Dnmt3aa but Not Dnmt3ab Is Required for Maintenance of Gametogenesis in Nile Tilapia (Oreochromis niloticus).

Dnmt3a, a de novo methyltransferase, is essential for mammalian germ line DNA methylation. Only one Dnmt3a is identified in mammals, and homozygous mutants of Dnmt3a are lethal, while two Dnmt3a paralogs, dnmt3aa and dnmt3ab, are identified in teleosts due to the third round of genome duplication, and homozygous mutants of dnmt3aa and dnmt3ab are viable in zebrafish. The expression patterns and roles of dnmt3aa and dnmt3ab in gonadal development remain poorly understood in teleosts. In this study, we elucidated the precise expression patterns of dnmt3aa and dnmt3ab in tilapia gonads. Dnmt3aa was highly expressed in oogonia, phase I and II oocytes and granulosa cells in ovaries and spermatogonia and spermatocytes in testes, while dnmt3ab was mainly expressed in ovarian granulosa cells and testicular spermatocytes. The mutation of dnmt3aa and dnmt3ab was achieved by CRISPR/Cas9 in tilapia. Lower gonadosomatic index (GSI), increased apoptosis of oocytes and spermatocytes and significantly reduced sperm quality were observed in dnmt3aa-/- mutants, while normal gonadal development was observed in dnmt3ab-/- mutants. Consistently, the expression of apoptotic genes was significantly increased in dnmt3aa-/- mutants. In addition, the 5-methylcytosine (5-mC) level in dnmt3aa-/- gonads was decreased significantly, compared with that of dnmt3ab-/- and wild type (WT) gonads. Taken together, our results suggest that dnmt3aa, not dnmt3ab, plays important roles in maintaining gametogenesis in teleosts.