CHANGE OF CYCLIN D2 MESSENGER-RNA EXPRESSION DURING MURINE TESTIS DEVELOPMENT DETECTED BY FRAGMENTED CDNA SUBTRACTION METHOD

Using subtractive hybridization and polymerase chain reaction, we deve loped a differential cloning system, the fragmented cDNA subtraction m ethod, that requires only small amounts of materials. The cloning syst em was used to isolate several cDNA fragments expressed more abundantl y in the premeiotic day 3 post-natal mouse testis than in the adult mo use testis. The isolated cDNA fragments included cDNA encoding the mur ine cyclin D2. Northern blot and in situ hybridization analyses reveal ed that, during testis development, cyclin D2 expression was most abun dant in the neonatal proliferating Sertoli cells. Those type A spermat ogonia that were thought to divide mitotically also expressed cyclin D 2 mRNA. Other spermatogenic cells, such as mitotically arrested gonocy tes in neonatal testis and meiotically dividing germ cells in adult te stis as well as adult Sertoli cells, were negative for the cyclin D2 s ignal. Adult W/W-v mutant mice lacking germ cells expressed cyclin D2 mRNA in terminally differentiated Sertoli cells. Elimination of germ c ells other than the undifferentiated type A spermatogonia by treating wild-type mice with an anti-c-kit monoclonal antibody did not result i n the expression of cyclin D2 in Sertoli cells. These results demonstr ate that there are lineage- and developmental-specific expression patt erns of cyclin D2 mRNA during mouse testis development. At the same ti me, it is suggested that primitive type A spermatogonia affect the cyc lin D2 expression of Sertoli cells.