Requirement of DDX6 mediated P-body formation for the function of NANOS2 in male germ cell differentiation

Saga Y

National Institute of Genetics, Mishima, Japan.

In embryonic male gonads, germ cells produce abundant mRNA-protein complex called P-body. The formation of P-bodies is one of sex specific characteristics and is associated with male germ cell differentiation in the embryonic stage. We reported previously that an RNA binding protein NANOS2 is recruited to P-body and plays an essential role for germ cell differentiation. However, it is not clear whether P-body formation is necessary for NANOS2 to control the target RNAs and whether all NANOS2 functions are dependent on P-bodies. To understand relationship between P-body formation and NANOS2 function, we deleted P-bodies by knocking out DDX6 gene, essential for P-body formation in a germ cell-specific manner. For this purpose, we produced an XY-ES cell line which contains a germ-cell specific inducible Cre recombinase together with Cre reporter and introduced floxed DDX6 alleles via Cas9-mediated homologous recombination. The results show that NANOS2-target Dazl was not repressed and male differentiation was not promoted similar to the case of NANOS2-null. On the other hand, some NANOS2-null properties, meiosis initiation and germ cell escape from seminiferous tubules are not observed in DDX6-KO germ cells, indicating that NANOS2 also works in a P-body-independent manner. This study also demonstrates that ES-mediated chimera method offers a powerful method as a standard genetic tool.