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J Biol Chem, Vol. 274, Issue 48, 34020-34028, November 26, 1999

Transcriptional Regulation of the Transforming Growth Factor-2 Promoter by cAMP-responsive Element-binding Protein (CREB) and Activating Transcription Factor-1 (ATF-1) Is Modulated by Protein Kinases and the Coactivators p300 and CREB-binding Protein

Michelle L. Kingsley-Kallesen^§, David Kelly, and Angie Rizzino^§

From the  Eppley Institute for Research in Cancer and Allied Diseases and the ^§ Department of Pathology and Microbiology, University of Nebraska Medical Center, Omaha, Nebraska 68198-6805

Transcription of the transforming growth factor-2 (TGF-2) gene is dependent on a cAMP-response element/activating transcription factor (CRE/ATF) site that is bound by CREB and ATF-1 as well as an E-box motif that is bound by upstream stimulatory factors 1 and 2 (USF1 and USF2). To identify additional factors involved in the expression of the TGF-2 gene, we employed F9 embryonal carcinoma (EC) cells, which express TGF-2 only after the cells differentiate. We show that overexpression of the transcription factors, CREB, ATF-1, USF1, and USF2 dramatically increases TGF-2 promoter activity in F9-differentiated cells. We further show that the coactivators p300 and CBP up-regulate the TGF-2 promoter when CREB and ATF-1 are expressed in conjunction with protein kinases that phosphorylate CREB on serine 133 and ATF-1 on serine 63. Importantly, we identify the presence of serine 133-phosphorylated CREB in the nucleus of F9-differentiated cells but not in the nucleus of F9 EC cells. This phosphorylated form is present in whole cell extracts of both the parental and differentiated cells, suggesting that nuclear accumulation of serine 133-phosphorylated CREB is regulated during differentiation of F9 EC cells and is likely to play an important role in the activation of the TGF-2 gene.

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