Vg1RBP phosphorylation does not affect its RNA binding or self-association. (A) Phosphorylation of Vg1RBP does not affect RNA binding. S10 extracts from stage VI oocytes (VI) or progesterone-matured eggs (E) and 0.75 pmol recombinant Vg1RBP variants were tested in a UV cross-linking assay using VLE RNA probe uniformly labeled with [α-32P]-UTP. Where indicated, proteins were prephosphorylated with recombinant in vitro activated p38α (+ p38α**). A parallel reaction with inactive p38α (+ p38α) was used as a control. The samples were resolved using SDS-PAGE and visualized by autoradiography. (B) Phosphorylation of Vg1RBP does not affect self association. S10 extracts from stage VI oocytes (VI) or progesterone-matured eggs (E) and 15 pmol recombinant Vg1RBP variants were cross-linked by addition of 1.5 mg/mL dimethyl suberimidate (DMS) in borate buffer (+) or borate buffer alone (−). Where indicated, proteins were prephosphorylated as above. Samples were resolved on phosphate SDS-PAGE gel and subjected to Western blot analysis using anti-Vg1RBP antiserum. Migration of monomer and dimer forms is indicated on the left.
