ABSTRACT
Here we report that ILK localizes in the mouse primary cilium, a sensory organelle required for signalling by the Hedgehog (Hh) pathway. Genetic or pharmacological inhibition of ILK blocks ciliary accumulation of the Hh pathway effector smoothened (Smo) and suppresses the induction of Gli transcription factor mRNAs by SHh. Conditional deletion of ILK or Smo also inhibits SHh-driven activation of Gli2 in the embryonic mouse cerebellum. ILK regulation of Hh signalling probably requires the physical interaction of ILK and Smo in the cilium, and we also show selective cilia-associated interaction of ILK with ß-arrestin, a known mediator of Smo-dependent signalling.
Subject(s)
Cerebellum/metabolism , Cilia/metabolism , Hedgehog Proteins/metabolism , Protein Serine-Threonine Kinases/metabolism , Receptors, G-Protein-Coupled/metabolism , Signal Transduction , Animals , Arrestins/metabolism , Cell Line , Cerebellum/embryology , Kruppel-Like Transcription Factors/genetics , Kruppel-Like Transcription Factors/metabolism , Mice , Protein Kinase Inhibitors/pharmacology , Protein Serine-Threonine Kinases/antagonists & inhibitors , Protein Serine-Threonine Kinases/genetics , RNA, Messenger/genetics , RNA, Messenger/metabolism , Receptors, G-Protein-Coupled/genetics , Smoothened Receptor , Zinc Finger Protein Gli2ABSTRACT
Testicular development is governed by the combined influence of hormones and proteins, including FSH, inhibins, activins and follistatin (FST). This study documents the expression of these proteins and their corresponding mRNAs, in testes and serum from mice aged 0 through 91 days post partum (dpp), using real-time PCR, in situ hybridisation, immunohistochemistry, ELISA and RIA. Serum immunoactive total inhibin and FSH levels were negatively correlated during development, with FSH levels rising and inhibin levels falling. Activin A production changed significantly during development, with subunit mRNA and protein levels declining rapidly after 4 dpp, while simultaneously levels of the activin antagonists, FST and inhibin/activin beta(C), increased. Inhibin/activin beta(A) and beta(B) subunit mRNAs were detected in Sertoli, germ and Leydig cells throughout testis development, with the beta(A) subunit also detected in peritubular myoid cells. The alpha, beta(A), beta(B) and beta(C) subunit proteins were detected in Sertoli and Leydig cells of developing and adult mouse testes. While beta(A) and beta(B) subunit proteins were observed in spermatogonia and spermatocytes in immature testes, beta(C) was localised to leptotene and zygotene spermatocytes in immature and adult testes. Nuclear beta(A) subunit protein was observed in primary spermatocytes and nuclear beta(C) subunit in gonocytes and round spermatids. The changing spatial and temporal distributions of inhibins and activins indicate that their modulated synthesis and action are important during onset of murine spermatogenesis. This study provides a foundation for evaluation of these proteins in mice with disturbed testicular development, enabling their role in normal and perturbed spermatogenesis to be more fully understood.
Subject(s)
Activins/biosynthesis , Follicle Stimulating Hormone/blood , Follistatin/biosynthesis , Inhibins/biosynthesis , Testis/metabolism , Activins/blood , Animals , Follicle Stimulating Hormone/biosynthesis , Follistatin/blood , Immunohistochemistry , Inhibins/blood , Male , Mice , Mice, Inbred C57BL , Mice, Inbred CBA , RNA, Messenger/analysis , RNA, Messenger/metabolism , Spermatocytes/chemistry , Spermatocytes/metabolism , Spermatogenesis/physiology , Spermatogonia/chemistry , Spermatogonia/metabolism , Spermatozoa/chemistry , Spermatozoa/metabolism , Testicular Diseases/metabolism , Testis/chemistry , Testis/physiologyABSTRACT
To achieve and maintain fertility, the adult mammalian testis produces many generations of sperm. While testicular integrity is established in the fetus and develops further in juvenile life, sperm production does not ensue until much later in life, following the onset of puberty. Signals from the transforming growth factor-beta superfamily of proteins are vital for governance of testis development and spermatogenesis, and this review discusses our current understanding of the mechanisms and processes in which they have been implicated with a focus on the fetal and juvenile testis.