MULTIPLE PROLACTIN (PRL) RECEPTOR CYTOPLASMIC RESIDUES AND STAT1 MEDIATE PRL SIGNALING TO THE INTERFERON REGULATORY FACTOR-I PROMOTER

The Nb2 PRL receptor (PRL-R) is known to mediate PRL signaling to the interferon (IFN) regulatory factor-1 (IRF-1) gene via the family of si gnal transducers and activators of transcription or Stats. To analyze the components of the PRL-R/Stat/lRF-1 signaling pathway, various PRL- R, Stat, and IRF-1-CAT reporter constructs were transiently cotransfec ted into COS cells, First, mutations in the IFN gamma-activated sequen ce (GAS), either multimerized or in the context of the 1.7-kb IRF-1 pr omoter, failed to mediate a PRL response, showing that the IRF-1 GAS i s a target of PRL signaling. Next, pairwise alanine substitutions into conserved residues in the proline-rich motif or Box 1 region and two tyrosine mutations, Y308F and Y382F, in the PRL-R intracellular domain all impaired PRL signaling to multimerized GAS or to the 1.7-kb IRF-1 promoter, Furthermore, these PRL-R mutants mediated reduced Stat1 bin ding to the IRF-1 GAS. Transfection of Stat1 further enhanced PRL sign aling to the IRF-1 promoter, suggesting that Stat1 is a positive media tor of PRL action, These studies show that both membrane proximal and distal residues of the PRL-R are involved in signaling to the IRF-1 ge ne. Further, Stat1 and the GAS element are important for PRL activatio n of the IRF-1 gene.