Rap1 reverses transcriptional repression of TGF-β type II receptor by a mechanism involving AP-1 in the human pancreatic cancer cell line, UK Pan-1

Kimberly D. Fralix, Shujie Zhao, Kolaparthi Venkatasubbarao, James W. Freeman

Research output: Contribution to journalArticlepeer-review

13 Scopus citations

Abstract

The TGF-β signaling pathway has potent anti-mitogenic effects in epithelial cells and loss of negative growth regulation is often associated with increased tumorigenicity. The human pancreatic ductal adenocarcinoma cell line, UK Pan-1, which expresses DPC4, is not highly responsive to TGF-β due to transcriptional repression of TGF-β-type II receptor (RII). Here, we show that UK Pan-1 cells transfected with a plasmid to overexpress rap1 protein (UK/rap1) causes an increase in RII transcription and restores sensitivity to TGF-β growth inhibition. The overexpression of rap1 was associated with diminished ras signaling as measured by ras binding domain (RBD)-binding assays. Electrophoretic mobility shift assays (EMSA) analysis revealed increased binding of nuclear proteins to a previously identified positive regulatory element (PRE1) of the RII promoter in rap1 transfected cells. Competition with an oligo containing the AP-1 consensus site was able to inhibit this binding of nuclear proteins to the PRE1 region. Further EMSA analysis using antibodies to various AP-1 components revealed that junB antibodies partially depleted the increase in binding to the PRE1 seen in UK/rap1 cells while antibodies to other AP-1 constituents such as c-jun, c-fos, and ATF-1 had no effect on binding. Consistent with this data, transient transfection of UK Pan-1 cells with junB resulted in greater RII transcription (twofold) as measured by RII-luciferase assay. Mutation of the AP-1 site inhibited junB-mediated or rap1-mediated increases in RII promoter activity. These data suggest that rap1 signaling may mediate an increase in RII transcription via increased binding of nuclear factors including junB to the PRE1 region of the RII promoter.

Original languageEnglish (US)
Pages (from-to)88-99
Number of pages12
JournalJournal of Cellular Physiology
Volume194
Issue number1
DOIs
StatePublished - Jan 1 2003

ASJC Scopus subject areas

  • Physiology
  • Clinical Biochemistry
  • Cell Biology

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