GCNF mediated DNA methylation and stem cell differentiation

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GCNF (germ cell nuclear factor) is expressed at the gastrulation stage during embryonic development. GCNF represses pluripotency genes such as Oct-3/4 and Nanog and induces DNA methylation of their regulatory regions. In this project, the specific monoclonal antibodies against GCNF have been newly prepared in order to analyze the endogenous GCNF protein during murine ES cell differentiation. By the co-immunoprecipitation study, interaction between endogenous GCNF and Dnmt3 proteins was confirmed. At the beginning of ES differentiation, the expression level of GCNF was low, but it interacts with Dnmt3a2 and Dnmt3b. At the later stage, GCNF was transiently upregulated and it interacts with Dnmt3a1. Mass spectrometry analysis has revealed that anti-GCNF immunoprecipitates contain Trim28 and PELP1 complex, but the further analyses will be required to assess its functional significance.