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Transforming growth factor-beta1 inhibits rat prolactin promoter activity in GH4 neuroendocrine cells


Transforming growth factor-beta1 inhibits rat prolactin promoter activity in GH4 neuroendocrine cells



Dna and Cell Biology 18(11): 863-873



ISSN/ISBN: 1044-5498

PMID: 10595400

DOI: 10.1089/104454999314863

The prototypic member of the transforming growth factor beta family is TGFbeta1, which is known to be important in extracellular matrix production, cell proliferation, and cell differentiation. Specifically in the pituitary lactotroph, TGFbeta1 inhibits prolactin (PRL) peptide secretion, PRL mRNA levels, and PRL gene transcription. To further elucidate the molecular details by which TGFbeta1 modulates PRL gene transcription, we used a transient transfection approach to characterize and to map the TGFbeta1 inhibitory response element of the rat (r) PRL promoter. Here, we show that TGFbeta1 selectively inhibits basal rPRL promoter activity in GH4 cells in a dose-responsive fashion, with an IC50 of 6 pM, and that this inhibition occurs within 6 h after TGFbeta1 addition. Using a series of 5' deletion promoter mutants, the TGFbeta1 inhibitory response was found to be unaffected by deletion to position - 116 and was abrogated by further deletion to -54 in the rPRL promoter. However, on the basis of data from site-specific and linker-scanning mutants of the rPRL promoter, it appears that no single element is sufficient to mediate the TGFbeta1 inhibitory effect. Sequence analysis of the -116/-54 region failed to reveal any sequence homology to previously characterized TGFbeta response elements. Finally, TGFbeta1 failed to alter significantly the endogenous levels of the cell-specific activator protein GHF-1/Pit-1, indicating that the TGFbeta1 inhibitory effect is not attributable to diminished levels of GHF-1/Pit-1. Taken together, these data indicate that the TGFbeta1 inhibitory response is more complex than previously appreciated, requiring more than one cis-acting element and not always acting via TTGG or GTCTAGAC sites.

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