A PIT-1 homeodomain mutant blocks the intranuclear recruitment of the CCAAT/enhancer binding protein α required for prolactin gene transcription

John F. Enwright, Margaret A. Kawecki-Crook, Ty C. Voss, Fred Schaufele, Richard N. Day

Research output: Contribution to journalArticle

48 Scopus citations

Abstract

The pituitary-specific homeodomain protein Pit-1 cooperates with other transcription factors, including CCAAT/enhancer binding protein α (C/EBPα), in the regulation of pituitary lactotrope gene transcription. Here, we correlate cooperative activation of prolactin (PRL) gene transcription by Pit-1 and C/EBPα with changes in the subnuclear localization of these factors in living pituitary cells. Transiently expressed C/EBPα induced PRL gene transcription in pituitary GHFT1-5 cells, whereas the coexpression of Pit-1 and C/EBPα in HeLa cells demonstrated their cooperativity at the PRL promoter. Individually expressed Pit-1 or C/EBPα fused to color variants of fluorescent proteins, occupied different subnuclear compartments in living pituitary cells. When coexpressed, Pit-1 recruited C/EBPα from regions of transcriptionally quiescent centromeric heterochromatin to the nuclear regions occupied by Pit-1. The homeodomain region of Pit-1 was necessary for the recruitment of C/EBPα. A point mutation in the Pit-1 homeodomain associated with the syndrome of combined pituitary hormone deficiency in humans also failed to recruit C/EBPα. This Pit-1 mutant functioned as a dominant inhibitor of PRL gene transcription and, instead of recruiting C/EBPα, was itself recruited by C/EBPα to centromeric heterochromatin. Together our results suggest that the intranuclear positioning of these factors determines whether they activate or silence PRL promoter activity.

Original languageEnglish (US)
Pages (from-to)209-222
Number of pages14
JournalMolecular Endocrinology
Volume17
Issue number2
DOIs
StatePublished - Feb 1 2003

ASJC Scopus subject areas

  • Molecular Biology
  • Endocrinology

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