Expression of cAMP Response Element Binding Protein (CREB)‐Binding Protein (CBP) and the Implication in Retinoic Acid‐Inducible Transcription Activation in Human Salivary Gland Adenocarcinoma Cell Line HSG

Abstract

In the process of retinoic acid (RA) signaling, retinoic acid receptor interacts with a coactivator complex composed of various transcription cofactors such as CREB‐binding protein (CBP)/p300 and p160 family member proteins represented by steroid receptor coactivator‐1 (SRC‐1)/NCoA1 and p300/CBP cointegrator protein (p/CIP)/ACTR. In order to investigate the relationship of CBP to the RA signaling in a human salivary gland (HSG) adenocarcinoma cell line, we examined the expression of CBP in the cells. Immunoprecipitation and immunoblotting of the nuclear extract of HSG cells with anti‐human CBP antibody showed a specific 270‐kDa band, indicating the expression of CBP in HSG cells. The immunocytochemical analysis confirmed the nuclear localization of CBP. The transfection of HSG cells with a luciferase reporter plasmid harboring an RA‐response element at the 5′‐upstream region of the reporter gene increased RA‐dependent luciferase activity approximately 3‐fold. Co‐transfection with a CBP‐expression plasmid and the luciferase reporter gene enhanced the RA‐dependent transcription activation approximately 10‐fold. The immunoprecipitates obtained with anti‐CBP antibody exhibited a histone acetyl‐transferase (HAT) activity 2‐fold higher than that obtained with the control antibody, whereas the HAT activity of the immunoprecipitates with anti‐SRC‐1 and anti‐p/CIP, which were used as comparisons, were only a little increased. The RA treatment had no effect on the level of HAT activity except in the case of using the immunoprecipitate obtained with anti‐RARα, in which case it increased the activity. These findings indicate that CBP expressed in HSG cells mediates the RA‐inducible growth and differentiation‐regulating transcription activation in concert with the retinoic acid receptors.