Glucocorticoid receptor/signal transducer and activator of transcription 5(STAT5) interactions enhance STAT5 activation by prolonging STAT5 DNA binding and tyrosine phosphorylation

The regulation of casein gene expression by both PRL and glucocorticoids ha s been a well studied paradigm for understanding how the signaling pathways regulated by these two hormones interact in the nucleus. Previous studies have demonstrated that the downstream effecters of these pathways, signal t ransducer and activator of transcription 5 (STAT5) and the glucocorticoid r eceptor (GR), are associated via protein-protein interactions and act syner gistically to enhance beta-casein gene transcription. Indirect immunofluore scence microscopy was used to demonstrate that PRL-activated STATE can tran slocate GR into the nucleus, and that ligand-bound GR can translocate STAT5 into the nucleus. This provided further support of an interaction between the two proteins. To better understand the mechanism of transcriptional syn ergy between STATE and GR, experiments were performed in cells transiently transfected with STATE alone or with STAT5 and GR. GR cotransfection enhanc ed the DNA-binding activity of STAT5 without affecting STAT5 protein levels . The enhancement of STAT5 DNA binding by GR resulted in the formation of a complex that exhibited prolonged DNA binding after PRL treatment. This was correlated with increased STAT5 tyrosine phosphorylation, suggesting that GR enhances STATE DNA binding by modulating the rate of STAT5 dephosphoryla tion. In contrast, cotransfection of the estrogen receptor resulted in an o verall decrease in STAT5 tyrosine phosphorylation, without changing the kin etics of dephosphorylation. Enhancement of STAT5 activity by GR is, therefo re, one component of the transcriptional synergy exhibited by STATE and GR at the beta-casein promoter and is an example of how transcription factors at a composite response element may modulate each other's activity.