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A Pit-1 threonine 220 phosphomimic reduces binding to monomeric DNA sites to inhibit Ras and estrogen stimulation of the prolactin gene promoter.

Authors: Jean, A  Gutierrez-Hartmann, A  Duval, DL 
Citation: Jean A, etal., Mol Endocrinol. 2010 Jan;24(1):91-103. doi: 10.1210/me.2009-0279. Epub 2009 Nov 3.
Pubmed: (View Article at PubMed) PMID:19887646
DOI: Full-text: DOI:10.1210/me.2009-0279

Pit-1 is a POU-homeodomain transcription factor that dictates the ontogeny of pituitary somatotrophs, lactotrophs, and thyrotrophs through regulation of their respective protein hormone genes: GH, prolactin (PRL), and TSHbeta. Although Pit-1 threonine 220 (T220) and serine 115 are protein kinase phospho-acceptor sites, the transcriptional role of Pit-1 phosphorylation remains unclear. In the rat PRL promoter (rPRL), Ras-stimulated transcription is mediated by binding of Ets-1 and Pit-1 at a composite site (FPIV). Ets-1 and Pit-1 physically interact, and Pit-1 T220 is a major Ets-1 contact point. T220 was mutated to aspartic acid (D, to mimic phosphorylation) or a neutral alanine (A), and DNA binding and transcriptional activity were tested. The Pit-1 T220D mutation reduced binding at monomeric Pit-1 sites (FPIV, PRL-1d), but not dimeric Pit-1 sites (FPI). Pit-1 T220A bound all sites with wild-type (WT) affinity. In transfections of HeLa cells, each Pit-1 mutant transcriptionally activated the -425rPRL promoter and cooperated with Ets-1 to WT levels. In contrast, Pit-1-mediated Ras activation of the -425 rPRL promoter was significantly inhibited by T220D. Finally, Pit-1 synergistic activation of the 2500-bp rPRL promoter with estrogen receptor was reduced by T220D compared with T220A and WT Pit-1. Thus, phosphorylation of Pit-1 T220 reduces binding to monomeric sites blunting Ras and estrogen/estrogen receptor stimulation of the rPRL gene promoter. Consequently, T220 phosphorylation of Pit-1 by protein kinase A, protein kinase C, or cell cycle-dependent kinases appears to serve as a regulatory switch, inhibiting Ras and estrogen/estrogen receptor regulatory pathways, while enhancing the cAMP/protein kinase A response, thus allowing a more precise integration of pituitary responses to distinct signaling stimuli.


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CRRD Object Information
CRRD ID: 8661635
Created: 2014-06-11
Species: All species
Last Modified: 2014-06-11
Status: ACTIVE


RGD is funded by grant HL64541 from the National Heart, Lung, and Blood Institute on behalf of the NIH.