Interactions between XIAP Associated Factor 1 and a Nuclear Co-Activator, CBP, in Colon Cancer Cells

Abstract

Background/Aims: XIAP-associated factor 1 (XAF1) is a nuclear protein. CBP, the cAMP response element binding protein (CREB)-binding protein, plays an important role as a multifunctional transcriptional co-activator. In this investigation, we aimed to study the putative interaction between XAF1 and CBP in colon cancer cells. Methods: Expressions of XAF1 and CBP were detected by Western blot and RT-PCR. The interaction between XAF1 and CBP was investigated by the glutathione S-transferase (GST) pull-down assay, colocalization and co-immunoprecipitation analysis. Cell proliferation was examined by cell number counting. Results: Both XAF1 and CBP were co-localized in the nuclei of colon cancer cells and they demonstrated a physical interaction, as revealed by GST pull-down assay and co-immunoprecipitation analysis. CBP I peptide (residues 1–1098) was the interacting domain for XAF1 binding. The functional implication of the interaction between XAF1 and CBP was demonstrated by the finding that cell growth inhibition by XAF1 was potentiated by cotransfection with CBP. Furthermore, a reporter assay demonstrated that cotransfection with XAF1 and CBP led to marked reduction in phorbol ester 12-O-tetradecanoylphorbol-13-acetate (PMA)-stimulated adaptor-related protein complex 1 activity. Conclusions: CBP is a novel binding partner of XAF1, and the interaction between XAF1 and CBP and their functional consequence were mediated by adaptor-related protein complex 1.