Transcriptional silencing of the inhibin-α gene in human gastric carcinoma cells

Kim,Young  Il; Shim,Jaejun; Kim,Byung-Ho; Lee,Sung  Jae; Lee,Ha  Kyu; Cho,Chunghee; Cho,Byung-Nam

doi:10.3892/ijo.2012.1472

Transcriptional silencing of the inhibin-α gene in human gastric carcinoma cells

Authors:
- Young Il Kim
- Jaejun Shim
- Byung-Ho Kim
- Sung Jae Lee
- Ha Kyu Lee
- Chunghee Cho
- Byung-Nam Cho
View Affiliations

Affiliations: Medical Science Research Institute, Kyung Hee University Medical Center, Seoul 130‑872, Republic of Korea, Department of Internal Medicine, Kyung Hee University School of Medicine, Seoul 130‑872, Republic of Korea, Department of Life Sciences, The Catholic University of Korea, Bucheon 420-743, Republic of Korea, Department of Life Science, Gwangju Institute of Science and Technology (K-JIST), Gwangju 500-712, Republic of Korea
Published online on: May 10, 2012 https://doi.org/10.3892/ijo.2012.1472
Pages: 690-700

Metrics: Total Views: 0 (Spandidos Publications: | PMC Statistics: )

Metrics: Total PDF Downloads: 0 (Spandidos Publications: | PMC Statistics: )

Cited By (CrossRef): 0 citations Loading Articles...

Abstract

Although inhibin was first identified as a hormone regulating pituitary FSH secretion, it was later recognized to act as a tumor suppressor in the gonad and adrenal glands. Recently, the alpha subunit of this dimeric hormone (inhibin‑α) was reported to be involved in prostate tumorigenesis. To identify additional roles outside the reproductive axis, we investigated inhibin‑α gene activity and subsequent cell fate in human gastric cancer cells. The results were as follows: all the gastric cancer cells had at least one of a set of abnormalities including hypermethylation of the promoter, mutation of the 5'‑UTR or allelic imbalance including LOH in the inhibin‑α gene. Hypermethylation of the promoter and mutation of the 5'‑UTR in inhibin‑α were observed in SNU‑1, SNU‑5 and SNU‑484 cells. LOH was observed in AGS, KATO III, SNU‑5, SNU‑484 and SNU‑668 cells. Treatment with 5‑AzaC, a demethylating agent, induced demethylation of the inhibin‑α promoter in the SNU‑1, SNU‑5 and SNU‑484 cell lines, with the CpG5 site being strongly influenced by 5‑AzaC. In addition, inhibin‑α mRNA and protein were maintained at low levels in most of the gastric cancer cell lines. These low levels of mRNA and protein expression could be increased in most lines by treatment with 5‑AzaC. These increased inhibin‑α expression levels seemed to be closely associated with apoptosis and suppression of cell growth. Taken together, our results reveal that the inhibin‑α gene is transcriptionally silenced in human gastric cancer cells, and that reactivation of the gene suppresses their growth characteristics. This suggests that inhibin‑α may have a more general tumor suppressor activity outside the reproductive axis.

View Figures

View References

1	Ying SY: Inhibins and activins: chemical properties and biological activity. Proc Soc Exp Biol Med. 186:253–264. 1987. View Article : Google Scholar : PubMed/NCBI
2	Ying SY: Inhibins, activins, follistatins: gonadal proteins modulating the secretion of follicle-stimulating hormone. Endocr Rev. 9:267–293. 1988. View Article : Google Scholar : PubMed/NCBI
3	Robertson DM, Giacometti M, Foulds LM, Lahnstein J, Goss NH, Hearn MTW and de Kretser DM: Isolation of inhibin α-subunit precursor proteins from bovine follicular fluid. Endocrinology. 125:2141–2149. 1989.
4	Hotten G, Neidhardt H, Schneider C and Pohl J: Cloning of a new member of the TGF-β family: a putative new activin β_C chain. Biochem Biophys Res Commun. 206:608–613. 1995.
5	Oda S, Nishimatsu S, Murakami K and Ueno N: Molecular cloning and functional analysis of a new activin β subunit: a dorsal mesoderm-inducing activity in Xenopus. Biochem Biophys Res Commun. 210:581–588. 1995.
6	Fang J, Yin W, Smiley E, Wang SQ and Bonadio J: Molecular cloning of the mouse activin β_E subunit gene. Biochem Biophys Res Commun. 228:669–674. 1996.
7	de Kretser DM, Hedger MP, Loveland KL and Phillips DJ: Inhibins, activins and follistatin in reproduction. Hum Reprod Update. 8:529–541. 2002.PubMed/NCBI
8	Findlay JK, Drummond AE, Dyson ML, Baillie AJ, Robertson DM and Ethier JF: Recruitment and development of the follicle; the roles of the transforming growth factor-β superfamily. Mol Cell Endocrinol. 191:35–43. 2002.
9	Juengel JL and McNatty KP: The role of proteins of the transforming growth factor-β superfamily in the intraovarian regulation of follicular development. Hum Reprod Update. 11:144–161. 2005.
10	Knight PG and Glister C: TGF-β superfamily members and ovarian follicle development. Reproduction. 132:191–206. 2006.
11	Chand AL, Ooi GT, Harrison CA, Shelling AN and Robertson DM: Functional analysis of the human inhibin α subunit variant A257T and its potential role in premature ovarian failure. Hum Reprod. 22:3241–3248. 2007.
12	Welt CK, Taylor AE, Fox J, Messerlian GM, Adams JM and Schneyer AL: Follicular arrest in polycystic ovary syndrome is associated with deficient inhibin A and B biosynthesis. J Clin Endocrinol Metab. 90:5582–5587. 2005. View Article : Google Scholar : PubMed/NCBI
13	Hamar BD, Buhimschi IA, Sfakianaki AK, Pettker CM, Magloire LK, Funai EF, Copel JA and Buhimschi CS: Serum and urine inhibin A but not free activin A are endocrine biomarkers of severe pre-eclampsia. Am J Obstet Gynecol. 195:1636–1645. 2006. View Article : Google Scholar : PubMed/NCBI
14	Tsigkou A, Marrelli D, Reis FM, et al: Total inhibin is a potential serum marker for epithelial ovarian cancer. J Clin Endocrinol Metab. 92:2526–2531. 2007. View Article : Google Scholar : PubMed/NCBI
15	Tsigkou A, Luisi S, Reis FM and Petraglia F: Inhibins as diagnostic markers in human reproduction. Adv Clin Chem. 45:1–29. 2008. View Article : Google Scholar : PubMed/NCBI
16	Cho BN, McMullen ML, Pei L, Yates CJ and Mayo KE: Reproductive deficiencies in transgenic mice expressing the rat inhibin α-subunit gene. Endocrinology. 142:4994–5004. 2001.PubMed/NCBI
17	Ahn JM, Jung HK, Cho C, Choi D, Mayo KE and Cho BN: Changes in the reproductive functions of mice due to injection of a plasmid expressing an inhibin α-subunit into muscle: a transient transgenic model. Mol Cell. 18:79–86. 2004.PubMed/NCBI
18	Luisi S, Florio P, Reis FM and Petraglia F: Inhibins in female and male reproductive physiology: role in gametogenesis, conception, implantation and early pregnancy. Hum Reprod Update. 11:123–135. 2005. View Article : Google Scholar : PubMed/NCBI
19	Risbridger GP, Schmitt JF and Robertson DM: Activins and inhibins in endocrine and other tumors. Endocr Rev. 22:836–858. 2001. View Article : Google Scholar : PubMed/NCBI
20	Matzuk MM, Finegold MJ, Su JG, Hsueh AJ and Bradley A: α-inhibin is a tumour-suppressor gene with gonadal specificity in mice. Nature. 360:313–319. 1992.
21	Matzuk MM and Bradley A: Identification and analysis of tumor suppressor genes using transgenic mouse models. Semin Cancer Biol. 5:37–45. 1994.PubMed/NCBI
22	Matzuk MM, Finegold MJ, Mather JP, Krummen L, Lu H and Bradley A: Development of cancer cachexia-like syndrome and adrenal tumors in inhibin-deficient mice. Proc Natl Acad Sci USA. 91:8817–8821. 1994. View Article : Google Scholar : PubMed/NCBI
23	Schmitt JF, Millar DS, Pedersen JS, et al: Hypermethylation of the inhibin α-subunit gene in prostate carcinoma. Mol Endocrinol. 16:213–220. 2002.
24	Balanathan P, Ball EMA, Wang H, Harris SE, Shelling AN and Risbridger GP: Epigenetic regulation of inhibin α-subunit gene in prostate cancer cell lines. J Mol Endocrinol. 32:55–67. 2004.
25	Balanathan P, Williams ED, Wang H, et al: Elevated level of inhibin-α subunit is pro-tumourigenic and pro-metastatic and associated with extracapsular spread in advanced prostate cancer. Br J Cancer. 100:1784–1793. 2009.
26	Imai K, Khandoker MAM, Yonai M, et al: Matrix metalloproteinases-2 and -9 activities in bovine follicular fluid of different-sized follicles: relationship to intra-follicular inhibin and steroid concentrations. Domest Anim Endocrinol. 24:171–183. 2003. View Article : Google Scholar
27	Jones RL, Findlay JK, Farnworth PG, Robertson DM, Wallace E and Salamonsen LA: Activin A and inhibin A differentially regulate human uterine matrix metalloproteinases: potential interactions during decidualization and trophoblast invasion. Endocrinology. 147:724–732. 2006. View Article : Google Scholar
28	Hempel N, How T, Dong M, Murphy SK, Fields TA and Blobe GC: Loss of betaglycan expression in ovarian cancer: role in motility and invasion. Cancer Res. 67:5231–5238. 2007. View Article : Google Scholar : PubMed/NCBI
29	Clark SJ, Harrison J, Paul CL and Frommer M: High sensitivity mapping of methylated cytosines. Nucleic Acids Res. 22:2990–2997. 1994. View Article : Google Scholar : PubMed/NCBI
30	Sundblad V, Chiauzzi VA, Andreone L, Campo S, Charreau EH and Dain L: Controversial role of inhibin α-subunit gene in the aetiology of premature ovarian failure. Hum Reprod. 21:1154–1160. 2006.
31	Jones PA and Baylin SB: 2002.The fundamental role of epigenetic events in cancer. Nat Rev Genet. 3:415–428. 2002.PubMed/NCBI
32	Song SH, Jong HS, Choi HH, Inoue H, Tanabe T, Kim NK and Bang YJ: Transcriptional silencing of cyclooxygenase-2 by hyper-methylation of the 5′ CpG island in human gastric carcinoma cells. Cancer Res. 61:4628–4635. 2001.PubMed/NCBI
33	Kuchler RJ: Development of animal cell populations in vitro. Biochemical Methods in Cell Culture and Virology. Kuchler RJ: Dowden, Hutchinson and Ross Inc. Press; Stroudsburg, PA: pp. 90–113. 1977
34	Garinis GA, Patrinos GP, Spanakis NE and Menounos PG: DNA hypermethylation: when tumour suppressor genes go silent. Hum Genet. 111:115–127. 2002. View Article : Google Scholar : PubMed/NCBI
35	Watson RH, Roy WJ, Davis M, Hitchcock A and Campbell IG: Loss of heterozygosity at the α-inhibin locus on chromosome 2q is not a feature of human granulosa cell tumors. Gynecol Oncol. 65:387–390. 1997.
36	Marozzi A, Porta C, Vegetti W, Crosignani PG, Tibiletti MG, Dalpra L and Ginelli E: Mutation analysis of the inhibin alpha gene in a cohort of Italian women affected by ovarian failure. Hum Reprod. 17:1741–1745. 2002. View Article : Google Scholar : PubMed/NCBI
37	Stenvers KL and Findlay JK: Inhibins: from reproductive hormones to tumor suppressors. Trends Endocrinol Metab. 21:174–180. 2010. View Article : Google Scholar : PubMed/NCBI
38	Barton DE, Yang-Feng TL, Mason AJ, Seeburg PH and Francke U: Mapping of genes for inhibin subunits α, β_A, and β_B on human and mouse chromosomes and studies of jsd mice. Genomics. 5:91–99. 1898.
39	Mellor SL, Richards MG, Pedersen JS, Robertson DM and Risbridger GP: Loss of the expression and localization of inhibin α-subunit in high grade prostate cancer. J Clin Endocrinol Metab. 83:969–975. 1998.
40	Vitale AM, Gonzalez OM, Parborell F, Irusta G, Campo S and Tesone M: Inhibin A increases apoptosis in early ovarian antral follicles of diethylstilbestrol-treated rats. Biol Reprod. 67:1989–1995. 2002. View Article : Google Scholar : PubMed/NCBI
41	Steller MD, Shaw TJ, Vanderhyden BC and Ethier JF: Inhibin resistance is associated with aggressive tumorigenicity of ovarian cancer cells. Mol Cancer Res. 3:50–61. 2005.PubMed/NCBI
42	Geng LY, Fang M, Yi JM, Jiang F, Moeen-ud-Din M and Yang LG: Effect of overexpression of inhibin α (1–32) fragment on bovine granulosa cell proliferation, apoptosis, steroidogenesis, and development of co-cultured oocytes. Theriogenology. 70:35–43. 2008.
43	Myers M, Middlebrook BS, Matzuk MM and Pangas SA: Loss of inhibin alpha uncouples oocyte-granulosa cell dynamics and disrupts postnatal folliculogenesis. Dev Biol. 334:458–467. 2009. View Article : Google Scholar : PubMed/NCBI
44	Oka M, Meacham AM, Hamazaki T, Rodić N, Chang LJ and Terada N: De novo DNA methyltransferases Dnmt3a and Dnmt3b primarily mediate the cytotoxic effect of 5-aza-2′-deoxycytidine. Oncogene. 24:3091–3099. 2005. View Article : Google Scholar
45	Fulda S and Debatin KM: 5-Aza-2′-deoxycytidine and IFN-γ cooperate to sensitize for TRAIL-induced apoptosis by upregulating caspase-8. Oncogene. 25:5125–5133. 2006.
46	Valdez BC, Li Y, Murray D, Corn P, Champlin RE and Andersson BS: 5-Aza-2′-deoxycytidine sensitizes busulfan-resistant myeloid leukemia cells by regulating expression of genes involved in cell cycle checkpoint and apoptosis. Leuk Res. 34:364–372. 2010.
47	Hanahan D and Weinberg RA: Hallmarks of cancer: the next generation. Cell. 144:646–674. 2011. View Article : Google Scholar : PubMed/NCBI