2 TESTICULAR CDNA CLONES SUPPRESSED BY GONADOTROPIN STIMULATION EXHIBIT ZP2-LIKE AND ZP3-LIKE STRUCTURES IN JAPANESE EEL

A single injection of human chorionic gonadotropin (HCG) can induce co mplete spermatogenesis in immature eel testes consisting of premitotic spermatogonia. To understand the regulatory mechanisms of spermatogen esis, we have applied a subtractive hybridization method to identify g enes in which changes in expression occur after HCG treatment in vivo. The subtraction was carried out 24 hours after HCG injection. Two up- regulated and six down regulated cDNA clones by HCG stimulation were i solated, and named eel spermatogenesis-related substance (eSRS) 1 to 8 . In this paper, down-regulated cDNA clones of eSRS3 and eSRS4 were se quenced. A homology search showed that eSRS3 and eSRS4 have amino acid sequences similar to those of the ZP-domains of zona pellucida sperm- binding protein (ZP)-2 and 3, respectively. Transcripts of eSRS3 and e SRS4 have been detected only in immature testes and ovaries. Both tran scripts disappeared immediately after HCG injection and were not detec ted in testes throughout the experimental period. To determine whether HCG action on down-regulation of eSRS3 and eSRS4 transcription is dir ect or mediated through 11-ketotestosterone (11-KT), a spermatogenesis -inducing steroid in eel, we investigated the effect of HCG and 11-KT on testicular eSRS3 and eSRS4 mRNA transcription in vitro. Northern bl ot analysis using poly(A)(+) RNA extracted from cultured testis showed that both HCG and 11-KT suppressed the mRNA transcription of both eSR S3 and eSRS4. We speculate that eSRS3 and eSRS4 may play important rol es in the prevention of spermatogenesis in the eel. (C) 1998 Wiley-Lis s, Inc.