Regulation of mouse preimplantation development: inhibition of synthesis of proteins in the two-cell embryo that require transcription by inhibitors of cAMP-dependent protein kinase.

Perturbing the changes in protein phosphorylation accompanying the first cleavage can inhibit the appearance of a set of proteins whose synthesis is inhibited by alpha-amanitin (transcription-requiring proteins, TRPs) (W. T. Poueymirou and R. M. Schultz, 1987, Dev. Biol. 121, 489-498); synthesis of the TRPs is likely to represent activation of transcription of the embryonic genome that occurs at the 2-cell stage during mouse development. In the present study, we report the effects of three different inhibitors of the cAMP-dependent protein kinase, N-[2-(methylamino)ethyl]-5-isoquinoline-sulfonamide (H8), (Rp)-cAMPs, and protein kinase inhibitor (PKI), each of which inhibits the kinase by a different mechanism, on cleavage of 2-cell embryos and synthesis of the TRPs. Two-cell embryos possess PK-A activity, which is inhibited by each of these inhibitors. Both H8 and (Rp)-cAMPs inhibit cleavage of 2-cell embryos in a concentration-dependent manner; similar concentrations of H7, which is a less potent inhibitor of PK-A, do not inhibit cleavage. H8 and (Rp)-cAMPS inhibit in a concentration-dependent manner TRP synthesis, whereas higher concentrations of H7 are required to inhibit TRP synthesis. Microinjected PKI also inhibits synthesis of the TRPs. In addition, H8 inhibits the accumulation of translatable messenger RNAs that are likely to encode for the TRPs. Last, H8, but not H7, inhibits the phosphorylation of a phosphoprotein in 2-cell embryos. Results of these studies suggest a role for protein phosphorylation catalyzed by cAMP-dependent protein kinase in regulating transcription in the early mouse embryo.