Prolactin-Independent Modulation of the β-Casein Response Element by Erk2 MAP Kinase

Abstract

The MAP kinases have been suggested to play a role in intracellular signalling by PRL. A reporter gene construct, PRE 3-CAT, which manifests PRL responsiveness through a Stat5-binding site (PRE), was induced by PRL in CHO cells expressing the PRL-R. A fusion protein (Gal4-Stat5 695), containing the C-terminal domain of Stat5a (amino acids 695-794) linked to the DNA-binding domain of Gal4 (Gal4 DBD), strongly activated transcription of a luciferase reporter gene. Therefore, the Stat5 C-terminus, which contains a potential MAP kinase phosphorylation site, exhibits a modular transactivating function. A kinase-defective mutant of Erk2 (iMAPK) caused a dose-dependent suppression of PRL-stimulated PRE 3-CAT, and also inhibited the induction of PRE 3-CAT by Jak2 over-expression. Correspondingly, over-expression of the MAP kinase activator v-Src increased the PRL-stimulated level of PRE 3-CAT. Gal4-Stat5 695 activity was not modulated by PRL or Jak2, consistent with the absence of the relevant tyrosine phosphorylation site at residue 694. Gal4-Stat5 695 was not inhibited by iMAPK, indicating that the C-terminal transactivation region of Stat5a is not sensitive to direct modulation of a MAP kinase pathway. These results suggest that alteration of Erk2 activity by growth factors may modulate PRL-induced gene expression by a mechanism upstream of Stat5.