The Urokinase-Type Plasminogen Activator Contributes to cAMP-Induced Steroidogenesis in MA-10 Leydig Cells

Leydig cells produce androgens which are essential for male sex differentiation and reproductive functions. Steroidogenesis, as well as expression of several genes in Leydig cells, are stimulated by LH/cAMP and repressed by AMP/AMPK. One of those genes is Plau, which codes for the urokinase-type plasminogen activator (uPA), a secreted serine protease. The role of uPA and the regulation of Plau expression in Leydig cells remain unknown. Using siRNA-mediated knockdown, uPA was required for maximal cAMP-induced STAR and steroid hormone production in MA-10 Leydig cells. Analysis of Plau mRNA levels and promoter activity revealed that its expression is strongly induced by cAMP; this induction is blunted by AMPK. The cAMP-responsive region was located, in part, in the proximal Plau promoter that contains a species-conserved GC box at −56 bp. The transcription factor Krüppel-like factor 6 (KLF6) activated the Plau promoter. Mutation of the GC box at −56 bp abolished KLF6-mediated activation and significantly reduced cAMP-induced Plau promoter activity. These data define a role for uPA in Leydig cell steroidogenesis and provide insights into the regulation of Plau gene expression in these cells.

[1]  Daniela Alfano,et al.  Modulation of Cellular Function by the Urokinase Receptor Signalling: A Mechanistic View , 2022, Frontiers in Cell and Developmental Biology.

[2]  J. Tremblay,et al.  Transcription Factors in the Regulation of Leydig Cell Gene Expression and Function , 2022, Frontiers in Endocrinology.

[3]  J. Tremblay,et al.  Mechanism of Action of an Environmentally Relevant Organochlorine Mixture in Repressing Steroid Hormone Biosynthesis in Leydig Cells , 2022, International journal of molecular sciences.

[4]  F. Bergeron,et al.  Growth hormone-induced STAT5B regulates Star gene expression through a cooperation with cJUN in mouse MA-10 Leydig cells. , 2021, Endocrinology.

[5]  J. Tremblay,et al.  Mechanisms of MEHP Inhibitory Action and Analysis of Potential Replacement Plasticizers on Leydig Cell Steroidogenesis , 2021, International journal of molecular sciences.

[6]  J. Tremblay,et al.  Identification of novel genes and pathways regulated by the orphan nuclear receptor COUP-TFII in mouse MA-10 Leydig cells , 2021, Biology of Reproduction.

[7]  J. Tremblay,et al.  The Nuclear Receptor COUP-TFII Regulates Amhr2 Gene Transcription via a GC-Rich Promoter Element in Mouse Leydig Cells , 2019, Journal of the Endocrine Society.

[8]  P. Tajik,et al.  Effect of extracellular matrix on testosterone production during in vitro culture of bovine testicular cells , 2017, Veterinary research forum : an international quarterly journal.

[9]  J. Tremblay.  Molecular regulation of steroidogenesis in endocrine Leydig cells , 2015, Steroids.

[10]  M. Di-Luoffo,et al.  MEF2 Cooperates With Forskolin/cAMP and GATA4 to Regulate Star Gene Expression in Mouse MA-10 Leydig Cells. , 2015, Endocrinology.

[11]  J. Tremblay,et al.  The Nuclear Receptor NR2F2 Activates Star Expression and Steroidogenesis in Mouse MA-10 and MLTC-1 Leydig Cells1 , 2014, Biology of reproduction.

[12]  J. Tremblay,et al.  In Vitro Exposure of Leydig Cells to an Environmentally Relevant Mixture of Organochlorines Represses Early Steps of Steroidogenesis1 , 2014, Biology of reproduction.

[13]  M. Ascoli,et al.  The ERK1/2 pathway regulates testosterone synthesis by coordinately regulating the expression of steroidogenic genes in Leydig cells , 2013, Molecular and Cellular Endocrinology.

[14]  J. Tremblay,et al.  The calcium signaling pathway regulates leydig cell steroidogenesis through a transcriptional cascade involving the nuclear receptor NR4A1 and the steroidogenic acute regulatory protein. , 2013, Endocrinology.

[15]  G. Ricci,et al.  Hepatocyte Growth Factor Is a Mouse Fetal Leydig Cell Terminal Differentiation Factor1 , 2012, Biology of reproduction.

[16]  J. Beavo,et al.  cAMP-Specific Phosphodiesterases 8A and 8B, Essential Regulators of Leydig Cell Steroidogenesis , 2012, Molecular Pharmacology.

[17]  I. Politis,et al.  Cloning of the 5' regulatory regions and functional characterization of the core promoters of ovine PLAU (u-PA) and SERPIN1 (PAI-1). , 2011, Gene.

[18]  S. Vandenberg,et al.  Crosstalk between the urokinase-type plasminogen activator receptor and EGF receptor variant III supports survival and growth of glioblastoma cells , 2011, Proceedings of the National Academy of Sciences.

[19]  C. Marshall,et al.  Regulation of cell signalling by uPAR , 2010, Nature Reviews Molecular Cell Biology.

[20]  J. Tremblay,et al.  cAMP-induced expression of the orphan nuclear receptor Nur77 in MA-10 Leydig cells involves a CaMKI pathway. , 2008, Journal of andrology.

[21]  F. Blasi,et al.  The urokinase receptor as an entertainer of signal transduction. , 2009, Frontiers in bioscience.

[22]  J. Tremblay,et al.  The orphan nuclear receptor NUR77 regulates hormone-induced StAR transcription in Leydig cells through cooperation with Ca2+/calmodulin-dependent protein kinase I. , 2008, Molecular endocrinology.

[23]  G. Ricci,et al.  Hepatocyte growth factor-modulated rat Leydig cell functions. , 2007, Journal of andrology.

[24]  M. Crippa.  Urokinase-type plasminogen activator. , 2007, The international journal of biochemistry & cell biology.

[25]  F. Odet,et al.  The mouse testis is the source of various serine proteases and serine proteinase inhibitors (SERPINs): Serine proteases and SERPINs identified in Leydig cells are under gonadotropin regulation. , 2006, Endocrinology.

[26]  G. Hosgood,et al.  Immunohistochemical staining of urokinase plasminogen activator-like and urokinase plasminogen activator receptor-like proteins in the urinary tract of healthy dogs. , 2006, American journal of veterinary research.

[27]  P. Crépieux,et al.  Extracellular signal-regulated kinases (ERK) 1, 2 are required for luteinizing hormone (LH)-induced steroidogenesis in primary Leydig cells and control steroidogenic acute regulatory (StAR) expression. , 2005, Reproduction, nutrition, development.

[28]  O. Söder,et al.  Extracellular signal-regulated kinases are involved in the acute activation of steroidogenesis in immature rat Leydig cells by human chorionic gonadotropin. , 2004, Endocrinology.

[29]  W. Folk,et al.  MAPK and JNK transduction pathways can phosphorylate Sp1 to activate the uPA minimal promoter element and endogenous gene transcription. , 2004, Blood.

[30]  F. Odet,et al.  Evidence for similar expression of protein C inhibitor and the urokinase-type plasminogen activator system during mouse testis development. , 2004, Endocrinology.

[31]  V. Papadopoulos,et al.  Differential utilization of the promoter of peripheral-type benzodiazepine receptor by steroidogenic versus nonsteroidogenic cell lines and the role of Sp1 and Sp3 in the regulation of basal activity. , 2004, Endocrinology.

[32]  G. Ricci,et al.  Functional role of hepatocyte growth factor receptor during sperm maturation. , 2002, Journal of andrology.

[33]  I. Huhtaniemi,et al.  Mechanisms of Epidermal Growth Factor Signaling: Regulation of Steroid Biosynthesis and the Steroidogenic Acute Regulatory Protein in Mouse Leydig Tumor Cells1 , 2002, Biology of reproduction.

[34]  E. S. Díaz,et al.  Effect of extracellular matrix proteins on in vitro testosterone production by rat Leydig cells , 2002, Molecular reproduction and development.

[35]  D. Stocco.  Tracking the role of a star in the sky of the new millennium. , 2001, Molecular endocrinology.

[36]  S. Friedman,et al.  1 in vascular endothelial cells β Zf 9 / COPEB activates latent TGF-Transcriptional activation of urokinase by the Krüppel-like factor , 2000 .

[37]  P. Abrahamsson,et al.  Factors of the plasminogen activator system in human testis, as demonstrated by in-situ hybridization and immunohistochemistry. , 1999, Molecular human reproduction.

[38]  J. Tremblay,et al.  Transcription factor GATA-4 enhances Müllerian inhibiting substance gene transcription through a direct interaction with the nuclear receptor SF-1. , 1999, Molecular endocrinology.

[39]  S. Jaramillo,et al.  Synergistic transcriptional activation of the mouse urokinase plasminogen activator (uPA) gene and of its enhancer activator protein 1 (AP1) site by cAMP and retinoic acid. , 1998, Biochemical Journal.

[40]  Y. Liu,et al.  Expression of plasminogen activator and inhibitor, urokinase receptor and inhibin subunits in rhesus monkey testes. , 1997, Molecular human reproduction.

[41]  P. Di Bonito,et al.  Hormonal regulation of urokinase- and tissue- type plasminogen activator in rat Sertoli cells. , 1995, Biology of reproduction.

[42]  D. Piscitelli,et al.  Identification of 3',5'-cyclic adenosine monophosphate-inducible nuclear factors binding to the human urokinase promoter in mouse Sertoli cells. , 1993, Molecular endocrinology.

[43]  G. Michalopoulos,et al.  Activation of hepatocyte growth factor by the plasminogen activators uPA and tPA. , 1993, The American journal of pathology.

[44]  D. Belin,et al.  Involvement of the plasminogen activator/plasmin proteolytic cascade in fertilization. , 1993, Developmental biology.

[45]  Y Fujii-Kuriyama,et al.  cDNA cloning and transcriptional properties of a novel GC box-binding protein, BTEB2. , 1993, Nucleic acids research.

[46]  W. Birchmeier,et al.  Extracellular proteolytic cleavage by urokinase is required for activation of hepatocyte growth factor/scatter factor. , 1992, The EMBO journal.

[47]  R. Umek,et al.  Regulated expression of three C/EBP isoforms during adipose conversion of 3T3-L1 cells. , 1991, Genes & development.

[48]  T. Lane,et al.  Adhesion, shape, proliferation, and gene expression of mouse Leydig cells are influenced by extracellular matrix in vitro. , 1991, Biology of reproduction.

[49]  P. Verde,et al.  Follicle-stimulating hormone and cyclic AMP induce transcription from the human urokinase promoter in primary cultures of mouse Sertoli cells. , 1990, Molecular endocrinology.

[50]  D. Belin,et al.  Regulation of urokinase- and tissue-type plasminogen activator gene expression in the rat seminiferous epithelium. , 1989, Molecular endocrinology.

[51]  J. Toppari,et al.  Testicular plasminogen activators during postnatal development in the rat. , 1986, Acta endocrinologica.

[52]  E. Waller,et al.  Hormonal stimulation alters the type of plasminogen activator produced by Sertoli cells. , 1986, Biology of reproduction.

[53]  S. Strickland,et al.  Studies on the hormonal regulation of plasminogen activator production in the rat ovary. , 1986, Endocrinology.

[54]  R. Miskin,et al.  Follicular plasminogen activator: involvement in ovulation. , 1985, Endocrinology.

[55]  A. Vaheri,et al.  The cellular target for the plasminogen activator, urokinase, in human fibroblasts - 66 000 dalton protein. , 1982, Biochimica et biophysica acta.

[56]  L. Liotta,et al.  Effect of plasminogen activator (urokinase), plasmin, and thrombin on glycoprotein and collagenous components of basement membrane. , 1981, Cancer research.

[57]  M. Ascoli.  Characterization of several clonal lines of cultured Leydig tumor cells: gonadotropin receptors and steroidogenic responses. , 1981, Endocrinology.

[58]  K. Catt,et al.  Intermediate role of adenosine 3':5'-cyclic monophosphate and protein kinase during gonadotropin-induced steroidogenesis in testicular interstitial cells. , 1977, Proceedings of the National Academy of Sciences of the United States of America.

[59]  M. Kloczewiak,et al.  Characterisation and comparison of macromolecular end products of fibrinogen and fibrin proteolysis by plasmin. , 1970, Biochimica et biophysica acta.

[60]  P. Hall,et al.  Conversion of cholesterol to androgens by rat testes: comparison of interstitial cells and seminiferous tubules. , 1969, Endocrinology.

[61]  L. Ewing,et al.  On the formation of testosterone by the perfused rabbit testis. , 1966, Canadian journal of biochemistry.