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Molecular and Cellular Biology, July 2004, p. 6525-6535, Vol. 24, No. 14
0270-7306/04/$08.00+0     DOI: 10.1128/MCB.24.14.6525-6535.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.

Transcriptional Coactivator PC4 Stimulates Promoter Escape and Facilitates Transcriptional Synergy by GAL4-VP16

Aya Fukuda,1,{dagger} Tomoyoshi Nakadai,1 Miho Shimada,1 Tohru Tsukui,1 Masahito Matsumoto,1 Yasuhisa Nogi,1 Michael Meisterernst,2 and Koji Hisatake1*

Department of Molecular Biology, Saitama Medical School, Moroyama, Iruma-gun, Saitama 350-0495, Japan,1 Gene Expression, Institute of Molecular Immunology, GSF-National Research Center for Environment and Health, Munich, Germany2

Received 24 December 2003/ Returned for modification 18 January 2004/ Accepted 14 April 2004

Positive cofactor 4 (PC4) is a coactivator that strongly augments transcription by various activators, presumably by facilitating the assembly of the preinitiation complex (PIC). However, our previous observation of stimulation of promoter escape in GAL4-VP16-dependent transcription in the presence of PC4 suggested a possible role for PC4 in this step. Here, we performed quantitative analyses of the stimulatory effects of PC4 on initiation, promoter escape, and elongation in GAL4-VP16-dependent transcription and found that PC4 possesses the ability to stimulate promoter escape in response to GAL4-VP16 in addition to its previously demonstrated effect on PIC assembly. This stimulatory effect of PC4 on promoter escape required TFIIA and the TATA box binding protein-associated factor subunits of TFIID. Furthermore, PC4 displayed physical interactions with both TFIIH and GAL4-VP16 through its coactivator domain, and these interactions were regulated distinctly by PC4 phosphorylation. Finally, GAL4-VP16 and PC4 stimulated both initiation and promoter escape to similar extents on the promoters with three and five GAL4 sites; however, they stimulated promoter escape preferentially on the promoter with a single GAL4 site. These results provide insight into the mechanism by which PC4 permits multiply bound GAL4-VP16 to attain synergy to achieve robust transcriptional activation.

* Corresponding author. Mailing address: Department of Molecular Biology, Saitama Medical School, 38 Morohongo, Moroyama, Iruma-gun, Saitama 350-0495, Japan. Phone: 81-49-276-1490. Fax: 81-49-294-9751. E-mail: kojihisa{at}saitama-med.ac.jp.

{dagger} Present address: Laboratory of Biochemistry and Molecular Biology, The Rockefeller University, New York, NY 10021.

Molecular and Cellular Biology, July 2004, p. 6525-6535, Vol. 24, No. 14
0022-538X/04/$08.00+0     DOI: 10.1128/MCB.24.14.6525-6535.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.



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