Abstract
BackgroundTwo stages of genome activation have been identified in the mouse embryo. Specifically, minor transcriptional activation is evident at the one-cell stage and a second major episode of activation occurs at the two-cell stage. Nuclear translocation of RNA polymerase II and phosphorylation of the C-terminal domain (CTD) of the largest enzyme subunit are major determinants of embryonic genome activation. P-TEFb, the Pol II CTD kinase, regulates transcriptional elongation via phosphorylation of the serine 2 residues of the CTD.ResultsHere, we show that the CDK9 and cyclin T1 subunits of P-TEFb are present in mouse oocytes and preimplantation embryos. Both proteins translocate to pronuclei at the late one-cell stage and are predominantly localized in nuclei at the two-cell stage. We additionally examine the effects of the CDK9-specific inhibitor, flavopiridol, on mouse preimplantation development. Our data show that treatment with the drug results in mislocalization of CDK9, cyclin T1, and phosphorylated Pol II, as well as developmental arrest at the two-cell stage.ConclusionsA change in CDK9 localization from the cytoplasm to the pronucleus occurs at the time of minor embryonic genome activation, and CDK9 accumulation at the two-cell stage is evident, concomitant with major transcriptional activation of the embryonic genome. Moreover, CDK9 inhibition triggers a developmental block at the two-cell stage. Our findings clearly indicate that CDK9 is essential for embryonic genome activation in the mouse.
Highlights
Two stages of genome activation have been identified in the mouse embryo
CDK9 and cyclin T1 in oocytes and embryos To determine if antibodies against CDK9 or cyclin T1 recognized the corresponding antigens in mouse embryos, we conducted a binding competition assay in which the antigen peptides were separately incubated with the appropriate antibodies prior to immunofluorescence staining
CDK9 and cyclin T1 levels were quantitated in mouse embryos
Summary
Two stages of genome activation have been identified in the mouse embryo. minor transcriptional activation is evident at the one-cell stage and a second major episode of activation occurs at the two-cell stage. The maternal-zygotic transition is a critical event in early mouse embryogenesis This transition transforms the highly differentiated oocyte into a totipotent blastomere, and is complete by the two-cell stage. During this transition, maternal mRNAs are degraded and the embryonic genome is activated [1]. Two transcriptional stages have been identified: a minor transcriptional wave at the one-cell stage, and a second major wave at the two-cell stage [2]. These findings are supported by the results of experiments
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