PROLACTIN REGULATION OF THE CALMODULIN-DEPENDENT PROTEIN-KINASE-III ELONGATION FACTOR-II SYSTEM IN THE RAT CORPUS-LUTEUM

AM(r) 100,000 phosphoprotein in the corpus luteum was identified as el ongation factor 2 (EF-2). Since prolactin (PRL) is necessary for optim al luteal development and protein synthesis, we determined whether thi s hormone affects the content and/or phosphorylation of EF-2 in the co rpus luteum. PRL treatment enhanced the Ca2+/calmodulin (CaM) dependen t phosphorylation of endogenous EF-2 in luteal cytoplasmic extracts. I mmunoblot analysis revealed that PRL had no effect on EF-2 levels, but examination of luteal EF-2 by two dimensional isoelectric focusing/SD S-polyacrylamide gel electrophoresis showed that PRL increased the rel ative amount of the most basic dephosphorylated forms of EF-2. This su ggests that PRL induces net dephosphorylation of the protein in vivo. Since EF-2 phosphorylation is regulated by both Ca2+/CaM-dependent kin ase III (CaM kinase III) and protein phosphatase 2A, we examined the e ffect of PRL on both enzymes. Paradoxically, PRL enhanced the in vitro activity of CaM kinase III, possibly reflecting increased kinase leve ls, but had no effect on phosphatase activity. These results suggest t hat PRL maintains luteal EF-2 in a relatively dephosphorylated state i n vivo by limiting the availability of Ca2+ and/or CaM to CaM kinase I II. These data provide strong evidence for a role of the EF-2/CaM kina se III system in PRL action in the corpus luteum.