Induction of relaxin messenger RNA expression in response to prolactin receptor activation requires protein kinase C delta signaling.
The ability of PRL or rat placental lactogen (rPL)-1 to induce relaxin mRNA expression was analyzed in a luteinized rat granulosa cell culture model. PRL receptor activation induced relaxin mRNA expression in a concentration- and time-dependent manner. High concentrations of PRL receptor agonist, equivalent to those of the second half of pregnancy in rats, were required to elicit relaxin mRNA expression. A 40-fold induction of relaxin mRNA was observed in cells treated 24 h with 1 microg/ml of rPL-1. Estrogen enhanced relaxin expression induced by PRL but did not affect relaxin expression on its own. PRL/rPL-1 induction of relaxin expression was independent of the extracellular regulated kinase (ERK) members of the mitogen-activated protein kinase (MAPK) pathway, based on the inability of the ERK kinase inhibitor PD98059 to block induction of relaxin expression. PRL/rPL-1 induction of relaxin expression required protein kinase C (PKC) delta, based on the ability of the preferential PKC delta inhibitor rottlerin to abolish induction of relaxin expression. Direct activation of PKC by phorbol myristate acetate, however, was not sufficient to promote induction of relaxin mRNA expression. Stats (signal transducers and activators of transcription) 3 and 5 DNA binding activities were induced by PRL/rPL-1 treatment of luteinized granulosa cells but only Stat 3 DNA binding was reduced by rottlerin. PRL/rPL-1 treatment of luteinized granulosa cells resulted in increased phosphorylation on tyrosine-705 and serine-727 of Stat 3, and these responses were reduced and blocked, respectively, by rottlerin. Tyrosine and serine phosphorylations of Stat 3 in the corpus luteum were also increased in the second half of pregnancy when PL levels are highest. Stat 3, but not Stat 1 or 5, coimmunoprecipitated with luteal PKC delta during pregnancy; Stat 3 transiently coimmunoprecipitated with PKC delta from luteinized granulosa cells in response to PRL receptor activation; and Stat 3/PKC delta complex formation required PKC delta kinase activity. Taken together, these results show that PKC delta is obligatory for PRL/rPL-1-dependent relaxin expression, that PKC delta complexes with Stat 3 in response to PRL receptor activation, and that PKC delta is involved in the regulation of Stat 3 phosphorylation downstream of the PRL receptor. These results demonstrate that PRL/rPL-1 promotes relaxin expression in luteal cells and that this event is mediated, at least in part, via PKC delta.
Peters
C A
CA
Department of Cell and Molecular Biology, Northwestern University Medical School, Chicago, Illinois 60611, USA.
Maizels
E T
ET
Robertson
M C
MC
Shiu
R P
RP
Soloff
M S
MS
Hunzicker-Dunn
M
M
D000098:Acetophenones / Q000494:pharmacology; D000818:Animals; D001578:Benzopyrans / Q000494:pharmacology; D004268:DNA-Binding Proteins / Q000378:metabolism; D004958:Estradiol / Q000494:pharmacology; D005260:Female; D005419:Flavonoids / Q000494:pharmacology; D015870:Gene Expression / Q000187:drug effects*; D007163:Immunosorbent Techniques; D007527:Isoenzymes / Q000378:metabolism*; D020929:Mitogen-Activated Protein Kinase Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D010766:Phosphorylation; D011247:Pregnancy; D011388:Prolactin / Q000494:pharmacology; D011493:Protein Kinase C / Q000378:metabolism*; D012333:RNA, Messenger / Q000096:biosynthesis*; D051381:Rats; D012065:Relaxin / Q000235:genetics*; D050796:STAT3 Transcription Factor; D015398:Signal Transduction; D015534:Trans-Activators / Q000378:metabolism
D000098:Acetophenones; D001578:Benzopyrans; D004268:DNA-Binding Proteins; D005419:Flavonoids; D007527:Isoenzymes; D012333:RNA, Messenger; D050796:STAT3 Transcription Factor; C494088:Stat3 protein, rat; D015534:Trans-Activators; D004958:Estradiol; D011388:Prolactin; D012065:Relaxin; C085746:rottlerin; D011493:Protein Kinase C; D020928:Mitogen-Activated Protein Kinases; D020929:Mitogen-Activated Protein Kinase Kinases; C093973:2-(2-amino-3-methoxyphenyl)-4H-1-benzopyran-4-one