Register      Login
Reproduction, Fertility and Development Reproduction, Fertility and Development Society
Vertebrate reproductive science and technology
RESEARCH ARTICLE

Expression patterns of activin, inhibin and follistatin variants in the adult male mouse reproductive tract suggest important roles in the epididymis and vas deferens

Wendy R. Winnall A D E , Hui Wu A , Mai A. Sarraj B , Peter A. W. Rogers C , David M. de Kretser A , Jane E. Girling C and Mark P. Hedger A
+ Author Affiliations
- Author Affiliations

A Centre for Reproduction and Development, Monash Institute of Medical Research, Monash University, Clayton, Vic. 3168, Australia.

B Prince Henry’s Institute of Medical Research, Clayton, Vic. 3168, Australia.

C Department of Obstetrics and Gynaecology, The University of Melbourne, Vic. 3010, Australia.

D Present address: Department of Microbiology and Immunology, University of Melbourne, Vic. 3010, Australia.

E Corresponding author. Email: wwinnall@unimelb.edu.au

Reproduction, Fertility and Development 25(3) 570-580 https://doi.org/10.1071/RD11287
Submitted: 10 November 2011  Accepted: 27 April 2012   Published: 18 May 2012

Abstract

Activin A and its inhibitors follistatin and inhibin play key roles in development and function of the male reproductive tract. Quantitative (q) polymerase chain reaction (PCR) was used to evaluate the expression of Inhba (the gene encoding activin A subunits), Inha and Inhbb (genes encoding the inhibin B subunits), as well as the genes for follistatin (Fst) and follistatin-like 3 (Fstl3) and the activin receptor subunits, in the male mouse reproductive tract. A qPCR assay that discriminated between the two follistatin variants of Fst288 (tissue-bound form) and Fst315 (circulating) was established. Activin A protein was measured by ELISA, whereas the inhibin α-subunit and total follistatin proteins were measured by radioimmunoassay (RIA). A screen of 22 tissues demonstrated tissue-specific regulation of the follistatin variants, with Fst288 highly expressed in the vas deferens and Fst315 most highly expressed in the skin. The expression of Fst288 and Fst315 and follistatin protein levels increased progressively from the testis through to the distal vas deferens. Inhba and the activin receptors were highly expressed in the epididymis, but activin A protein was elevated in both the epididymis and vas deferens. Inhibin α-subunit mRNA and protein and Inhbb expression were highest in the testis. These results indicate a role for activin A within the epididymis, but also that activin A bioactivity may be increasingly inhibited by follistatin distally along the male reproductive tract.

Additional keywords: Fst288, Fst315, real-time polymerase chain reaction (PCR).


References

Anawalt, B. D., Bebb, R. A., Matsumoto, A. M., Groome, N. P., Illingworth, P. J., McNeilly, A. S., and Bremner, W. J. (1996). Serum inhibin B levels reflect Sertoli cell function in normal men and men with testicular dysfunction. J. Clin. Endocrinol. Metab. 81, 3341–3345.
Serum inhibin B levels reflect Sertoli cell function in normal men and men with testicular dysfunction.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DyaK28Xls1OrtLw%3D&md5=afef3e29ce44cd5e1e74454b286eb44dCAS | 8784094PubMed |

Anderson, R. A., Evans, L. W., Irvine, D. S., McIntyre, M. A., Groome, N. P., and Riley, S. C. (1998). Follistatin and activin A production by the male reproductive tract. Hum. Reprod. 13, 3319–3325.
Follistatin and activin A production by the male reproductive tract.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DyaK1MXltFOmtQ%3D%3D&md5=6502037f44baa916e6989ce4cb9545a3CAS | 9886507PubMed |

Bahathiq, A. O., Stewart, R. L., Baxter, L., Wells, M., Moore, H. D., and Ledger, W. L. (2005). Tissue immunoexpression and messenger ribonucleic acid localization of inhibin/activin subunit in human epididymis. Fertil. Steril. 83, 78–85.
Tissue immunoexpression and messenger ribonucleic acid localization of inhibin/activin subunit in human epididymis.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD2MXisVOqsrc%3D&md5=6e05fd24fa5e53a2be2e7edd97825012CAS | 15652891PubMed |

Chang, H., Lau, A. L., and Matzuk, M. M. (2001). Studying TGF-beta superfamily signaling by knockouts and knockins. Mol. Cell. Endocrinol. 180, 39–46.
Studying TGF-beta superfamily signaling by knockouts and knockins.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD3MXltVarurc%3D&md5=f0953f5b596f2cb3b324c0e8c378d80eCAS | 11451570PubMed |

Craythorn, R. G., Girling, J. E., Hedger, M. P., Rogers, P. A., and Winnall, W. R. (2009). An RNA spiking method demonstrates that 18S rRNA is regulated by progesterone in the mouse uterus. Mol. Hum. Reprod. 15, 757–761.
An RNA spiking method demonstrates that 18S rRNA is regulated by progesterone in the mouse uterus.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD1MXht1Ortr%2FJ&md5=01e35a4bb1780ad12f2f6e32f123fb3fCAS | 19602508PubMed |

Da Silva, N., Cortez-Retamozo, V., Reinecker, H. C., Wildgruber, M., Hill, E., Brown, D., Swirski, F. K., Pittet, M. J., and Breton, S. (2011). A dense network of dendritic cells populates the murine epididymis. Reproduction 141, 653–663.
A dense network of dendritic cells populates the murine epididymis.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BC3MXmvFCisrk%3D&md5=821d60ad2acfe0fd5f053c08c12d5c94CAS | 21310816PubMed |

de Kretser, D. M., Buzzard, J. J., Okuma, Y., O’Connor, A. E., Hayashi, T., Lin, S. Y., Morrison, J. R., Loveland, K. L., and Hedger, M. P. (2004). The role of activin, follistatin and inhibin in testicular physiology. Mol. Cell. Endocrinol. 225, 57–64.
The role of activin, follistatin and inhibin in testicular physiology.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD2cXnvV2mtbg%3D&md5=aaa0cfea8b6189b119e309654cdd756cCAS | 15451568PubMed |

Eichberger, T., Kaser, A., Pixner, C., Schmid, C., Klingler, S., Winklmayr, M., Hauser-Kronberger, C., Aberger, F., and Frischauf, A. M. (2008). GLI2-specific transcriptional activation of the bone morphogenetic protein/activin antagonist follistatin in human epidermal cells. J. Biol. Chem. 283, 12 426–12 437.
GLI2-specific transcriptional activation of the bone morphogenetic protein/activin antagonist follistatin in human epidermal cells.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD1cXltVyqs74%3D&md5=945cd274e2a813d47b6d518d53e04eb7CAS |

Esch, F. S., Shimasaki, S., Mercado, M., Cooksey, K., Ling, N., Ying, S., Ueno, N., and Guillemin, R. (1987). Structural characterization of follistatin: a novel follicle-stimulating hormone release-inhibiting polypeptide from the gonad. Mol. Endocrinol. 1, 849–855.
Structural characterization of follistatin: a novel follicle-stimulating hormone release-inhibiting polypeptide from the gonad.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DyaL1MXhs1Kht7k%3D&md5=84a783d2e7e8f26e52ae471b1fdebaf7CAS | 3153465PubMed |

Feng, Z. M., Madigan, M. B., and Chen, C. L. (1993). Expression of type II activin receptor genes in the male and female reproductive tissues of the rat. Endocrinology 132, 2593–2600.
Expression of type II activin receptor genes in the male and female reproductive tissues of the rat.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DyaK3sXltlWrsLo%3D&md5=b1905b102154224a961e651f27f12401CAS | 7916681PubMed |

Graham, H., and Peng, C. (2006). Activin receptor-like kinases: structure, function and clinical implications. Endocr. Metab. Immune Disord. Drug Targets 6, 45–58.
| 1:CAS:528:DC%2BD28XisVWkurs%3D&md5=ac126b2147e27ee5f47a67cc75a31bd8CAS | 16611164PubMed |

Hakovirta, H., Kaipia, A., Soder, O., and Parvinen, M. (1993). Effects of activin-A, inhibin-A, and transforming growth factor-beta 1 on stage-specific deoxyribonucleic acid synthesis during rat seminiferous epithelial cycle. Endocrinology 133, 1664–1668.
Effects of activin-A, inhibin-A, and transforming growth factor-beta 1 on stage-specific deoxyribonucleic acid synthesis during rat seminiferous epithelial cycle.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DyaK2cXotFE%3D&md5=25e501a19155943dddfc99d3f7bf3b33CAS | 8404607PubMed |

Hedger, M. P. (2011). Immunophysiology and pathology of inflammation in the testis and epididymis. J. Androl. 32, 625–640.
Immunophysiology and pathology of inflammation in the testis and epididymis.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BC3MXhsVCru73O&md5=c353d004cffbb103189e06b4d0683ecdCAS | 21764900PubMed |

Hedger, M. P., Winnall, W. R., Phillips, D. J., and de Kretser, D. M. (2011). The regulation and functions of activin and follistatin in inflammation and immunity. Vitam. Horm. 85, 255–297.
The regulation and functions of activin and follistatin in inflammation and immunity.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BC3MXovVCit7g%3D&md5=6270c49f568bf37b219f58a8a1bd4943CAS | 21353885PubMed |

Hübner, G., Hu, Q., Smola, H., and Werner, S. (1996). Strong induction of activin expression after injury suggests an important role of activin in wound repair. Dev. Biol. 173, 490–498.
Strong induction of activin expression after injury suggests an important role of activin in wound repair.Crossref | GoogleScholarGoogle Scholar | 8606007PubMed |

Knight, P. G., Muttukrishna, S., and Groome, N. P. (1996). Development and application of a two-site enzyme immunoassay for the determination of ‘total’ activin-A concentrations in serum and follicular fluid. J. Endocrinol. 148, 267–279.
| 1:CAS:528:DyaK28XhtVCrsrg%3D&md5=f46623bd2d9918290ca514da68998686CAS | 8699141PubMed |

Lerch, T. F., Shimasaki, S., Woodruff, T. K., and Jardetzky, T. S. (2007). Structural and biophysical coupling of heparin and activin binding to follistatin isoform functions. J. Biol. Chem. 282, 15 930–15 939.
Structural and biophysical coupling of heparin and activin binding to follistatin isoform functions.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD2sXltl2nsLc%3D&md5=8ff941159c771bf244c46f7bb26d5c50CAS |

Livak, K. J., and Schmittgen, T. D. (2001). Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)) method. Methods 25, 402–408.
Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)) method.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD38XhtFelt7s%3D&md5=c02316d2aa02af4c464251269a77c9c9CAS | 11846609PubMed |

Mather, J. P., Attie, K. M., Woodruff, T. K., Rice, G. C., and Phillips, D. M. (1990). Activin stimulates spermatogonial proliferation in germ-Sertoli cell cocultures from immature rat testis. Endocrinology 127, 3206–3214.
Activin stimulates spermatogonial proliferation in germ-Sertoli cell cocultures from immature rat testis.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DyaK3MXlsF2gsA%3D%3D&md5=28f27e81cc83e8808dbddb444cd5f135CAS | 2249646PubMed |

Mathews, L. S., and Vale, W. W. (1991). Expression cloning of an activin receptor, a predicted transmembrane serine kinase. Cell 65, 973–982.
Expression cloning of an activin receptor, a predicted transmembrane serine kinase.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DyaK38XjsFCktw%3D%3D&md5=d016575cff4ff1686603ae61230fbd7bCAS | 1646080PubMed |

Matzuk, M. M., Kumar, T. R., and Bradley, A. (1995a). Different phenotypes for mice deficient in either activins or activin receptor type II. Nature 374, 356–360.
Different phenotypes for mice deficient in either activins or activin receptor type II.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DyaK2MXksFKnu7Y%3D&md5=f0609bb4c327f5a715e3aa33b3e36fbfCAS | 7885474PubMed |

Matzuk, M. M., Lu, N., Vogel, H., Sellheyer, K., Roop, D. R., and Bradley, A. (1995b). Multiple defects and perinatal death in mice deficient in follistatin. Nature 374, 360–363.
Multiple defects and perinatal death in mice deficient in follistatin.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DyaK2MXksFKnu7c%3D&md5=0ac6a19e2912e4990039b0e0e59c4707CAS | 7885475PubMed |

Mendis, S. H., Meachem, S. J., Sarraj, M. A., and Loveland, K. L. (2011). Activin A balances Sertoli and germ cell proliferation in the fetal mouse testis. Biol. Reprod. 84, 379–391.
Activin A balances Sertoli and germ cell proliferation in the fetal mouse testis.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BC3MXhsVelt7w%3D&md5=8b37a75a4d3c99446bb566e7ed443f1dCAS | 20926807PubMed |

Michel, U., Albiston, A., and Findlay, J. K. (1990). Rat follistatin: gonadal and extragonadal expression and evidence for alternative splicing. Biochem. Biophys. Res. Commun. 173, 401–407.
Rat follistatin: gonadal and extragonadal expression and evidence for alternative splicing.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DyaK3MXitFequw%3D%3D&md5=1c4d6ccb7809c27786616e437f52268cCAS | 1979488PubMed |

Nakamura, T., Takio, K., Eto, Y., Shibai, H., Titani, K., and Sugino, H. (1990). Activin-binding protein from rat ovary is follistatin. Science 247, 836–838.
Activin-binding protein from rat ovary is follistatin.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DyaK3cXhsFajsbg%3D&md5=9b57960804c672f6da318e369245eed7CAS | 2106159PubMed |

Nistal, M., Gonzalez-Peramato, P., and De Miguel, M. P. (2010). Immunodetection of inhibin in the human testis and epididymis during normal development and in non-tumoural testicular lesions. Reprod. Fertil. Dev. 22, 558–563.
Immunodetection of inhibin in the human testis and epididymis during normal development and in non-tumoural testicular lesions.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BC3cXisFOgsb4%3D&md5=33cb55e6b7b38bc6dd9b59414c743e34CAS | 20188029PubMed |

O’Connor, A. E., McFarlane, J. R., Hayward, S., Yohkaichiya, T., Groome, N. P., and de Kretser, D. M. (1999). Serum activin A and follistatin concentrations during human pregnancy: a cross-sectional and longitudinal study. Hum. Reprod. 14, 827–832.
Serum activin A and follistatin concentrations during human pregnancy: a cross-sectional and longitudinal study.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DyaK1MXisV2qtr0%3D&md5=29da6964045d0781c65736c56cb0f9cfCAS | 10221721PubMed |

Okuma, Y., O’Connor, A. E., Hayashi, T., Loveland, K. L., de Kretser, D. M., and Hedger, M. P. (2006). Regulated production of activin A and inhibin B throughout the cycle of the seminiferous epithelium in the rat. J. Endocrinol. 190, 331–340.
Regulated production of activin A and inhibin B throughout the cycle of the seminiferous epithelium in the rat.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD28Xpt1WjtLw%3D&md5=c5bfd695381cabc0339d488ee738ab94CAS | 16899566PubMed |

Robertson, D. M., Hayward, S., Irby, D., Jacobsen, J., Clarke, L., McLachlan, R. I., and de Kretser, D. M. (1988). Radioimmunoassay of rat serum inhibin: changes after PMSG stimulation and gonadectomy. Mol. Cell. Endocrinol. 58, 1–8.
Radioimmunoassay of rat serum inhibin: changes after PMSG stimulation and gonadectomy.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DyaL1cXkvVKns7k%3D&md5=0a56df7f543ff49346176247276afdf4CAS | 3208985PubMed |

Schneider, O., Nau, R., and Michel, U. (2000). Comparative analysis of follistatin-, activin beta A- and activin beta B-mRNA steady-state levels in diverse porcine tissues by multiplex S1 nuclease analysis. Eur. J. Endocrinol. 142, 537–544.
Comparative analysis of follistatin-, activin beta A- and activin beta B-mRNA steady-state levels in diverse porcine tissues by multiplex S1 nuclease analysis.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD3cXjslels78%3D&md5=9bd8e5e61293d5c630b9c2bde91e9e09CAS | 10802535PubMed |

Schneyer, A., Sidis, Y., Xia, Y., Saito, S., del Re, E., Lin, H. Y., and Keutmann, H. (2004). Differential actions of follistatin and follistatin-like 3. Mol. Cell. Endocrinol. 225, 25–28.
Differential actions of follistatin and follistatin-like 3.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD2cXnvV2mtbw%3D&md5=49e31b65d4cb4a39756a73d9c4810285CAS | 15451564PubMed |

Shimasaki, S., Koga, M., Esch, F., Cooksey, K., Mercado, M., Koba, A., Ueno, N., Ying, S. Y., Ling, N., and Guillemin, R. (1988). Primary structure of the human follistatin precursor and its genomic organization. Proc. Natl Acad. Sci. USA 85, 4218–4222.
Primary structure of the human follistatin precursor and its genomic organization.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DyaL1MXhvFajsQ%3D%3D&md5=1a475c8a7bcc0210cbb362d61cf6d7cfCAS | 3380788PubMed |

Sugino, K., Kurosawa, N., Nakamura, T., Takio, K., Shimasaki, S., Ling, N., Titani, K., and Sugino, H. (1993). Molecular heterogeneity of follistatin, an activin-binding protein. Higher affinity of the carboxyl-terminal truncated forms for heparan sulfate proteoglycans on the ovarian granulosa cell. J. Biol. Chem. 268, 15 579–15 587.
| 1:CAS:528:DyaK3sXmtFOmu7w%3D&md5=b5c864f2b8ac936b5c33637a30c0abafCAS |

Thomas, T. Z., Chapman, S. M., Hong, W., Gurusingfhe, C., Mellor, S. L., Fletcher, R., Pedersen, J., and Risbridger, G. P. (1998). Inhibins, activins, and follistatins: expression of mRNAs and cellular localization in tissues from men with benign prostatic hyperplasia. Prostate 34, 34–43.
Inhibins, activins, and follistatins: expression of mRNAs and cellular localization in tissues from men with benign prostatic hyperplasia.Crossref | GoogleScholarGoogle Scholar | 1:STN:280:DyaK1c%2FovF2rtQ%3D%3D&md5=36735fb36d18a8426fc711fd14f2c222CAS | 9428386PubMed |

Tomaszewski, J., Joseph, A., Archambeault, D., and Yao, H. H. (2007). Essential roles of inhibin beta A in mouse epididymal coiling. Proc. Natl Acad. Sci. USA 104, 11 322–11 327.
Essential roles of inhibin beta A in mouse epididymal coiling.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD2sXnvFOmsr4%3D&md5=3e2e9cad3c1567ce81d75ec4e55206b3CAS |

Tortoriello, D. V., Sidis, Y., Holtzman, D. A., Holmes, W. E., and Schneyer, A. L. (2001). Human follistatin-related protein: a structural homologue of follistatin with nuclear localization. Endocrinology 142, 3426–3434.
Human follistatin-related protein: a structural homologue of follistatin with nuclear localization.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD3MXls1Sqsbo%3D&md5=b3773e6c104207dd952a8b88bc4a9e43CAS | 11459787PubMed |

Tsuchida, K., Arai, K. Y., Kuramoto, Y., Yamakawa, N., Hasegawa, Y., and Sugino, H. (2000). Identification and characterization of a novel follistatin-like protein as a binding protein for the TGF-beta family. J. Biol. Chem. 275, 40 788–40 796.
Identification and characterization of a novel follistatin-like protein as a binding protein for the TGF-beta family.Crossref | GoogleScholarGoogle Scholar | 1:STN:280:DC%2BD3M7gtlamsQ%3D%3D&md5=6b9d20da79a2c5df15de03e7663be472CAS |

Veeramachaneni, D. N., Schanbacher, B. D., and Amann, R. P. (1989). Immunolocalization and concentrations of inhibin alpha in the ovine testis and excurrent duct system. Biol. Reprod. 41, 499–503.
Immunolocalization and concentrations of inhibin alpha in the ovine testis and excurrent duct system.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DyaK3cXpt1U%3D&md5=1cec795f2296fd2015fbc387d10947e8CAS | 2590716PubMed |

Winnall, W. R., Muir, J. A., Liew, S., Hirst, J. J., Meachem, S. J., and Hedger, M. P. (2009). Effects of chronic celecoxib on testicular function in normal and lipopolysaccharide-treated rats. Int. J. Androl. 32, 542–555.
| 1:CAS:528:DC%2BD1MXht1OqsbbP&md5=72df7299eaf5c18c34320bd037f2b91eCAS | 18522674PubMed |

Zhang, T., Guo, C. X., Hu, Z. Y., and Liu, Y. X. (1997). Localization of plasminogen activator and inhibitor, LH and androgen receptors and inhibin subunits in monkey epididymis. Mol. Hum. Reprod. 3, 945–952.
Localization of plasminogen activator and inhibitor, LH and androgen receptors and inhibin subunits in monkey epididymis.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DyaK1cXktlWmsg%3D%3D&md5=b17509d8cdd27a6c149a1287d2bf973cCAS | 9433919PubMed |