI: 10.1111/j.1460-9568.2008.06139.x] [Citation(s) in RCA: 34] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/28/2022]
Abstract
WW domain-containing oxidoreductase (named WWOX, FOR or WOX1) is a pro-apoptotic protein and tumor suppressor. Animals treated with dopaminergic neurotoxin 1-methyl-4-phenyl-pyridinium (MPP+) develop Parkinson's disease (PD)-like symptoms. Here we investigated whether WOX1 is involved in MPP+-induced neurodegeneration. Upon insult with MPP+ in rat brains, WOX1 protein was upregulated and phosphorylated at Tyr33 (or activated) in the injured neurons in the striatum and cortex ipsilaterally to intoxication, as determined by immunohistochemistry and Western blotting. Also, WOX1 was present in the condensed nuclei and damaged mitochondria of degenerative neurons, as revealed by transmission immunoelectron microscopy. Time-lapse microscopy revealed that MPP+ induced membrane blebbing and shrinkage of neuroblastoma SK-N-SH cells. Dominant-negative WOX1, a potent inhibitor of Tyr33 phosphorylation, abolished this event, indicating a critical role of the phosphorylation in apoptosis. c-Jun N-terminal kinase (JNK1) is known to bind and counteract the apoptotic function of WOX1. Suppression of JNK1 function by a dominant-negative spontaneously induced WOX1 activation. WOX1 physically interacted with JNK1 in SK-N-SH cells and rat brain extracts. MPP+ rapidly increased the binding, followed by dissociation, which is probably needed for WOX1 to exert apoptosis. We synthesized a short Tyr33-phosphorylated WOX1 peptide (11 amino acid residues). Interestingly, this peptide blocked MPP+-induced neuronal death in the rat brains, whereas non-phospho-WOX1 peptide had no effect. Together, activated WOX1 plays an essential role in the MPP+-induced neuronal death. Our synthetic phospho-WOX1 peptide prevents neuronal death, suggestive of its therapeutic potential in mitigating the symptoms of PD.
Collapse
Affiliation(s)
- Chen-Peng Lo
- Institute of Basic Medical Sciences, National Cheng Kung University, Tainan, Taiwan 70101, ROC
| | | | | | | | | | | | | | | | | |
Collapse
|
|
17
|
Dias EP, Pimenta FJ, Sarquis MS, Dias Filho MA, Aldaz CM, Fujii JB, Gomez RS, De Marco L. Association between decreased WWOX protein expression and thyroid cancer development. Thyroid 2007; 17:1055-9. [PMID: 18047428 PMCID: PMC4150466 DOI: 10.1089/thy.2007.0232] [Citation(s) in RCA: 15] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 11/12/2022]
Abstract
CONTEXT Chromosomal fragile sites are often related to cancer development. The WW domain-containing oxidoreductase gene (WWOX) spans the second most common chromosomal fragile site (FRA16D) and encodes an important proapoptotic protein. OBJECTIVE To verify our hypothesis that underexpression of WWOX could contribute to malignant transformation of the thyroid cells. METHOD We compared WWOX expression among follicular adenomas (FAs) and differentiated thyroid carcinomas [follicular thyroid carcinomas (FTCs) and papillary thyroid carcinomas (PTCs)] in 53 thyroid tumors resected from patients submitted to total thyroidectomy. DESIGN Multiple fields of tumor areas of FAs, FTCs, and PTCs as well as normal thyroid tissue were stained with WWOX antiserum, and classified by the extent of staining (percentage of cells staining) and staining intensity. MAIN OUTCOME PTCs showed a significantly decreased expression of WWOX when compared to FAs and FTCs. Further, using a unique model of comparison in patients in whom FAs and PTCs were concomitantly present, we detected the same result (i.e., no expression in PTCs). CONCLUSION We conclude that WWOX underexpression is an important step that might increase the vulnerability to the carcinogenesis process in PTCs.
Collapse
Affiliation(s)
- Eduardo P Dias
- Department of Pharmacology, Universidade Federal de Minas Gerais, Belo Horizonte, Brazil.
| | | | | | | | | | | | | | | |
Collapse
|
|
18
|
Chang NS, Hsu LJ, Lin YS, Lai FJ, Sheu HM. WW domain-containing oxidoreductase: a candidate tumor suppressor. Trends Mol Med 2006; 13:12-22. [PMID: 17142102 DOI: 10.1016/j.molmed.2006.11.006] [Citation(s) in RCA: 99] [Impact Index Per Article: 5.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/05/2006] [Revised: 11/03/2006] [Accepted: 11/22/2006] [Indexed: 10/23/2022]
Abstract
Common fragile site gene WWOX encodes a candidate tumor suppressor WW domain-containing oxidoreductase. Alteration of this gene, along with dramatic downregulation of WWOX protein, is shown in the majority of invasive cancer cells. Ectopic WWOX exhibits proapoptotic and tumor inhibitory functions in vitro and in vivo, probably interacting with growth regulatory proteins p53, p73 and others. Hyaluronidases regulate WWOX expression, increase cancer invasiveness and seem to be involved in the development of hormone-independent growth of invasive cancer cells. Estrogen and androgen stimulate phosphorylation and nuclear translocation of WWOX, although binding of WWOX to these sex hormones is unknown. We propose that suppression of WWOX expression by overexpressed hyaluronidases might contribute in part to the development of hormone independence in invasive cancer.
Collapse
Affiliation(s)
- Nan-Shan Chang
- Institute of Molecular Medicine, National Cheng Kung University Medical College, Tainan, Taiwan 70101, Republic of China.
| | | | | | | | | |
Collapse
|
|
19
|
Re A, Cora D, Puliti AM, Caselle M, Sbrana I. Correlated fragile site expression allows the identification of candidate fragile genes involved in immunity and associated with carcinogenesis. BMC Bioinformatics 2006; 7:413. [PMID: 16981993 PMCID: PMC1601973 DOI: 10.1186/1471-2105-7-413] [Citation(s) in RCA: 18] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/27/2006] [Accepted: 09/18/2006] [Indexed: 01/26/2023] Open
Abstract
Background Common fragile sites (cfs) are specific regions in the human genome that are particularly prone to genomic instability under conditions of replicative stress. Several investigations support the view that common fragile sites play a role in carcinogenesis. We discuss a genome-wide approach based on graph theory and Gene Ontology vocabulary for the functional characterization of common fragile sites and for the identification of genes that contribute to tumour cell biology. Results Common fragile sites were assembled in a network based on a simple measure of correlation among common fragile site patterns of expression. By applying robust measurements to capture in quantitative terms the non triviality of the network, we identified several topological features clearly indicating departure from the Erdos-Renyi random graph model. The most important outcome was the presence of an unexpected large connected component far below the percolation threshold. Most of the best characterized common fragile sites belonged to this connected component. By filtering this connected component with Gene Ontology, statistically significant shared functional features were detected. Common fragile sites were found to be enriched for genes associated to the immune response and to mechanisms involved in tumour progression such as extracellular space remodeling and angiogenesis. Moreover we showed how the internal organization of the graph in communities and even in very simple subgraphs can be a starting point for the identification of new factors of instability at common fragile sites. Conclusion We developed a computational method addressing the fundamental issue of studying the functional content of common fragile sites. Our analysis integrated two different approaches. First, data on common fragile site expression were analyzed in a complex networks framework. Second, outcomes of the network statistical description served as sources for the functional annotation of genes at common fragile sites by means of the Gene Ontology vocabulary. Our results support the hypothesis that fragile sites serve a function; we propose that fragility is linked to a coordinated regulation of fragile genes expression.
Collapse
Affiliation(s)
- Angela Re
- Dipartimento di Fisica Teorica dell'Università degli Studi di Torino e INFN, Via P. Giuria 1 – I 10125 Torino, Italy
| | - Davide Cora
- Dipartimento di Fisica Teorica dell'Università degli Studi di Torino e INFN, Via P. Giuria 1 – I 10125 Torino, Italy
- Centro Interdipartimentale Sistemi Complessi in Biologia e Medicina Molecolare, Via Accademia Albertina 13 – I 10123 Torino, Italy
| | - Alda Maria Puliti
- Dipartimento di Biologia dell'Università degli Studi di Pisa, Via San Giuseppe 22 – I 56126 Pisa, Italy
- Laboratorio di Genetica Molecolare, Istituto G.Gaslini, L.go Gaslini 5 – I 16147 Genova, Italy
| | - Michele Caselle
- Dipartimento di Fisica Teorica dell'Università degli Studi di Torino e INFN, Via P. Giuria 1 – I 10125 Torino, Italy
- Centro Interdipartimentale Sistemi Complessi in Biologia e Medicina Molecolare, Via Accademia Albertina 13 – I 10123 Torino, Italy
| | - Isabella Sbrana
- Dipartimento di Biologia dell'Università degli Studi di Pisa, Via San Giuseppe 22 – I 56126 Pisa, Italy
| |
Collapse
|