Evidence for a role of glycogen synthase kinase-3 beta in rodent spermatogenesis.
Glycogen synthase kinase-3 beta (GSK-3 beta) regulates cell metabolism, cell cycle, and cell fate through the phosphorylation of a diverse array of substrates. Herein, we provide evidence that supports a role for GSK-3 in mammalian meiosis and spermatogenesis. Immunostaining of testis sections showed that while GSK-3 alpha was ubiquitous in the seminiferous tubules, GSK-3 beta was expressed in premeiotic type B spermatogonia, in both meiotic preleptotene and leptotene spermatocytes, as well as in Sertoli cells in both the mouse and rat. Thus, GSK-3 beta is expressed in germ cells entering meiosis. In addition, intense immunoreactivity was detected in rat step 6 though 11 spermatids. In situ hybridization (ISH) in rat testis confirmed the immunostaining pattern in leptotene and spermatids and showed a GSK-3 beta messenger RNA (mRNA) signal in some pachytene spermatocytes. The restricted pattern of expression suggests cell-specific regulation of Gsk-3 beta mRNA. To determine whether GSK-3 is required for meiosis entry, rat stage VIIa seminiferous tubule segments were cultured with selective small-molecule GSK-3 inhibitors. These compounds markedly and dose-dependently suppressed meiotic synthesis (S)-phase DNA. Since a yeast GSK-3 homolog, Rim11p (regulator of inducer of meiosis), is pivotal to meiosis entry, we tested whether GSK-3 beta complements Rim11p function in meiosis. Rim11p phosphorylates transcription factors Ume6p (unscheduled meiotic gene expression) and Ime1p (inducer of meiosis) to induce meiosis entry. Overexpression of murine GSK-3 beta in a rim11 mutant yeast failed to rescue the sporulation defect. Our finding that GSK-3 beta interacted only with Ume6p but not with IME1 in a yeast 2-hybrid assay suggests that noncomplementation reflects partial divergence in substrate specificity. This work provides the basis for future studies of GSK-3 beta signaling in mammalian meiosis and spermatogenesis.
Department of Medicine, Division of Endocrinology, Harbor-UCLA Medical Center and Research and Education Institute, Torrance, California 90509, USA.
Taylor B
TB
Guo
Kam C
KC
Chan
Harri
H
Hakovirta
Yang
Y
Xiao
Jorma
J
Toppari
Aaron P
AP
Mitchell
Wael A
WA
Salameh
D000595:Amino Acid Sequence; D000818:Animals; D015152:Blotting, Northern; D015153:Blotting, Western; D004268:DNA-Binding Proteins / Q000502:physiology; D038362:Glycogen Synthase Kinase 3 / Q000502:physiology*; D000071679:Glycogen Synthase Kinase 3 beta; D017403:In Situ Hybridization; D047908:Intracellular Signaling Peptides and Proteins; D008297:Male; D008540:Meiosis / Q000187:drug effects / Q000502:physiology; D051379:Mice; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000502:physiology; D009924:Organ Culture Techniques; D017346:Protein Serine-Threonine Kinases / Q000235:genetics; D051381:Rats; D012097:Repressor Proteins; D020133:Reverse Transcriptase Polymerase Chain Reaction; D029701:Saccharomyces cerevisiae Proteins / Q000502:physiology; D012671:Seminiferous Tubules / Q000187:drug effects / Q000502:physiology; D017386:Sequence Homology, Amino Acid; D013091:Spermatogenesis / Q000187:drug effects / Q000502:physiology*; D013094:Spermatozoa / Q000378:metabolism; D013737:Testis / Q000378:metabolism*; D014157:Transcription Factors / Q000502:physiology; D020798:Two-Hybrid System Techniques; D015003:Yeasts / Q000235:genetics
D004268:DNA-Binding Proteins; C080326:IME1 protein, S cerevisiae; D047908:Intracellular Signaling Peptides and Proteins; D009687:Nuclear Proteins; D012097:Repressor Proteins; D029701:Saccharomyces cerevisiae Proteins; D014157:Transcription Factors; C089979:UME6 protein, S cerevisiae; D000071679:Glycogen Synthase Kinase 3 beta; D017346:Protein Serine-Threonine Kinases; C090458:RIM11 protein, S cerevisiae; D038362:Glycogen Synthase Kinase 3