Phosphorylation of Caprin by Chk1(Grapes) May Control the Cell Cycle at the Mid-Blastula Transition.

Phosphorylation of Caprin by Chk1(Grapes) May Control the Cell Cycle at the Mid-Blastula Transition. Helen X. Chen^1,3, Ophelia Papoulas^2,3, Paul Macdonald^1,3. 1) Section of Molecular Cell and Developmental Biology; 2) Center for Systems and Synthetic Biology; 3) The Institute for Cellular and Molecular Biology, The University of Texas at Austin, Austin, TX 78712.

The mid-blastula transition (MBT) is the first embryonic development event requiring zygotic gene activity. One feature of the MBT is a slowing of the cell cycle. Previously, we reported that two translational regulators, Drosophila Fragile X mental retardation protein (dFMRP; FMR1) and Cytoplasmic activation/proliferation-associated protein (Caprin; Capr) collaborate to control the cell cycle at the MBT by mediating the normal repression of maternal Cyclin B mRNA. It remains unclear how the proper timing of the MBT is determined, and whether regulation of Capr or FMR1 activity might contribute to timing. Using precisely staged Drosophila embryo extracts we show that Capr is phosphorylated in multiple domains, with changes in phosphorylation state coinciding with the MBT. Three lines of evidence demonstrate that one responsible kinase for MBT-specific phosphorylation is Chk1/Grapes (Grp). First, immunodepletion of Chk1 from embryo extracts greatly reduces phosphorylation of the central domain of Capr (Capr298-630). Second, purified Chk1 phosphorylates Capr298-630 in vitro. Third, mutation of the serine within Capr298-630 that is phosphorylated by purified Chk1 blocks phosphorylation of Capr298-630 in embryo extracts. A role for Chk1 in MBT timing is suggested by a genetic interaction. Reducing the levels of both Capr and FMR1 in embryos from mothers heterozygous for mutation of each gene has no effect on MBT timing. However, the combination of loss of Capr activity and reduction of FMR1 activity results in altered MBT timing with a precocious mitosis. A similar phenotype was obtained when, in the background of reduced Capr and FMR1, the level of Chk1 was reduced. This raises the possibility that Capr phosphorylation by Chk1 contributes to control of MBT timing. Since Capr and FMR1 control the levels of Cyclin B, the precocious mitosis might be due to altered Cyclin B regulation.