Animal Bioscience

Asian Australasian Association of Animal Production Societies (아세아태평양축산학회)
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Functional characterization of naturally-occurring constitutively activating/inactivating mutations in equine follicle-stimulating hormone receptor

  • Byambaragchaa, Munkhzaya (Animal Biotechnology, Graduate School of Future Convergence Technology, Hankyong National University) ;
  • Ahn, Tae-Young (Animal Biotechnology, Graduate School of Future Convergence Technology, Hankyong National University) ;
  • Choi, Seung-Hee (Animal Biotechnology, Graduate School of Future Convergence Technology, Hankyong National University) ;
  • Kang, Myung-Hwa (Department of Food Science and Nutrition, Hoseo University) ;
  • Min, Kwan-Sik (Animal Biotechnology, Graduate School of Future Convergence Technology, Hankyong National University)
  • Received : 2021.05.27
  • Accepted : 2021.07.12
  • Published : 2022.03.01
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Abstract

Objective: Follicle-stimulating hormone (FSH) is the central hormone involved in mammalian reproduction, maturation at puberty, and gamete production that mediates its function by control of follicle growth and function. The present study investigated the mutations involved in the regulation of FSH receptor (FSHR) activation. Methods: We analyzed seven naturally-occurring mutations that were previously reported in human FSHR (hFSHR), in the context of equine FSHR (eFSHR); these include one constitutively activation variant, one allelic variant, and five inactivating variants. These mutations were introduced into wild-type eFSHR (eFSHR-wt) sequence to generate mutants that were designated as eFSHR-D566G, -A306T, -A189V, -N191I, -R572C, -A574V, and -R633H. Mutants were transfected into PathHunter EA-parental CHO-K1 cells expressing β-arrestin. The biological function of mutants was analyzed by quantitating cAMP accumulation in cells incubated with increasing concentrations of FSH. Results: Cells expressing eFSHR-D566G exhibited an 8.6-fold increase in basal cAMP response, as compared to that in eFSHR-wt. The allelic variation mutant eFSHR-A306T was not found to affect the basal cAMP response or half maximal effective concentration (EC50) levels. On the other hand, eFSHR-D566G and eFSHR-A306T displayed a 1.5- and 1.4-fold increase in the maximal response, respectively. Signal transduction was found to be completely impaired in case of the inactivating mutants eFSHR-A189V, -R572C, and -A574V. When compared with eFSHR-wt, eFSHR-N191I displayed a 5.4-fold decrease in the EC50 levels (3,910 ng/mL) and a 2.3-fold decrease in the maximal response. In contrast, cells expressing eFSHR-R633H displayed in a similar manner to that of the cells expressing the eFSHR-wt on signal transduction and maximal response. Conclusion: The activating mutant eFSHR-D566G greatly enhanced the signal transduction in response to FSH, in the absence of agonist treatment. We suggest that the state of activation of the eFSHR can modulate its basal cAMP accumulation.

Keywords

References

  1. Simoni M, Gromoll J, Hoppner W, et al. Mutational analysis of the follicle-stimulating hormone (FSH) receptor in normal and infertile men: identification and characterization of two discrete FSH receptor isoforms. J Clin Endocrinol Metab 1999;84:751-5. https://doi.org/10.1210/jcem.84.2.5500
  2. Min KS, Liu X, Fabritz J, Jaquette J, Abell AN, Ascoli M. Mutations that induce constitutive activations and mutations that impair signal transduction modulate the basal and/or agonist-stimulated internalization of the lutropin/choriogonadotropin receptor. J Biol Chem 1998;273:34911-9. https://doi.org/10.1074/jbc.273.52.34911
  3. Krishnamurthy H, Kish H, Shi M, et al. Postendocytotic trafficking of the follicle-stimulating hormone (FSH)-FSH receptor complex. Mol Endocrinol 2003;17:2162-76. https://doi.org/10.1210/me.2003-0118
  4. Bhaskaran RS, Ascoli M. The post-endocytotic fate of the gonadotropin receptors is an important determinant of the desensitization of gonadotropin responses. J Mol Endocrinol 2005;34:447-57. https://doi.org/10.1677/jme.1.01745
  5. Banerjee AA, Mahale SD. Role of the extracellular and intracellular loops of follicle-stimulating hormone receptor in its function. Front Endocrinol 2015;6:110. https://doi.org/10.3389/fendo.2015.00110
  6. Aittomaki K, Dieguez-Lucena JL, Pakarinen P, et al. Mutation in the follicle-stimulating hormone receptor gene causes hereditary hypergonadotropic ovarian failure. Cell 1995;82:956-68. https://doi.org/10.1016/0092-8674(95)90275-9
  7. Gromoll J, Simoni M, Nieschlag E. An activating mutation of the follicle-stimulating hormone receptor autonomously sustains spermatogenesis in a hypophysectomized man. J Clin Endocrinol Metab 1996;84:1367-70. https://doi.org/10.1210/jcem.81.4.8636335
  8. Gromoll J, Simoni M, Nordhoff V, Behre HM, De Geyter C, Nieschlag E. Functional and clinical consequences of mutations in the FSH receptor. Mol Cell Endocrinol 1996;125: 177-82. https://doi.org/10.1016/s0303-7207(96)03949-4
  9. Tao YX, Abell AN, Liu X, Nakamura K, Segaloff DL. Constitutive activation of G protein-coupled receptors as a result of selective substitution of a conserved leucine residue in transmembrane helix III. Mol Endocrinol 2000;14:1272-82. https://doi.org/10.1210/mend.14.8.0503
  10. Tao YX, Mizrachi D, Segaloff DL. Chimeras of the rat and human FSH receptors (FSHRs) identify residues that permit or suppress transmembrane 6 mutation-induced constitutive activation of the FSHR via rearrangements of hydrophobic interactions between helices 6 and 7. Mol Endocrinol 2002;16:1881-92. https://doi.org/10.1210/me.2001-0199
  11. Zhang M, Tao YX, Ryan GL, Feng X, Fanelli F, Segaloff DL. Intrinsic differences in the response of the human lutropin receptor versus the human follitropin receptor to activating mutations. J Biol Chem 2007;282:25527-39. https://doi.org/10.1074/jbc.M703500200
  12. Ghezelayagh Z, Totonchi M, Zarei-Moradi S, et al. The impact of genetic variation and gene expression level of the follicle-stimulating hormone receptor on ovarian reserve. Cell J 2018; 19:620-6. https://doi.org/10.22074/cellj.2018.4183
  13. Simoni M, Gromoll J, Nieschlag E. The follicle-stimulating hormone receptor: biochemistry, molecular biology, physiology, and pathophysiology. Endocr Rev 1997;18:739-73. https://doi.org/10.1210/edrv.18.6.0320
  14. Kleinau G, Krause G. Thyrotropin and homologous glycoprotein hormone receptors: structural and functional aspects of extracellular signaling mechanisms. Endocr Rev 2009;30:133-51. https://doi.org/10.1210/er.2008-0044
  15. Laue L, Chan WY, Hsueh AJ, et al. Genetic heterogeneity of constitutively activating mutations of the human luteinizing hormone receptor in familial male-limited precocious puberty. Proc Natl Acad Sci USA 1995;92:1906-10. https://doi.org/10.1073/pnas.92.6.1906
  16. Kraaij R, Post M, Kremer H, et al. A missense mutation in the second transmembrane segment of the luteinizing hormone receptor causes familial male-limited precocious puberty. J Clin Endcrinol Metab 1995;80:3168-72. https://doi.org/10.1210/jcem.80.11.7593421
  17. Whitney EA, Layman LC, Chan PJ, Lee A, Peak DB, McDonough PG. The follicle-stimulating hormone receptor gene is polymorphic in premature ovarian failure and normal controls. Fertil Steril 1995;64:518-24. https://doi.org/10.1016/s0015-0282(16)57786-3
  18. Fuller PJ, Verity K, Shen Y, Mamers P, Jobling T, Burger HG. No evidence of a role for mutations or polymorphisms of the follicle-stimulating hormone receptor in ovarian granulosa cell tumors. J Clin Endocrinol Metab 1998;83:274-9. https://doi.org/10.1210/jcem.83.1.4509
  19. Bas F, Pescovitz OH, Steinmetz R. No activating mutations of FSH receptor in four children with ovarian juvenile granulosa cell tumors and the association of these tumors with central precocious puberty. J Pediatr Adolesc Gynecol 2009;22:173-9. https://doi.org/10.1016/j.jpag.2008.10.003
  20. Aittomaki K, Herva R, Stenman UH, et al. Clinical features of primary ovarian failure caused by a point mutation in the follicle-stimulating hormone receptor gene. J Clin Endocrinol Metab 1996;81:3722-6. https://doi.org/10.1210/jcem.81.10.8855829
  21. Tapanainen JS, Aittomaki K, Min J, Vaskivuo T, Huhtaniemi IT. Men homozygous for an inactivating mutation of the follicle-stimulating hormone (FSH) receptor gene present variable suppression of spermatogenesis and fertility. Nat Genet 1997;15:205-6. https://doi.org/10.1038/ng0297-205
  22. Liu X, Davis D, Segaloff DL. Disruption of potential sites for N-linked glycosylation does not impair hormone binding to the lutropin/choriogonadotropin receptor if Asn-173 is left intact. J Biol Chem 1993;268:1513-6. https://doi.org/10.1016/S0021-9258(18)53881-3
  23. Davis D, Liu X, Segaloff DL. Identification of the sites of N-linked glycosylation on the follicle-stimulating hormone (FSH) receptor and assessment of their role in FSH receptor function. Mol Endocrinol 1995;9:159-70. https://doi.org/10.1210/mend.9.2.7776966
  24. Beau I, Touraine P, Meduri G, et al. A novel phenotype related to partial loss of function mutations of the follicle stimulating hormone receptor. J Clin Invest 1998;102:1352-9. https://doi.org/10.1172/JCI3795
  25. Desai SS, Achrekar SK, Sahasrabuddhe KA, et al. Functional characterization of two naturally occurring mutations (Val514Ala and Ala575Val) in follicle-stimulating hormone receptor. J Clin Endocrinol Metab 2015;100:E635-45. https://doi.org/10.1210/jc.2014-3662
  26. Touraine P, Beau I, Gougeon A, et al. New natural inactivating mutations of the follicle-stimulating hormone receptor: correlations between receptor function and phenotype. Mol Endocrinol 1999;13:1844-54. https://doi.org/10.1210/mend.13.11.0370
  27. Hugon-Rodin J, Sonigo C, Gompel A, et al. First mutation in the FSHR cytoplasmic tail identified in a non-pregnant woman with spontaneous ovarian hyperstimulation syndrome. BMC Med Genet 2017;18:44. https://doi.org/10.1186/s12881-017-0407-6
  28. Desai SS, Achrekar SK, Pathak BR, et al. Follicle-stimulating hormone receptor polymorphism (G-29A) is associated with altered level of receptor expression in granulosa cells. J Clin Endocrinol Metab 2011;96:2805-12. https://doi.org/10.1210/jc.2011-1064
  29. Katari S, Wood-Trageser MA, Jiang H, et al. Novel inactivating mutation of the FSH receptor in two siblings of Indian origin with premature ovarian failure. J Clin Endocrinol Metab 2015;100:2154-7. https://doi.org/10.1210/jc.2015-1401
  30. Bramble MS, Goldstein EH, Lipson A, et al. A novel follicle-stimulating hormone receptor mutation causing primary ovarian failure: a fertility application of whole exome sequencing. Hum Reprod 2016;31:905-14. https://doi.org/10.1093/humrep/dew025
  31. Liu H, Xu X, Han T, et al. A novel homozygous mutation in the FSHR gene is causative for primary ovarian insufficiency. Fertil Steril 2017;108:1050-5. https://doi.org/10.1016/j.fertnstert.2017.09.010
  32. Byambaragchaa M, Kim JS, Park HK, et al. Constitutive activation and inactivation of mutations inducing cell surface loss of receptor and impairing of signal transduction of agonist-stimulated eel follicle-stimulating hormone receptor. Int J Mol Sci 2020;21:7075. https://doi.org/10.3390/ijms21197075
  33. Byambaragchaa M, Kim DJ, Kang MH, Min KS. Site specificity of eel luteinizing hormone N-linked oligosaccharides in signal transduction. Gen Comp Endocrinol 2018;268:50-6. https://doi.org/10.1016/j.ygcen.2018.07.015
  34. Morkrosiryski J, Mrimurer TM, Silvertsen B, Schwartz TW, Holst B. Modulation of constitutive activity and signaling bias of the ghrelin receptor by conformational constraint in the second extracellular loop. J Biol Chem 2012;287:33488-502. https://doi.org/10.1074/jbc.M112.383240
  35. Kudo M, Osuga Y, Kobilka BK, Hsueh AJW. Transmembrane regions V and VI of the human luteinizing hormone receptor are required for constitutive activation by a mutation in the third intracellular loop. J Biol Chem 1996;271:22470-8. https://doi.org/10.1074/jbc.271.37.22470
  36. Bradbury FA, Kawate N, Foster CM, Menon KMJ. Posttranslational processing in the Golgi plays a critical role in the trafficking of the luteinizing hormone/human chorionic gonadotropin receptor to the cell surface. J Biol Chem 1997;272:5921-6. https://doi.org/10.1074/jbc.272.9.5921
  37. Nakamura K, Krupnick JG, Benovic JL, Ascoli M. Signaling and phosphorylation-impaired mutants of the rat follitropin receptor reveal an activation- and phosphorylation-independent but arrestin-dependent pathway for internalization. J Biol Chem 1998;273:24346-54. https://doi.org/10.1074/jbc.273.38.24346
  38. Schulz A, Schoneberg T, Paschke R, Schultz G, Gudermann T. Role of the third intracellular loop for the activation of gonadotropin receptors. Mol Endocrinol 1999;13:181-90. https://doi.org/10.1210/mend.13.2.0233
  39. Haywood M, Tymchenko N, Spaliviero J, et al. An activated human follicle-stimulating hormone (FSH) receptor stimulates FSH-like activity in gonadotropin-deficient transgenic mice. Mol Endocrinol 2002;16:2582-91. https://doi.org/10.1210/me.2002-0032
  40. Nordhoff V, Gromall J, Foppiani L, et al. Targeted expression of human FSH receptor Asp567Gly mutant mRNA in testis of transgenic mice: role of the human FSH receptor promoter. Asian J Androl 2003;5:267-75.
  41. Allan CM, Lim P, Robson M, Spaliviero J, Handelsman DJ. Transgenic mutant D567G but not wild-type human FSH receptor overexpression provides FSH-independent and promiscuous glycoprotein hormone Sertoli cell signaling. Am J Physiol Endocrinol Metab 2009;296:E1022-8. https://doi.org/10.1152/ajpendo.90941.2008
  42. Loutradis D, Patsoula E, Minas V, et al. FSH receptor gene polymorphisms have a role for different ovarian response to stimulation in patients entering IVF/ICSI-ET programs. J Assist Reprod Genet 2006;23:177-84. https://doi.org/10.1007/s10815-005-9015-z
  43. De Koning CH, Benjamins T, Harms P, et al. The distribution of FSH receptor isoforms is related to basal FSH levels in subfertile women with normal menstrual cycles. Hum Reprod 2006;21:443-6. https://doi.org/10.1093/humrep/dei317
  44. Du J, Zhang W, Guo L, et al. Two FSHR variants, haplotypes and meta-analysis in Chinese women with premature ovarian failure and polycystic ovary syndrome. Mol Genet Metab 2010;100:292-5. https://doi.org/10.1016/j.ymgme.2010.03.018
  45. Yarney TA, Sairam MR, Khan H, Ravindranath N, Payne S, Seidah NG. Molecular cloning and expression of the ovine testicular follicle stimulating hormone receptor. Mol Cell Endocrinol 1993;93:219-26. https://doi.org/10.1016/0303-7207(93)90127-6
  46. Houde A, Lambert A, Saumande J, Silversides DW, Lussier JG, Saumande J. Structure of the bovine follicle-stimulating hormone receptor complementary DNA and expression in bovine tissues. Mol Reprod Dev 1994;39:127-35. https://doi.org/10.1002/mrd.1080390202
  47. Robert P, Amsellem S, Christophe S, et al. Cloning and sequencing of the equine testicular follitropin receptor. Biochem Biophys Res Commun 1994;201:201-7. https://doi.org/10.1006/bbrc.1994.1689
  48. Rannikko A, Pakainen P, Manna PR, et al. Functional characterization of the human FSH receptor with an inactivating Ala189Val mutation. Mol Hum Reprod 2002;8:311-7. https://doi.org/10.1093/molehr/8.4.311
  49. Tao YX. Inactivating mutations of G protein-coupled receptors and diseases: structure-function insights and therapeutic implications. Pharmacol Ther 2006;111:949-73. https://doi.org/10.1016/j.pharmthera.2006.02.008