Vitamin A regulation of BMP4 expression in the male germ line.

The molecular mechanisms leading to male infertility in vitamin A deficient (VAD) rodents have never been fully elucidated. Here, we report an interaction between BMP4 and retinoid signaling pathways in germ cells that may help clarify the biochemical basis of VAD. Adult germ cells, in particular spermatogonia, expressed BMP4 at both the mRNA and protein levels. BMP4 expression was significantly up-regulated in the testes of VAD mice and was down-regulated in freshly isolated germ cells and VAD testes by retinol, but not retinoic acid. The retinoid-responsive gene, RARbeta, was not induced in germ cells following retinoid treatment. Examination of BMP4 promoter usage in spermatogonia and the VAD testis revealed that germ cells utilize the recently characterized BMP4 intron 2 promoter, in addition to the classical 1A and 1B promoters. The observed decrease in BMP4 in response to retinol was mediated by the 1A and intron 2 promoters of the BMP4 gene. Our results reflect a direct requirement for retinoids by germ cells for the resumption of spermatogenesis in VAD animals via mechanisms that involve the suppression of BMP4 expression.

[1]  W. Hembree,et al.  Spermatogenic response to vitamin A in vitamin A deficient rats. , 1979, Biology of reproduction.

[2]  B. Hogan,et al.  Bone morphogenetic protein 4 in the extraembryonic mesoderm is required for allantois development and the localization and survival of primordial germ cells in the mouse , 2001, Proceedings of the National Academy of Sciences of the United States of America.

[3]  G. Yamada,et al.  Retinoic acid teratogenicity: the role of goosecoid and BMP-4. , 1999, Cellular and Molecular Biology.

[4]  K. Umesono,et al.  Cell-Type-Specific Regulation of the Retinoic Acid Receptor Mediated by the Orphan Nuclear Receptor TLX , 2000, Molecular and Cellular Biology.

[5]  P. Chomczyński,et al.  Single-step method of RNA isolation by acid guanidinium thiocyanate-phenol-chloroform extraction. , 1987, Analytical biochemistry.

[6]  P. Chambon,et al.  Synergistic activation of retinoic acid (RA)-responsive genes and induction of embryonal carcinoma cell differentiation by an RA receptor alpha (RAR alpha)-, RAR beta-, or RAR gamma-selective ligand in combination with a retinoid X receptor-specific ligand , 1995, Molecular and cellular biology.

[7]  H. Stunnenberg,et al.  Identification of a retinoic acid responsive element in the retinoic acid receptor & beta;gene , 1990, Nature.

[8]  P. Rossi,et al.  Developmental expression of BMP4/ALK3/SMAD5 signaling pathway in the mouse testis: a potential role of BMP4 in spermatogonia differentiation , 2003, Journal of Cell Science.

[9]  C. Ho,et al.  Overexpression of iNOS and down-regulation of BMPs-2, 4 and 7 in retinoic acid induced cleft palate formation. , 2004, Histology and histopathology.

[10]  Y. Mishina,et al.  Developmental expression and function of Bmp4 in spermatogenesis and in maintaining epididymal integrity. , 2004, Developmental biology.

[11]  B. Hogan,et al.  Bone morphogenetic protein 8A plays a role in the maintenance of spermatogenesis and the integrity of the epididymis. , 1998, Development.

[12]  T. Itano,et al.  Altered distribution of Sertoli cell vimentin and increased apoptosis in cryptorchid rats. , 2002, Journal of pediatric surgery.

[13]  N. Schultz,et al.  A multitude of genes expressed solely in meiotic or postmeiotic spermatogenic cells offers a myriad of contraceptive targets , 2003, Proceedings of the National Academy of Sciences of the United States of America.

[14]  D. D. de Rooij,et al.  Differential expression pattern of retinoid X receptors in adult murine testicular cells implies varying roles for these receptors in spermatogenesis. , 1998, Biology of reproduction.

[15]  H. Towbin,et al.  Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. 1979. , 1992, Biotechnology.

[16]  Retinoids , 1999, Handbook of Experimental Pharmacology.

[17]  P. Chambon,et al.  Multiple isoforms of the mouse retinoic acid receptor alpha are generated by alternative splicing and differential induction by retinoic acid. , 1991, The EMBO journal.

[18]  R. W. Padgett,et al.  Transforming growth factor β signaling mediators and modulators , 2000 .

[19]  D. G. Rooij,et al.  Retinoic acid is able to reinitiate spermatogenesis in vitamin A-deficient rats and high replicate doses support the full development of spermatogenic cells. , 1991 .

[20]  H. Towbin,et al.  Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. , 1979, Proceedings of the National Academy of Sciences of the United States of America.

[21]  J. Lehmann,et al.  A Retinoic Acid Receptor-Specific Element Controls the Retinoic Acid Receptor-β Promoter , 1990 .

[22]  M. Glozak,et al.  Specific induction of apoptosis in P19 embryonal carcinoma cells by retinoic acid and BMP2 or BMP4. , 1996, Developmental biology.

[23]  J. Rodríguez-León,et al.  Retinoic acid regulates programmed cell death through BMP signalling , 1999, Nature Cell Biology.

[24]  T. Tsuda,et al.  Sonic hedgehog and bone morphogenetic protein 4 expressions in the hindgut region of murine embryos with anorectal malformations. , 2004, Journal of pediatric surgery.

[25]  A. Cooney,et al.  The Mouse Bone Morphogenetic Protein-4 Gene , 1995, The Journal of Biological Chemistry.

[26]  G. Enders,et al.  Differentiation of murine premigratory primordial germ cells in culture. , 1999, Biology of reproduction.

[27]  P. Chambon,et al.  Developmental analysis of the retinoic acid-inducible RAR-beta 2 promoter in transgenic animals. , 1991, Development.

[28]  B. Hogan,et al.  The gene encoding bone morphogenetic protein 8B is required for the initiation and maintenance of spermatogenesis in the mouse. , 1996, Genes & development.

[29]  D. Wolgemuth,et al.  Role of retinoid signaling in the regulation of spermatogenesis , 2004, Cytogenetic and Genome Research.

[30]  K. Kim,et al.  Cellular and subcellular localization of six retinoid receptors in rat testis during postnatal development: identification of potential heterodimeric receptors. , 1999, Biology of reproduction.

[31]  M. Dym,et al.  Spermatogenic cells of the prepuberal mouse: isolation and morphological characterization , 1977, The Journal of cell biology.

[32]  A. Geiser,et al.  Regulation of the promoters for the human bone morphogenetic protein 2 and 4 genes. , 2000, Gene.

[33]  R. Habert,et al.  Retinoic acid receptors and retinoid X receptors in the rat testis during fetal and postnatal development: immunolocalization and implication in the control of the number of gonocytes. , 1999, Biology of reproduction.

[34]  B. Hogan,et al.  Bmp4 is required for the generation of primordial germ cells in the mouse embryo. , 1999, Genes & development.

[35]  P. Chambon,et al.  Mouse retinoic acid receptor alpha 2 isoform is transcribed from a promoter that contains a retinoic acid response element. , 1991, Proceedings of the National Academy of Sciences of the United States of America.

[36]  U. K. Laemmli,et al.  Cleavage of Structural Proteins during the Assembly of the Head of Bacteriophage T4 , 1970, Nature.

[37]  P. Chambon,et al.  Abnormal spermatogenesis in RXR beta mutant mice. , 1996, Genes & development.

[38]  M. Glozak,et al.  Retinoic acid- and bone morphogenetic protein 4-induced apoptosis in P19 embryonal carcinoma cells requires p27. , 2001, Experimental cell research.

[39]  Zhaoyuan Hu,et al.  Disrupted expression of intermediate filaments in the testis of rhesus monkey after experimental cryptorchidism. , 2004, International journal of andrology.

[40]  R. Evans,et al.  Characterization of an autoregulated response element in the mouse retinoic acid receptor type beta gene. , 1990, Proceedings of the National Academy of Sciences of the United States of America.

[41]  Deborah L. Thompson,et al.  Retinoic Acid Repression of Bone Morphogenetic Protein 4 in Inner Ear Development , 2003, Molecular and Cellular Biology.

[42]  G. Duester,et al.  Retinoid activation of retinoic acid receptor but not retinoid X receptor is sufficient to rescue lethal defect in retinoic acid synthesis , 2003, Proceedings of the National Academy of Sciences of the United States of America.

[43]  X. Liu,et al.  Requirement of Bmp8b for the generation of primordial germ cells in the mouse. , 2000, Molecular endocrinology.

[44]  B. Zirkin,et al.  Reduced intratesticular testosterone concentration alters the polymerization state of the Sertoli cell intermediate filament cytoskeleton by degradation of vimentin. , 2003, Endocrinology.

[45]  D. Wolgemuth,et al.  Genetic and Molecular Approaches to Understanding the Role of Retinoids in Mammalian Spermatogenesis , 1999 .

[46]  C. Mendelsohn,et al.  Regulation of retinoic acid signaling during lung morphogenesis. , 2000, Development.

[47]  Z. Xu,et al.  Mutation in Bmp7 exacerbates the phenotype of Bmp8a mutants in spermatogenesis and epididymis. , 2001, Developmental biology.

[48]  Thomas D. Schmittgen,et al.  Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)) Method. , 2001, Methods.

[49]  P. Chambon,et al.  High postnatal lethality and testis degeneration in retinoic acid receptor alpha mutant mice. , 1993, Proceedings of the National Academy of Sciences of the United States of America.