Transcriptional activation of the type II transforming growth factor-beta receptor gene upon differentiation of embryonal carcinoma cells.
Previously, it has been shown that differentiation of embryonal carcinoma (EC) cells turns on the expression of functional transforming growth factor type-beta receptors. Here, we show that the type II receptor (TbetaR-II) gene is activated at the transcriptional level when EC cells differentiate. We show that the differentiated cells, but not the parental EC cells, express transcripts for TbetaR-II. In addition, the expression of TbetaR-II promoter/reporter gene constructs are elevated dramatically when EC cells differentiate and we identify at least two positive and two negative regulatory regions in the 5' flanking region of the TbetaR-II gene. Moreover, we identify a cAMP response element/activating transcription factor site that acts as a positive cis-regulatory element in the TbetaR-II promoter, and we demonstrate that the transcription factor ATF-1 binds to this site and strongly stimulates the expression of the TbetaR-II promoter/reporter gene constructs when ATF-1 is overexpressed in EC-derived differentiated cells. Equally important, we identify a negative regulatory element in a 53-base pair region that had previously been shown to inhibit strongly the expression of TbetaR-II promoter/reporter gene constructs. Specifically, we demonstrate that this region, which contains an inverted CCAAT box motif, binds the transcription factor complex NF-Y (also referred to as CBF) in vitro. Furthermore, expression of a dominant-negative NF-YA mutant protein, which prevents DNA binding by NF-Y, enhances TbetaR-II promoter expression. Together, these studies suggest that the transcription factors ATF-1 and NF-Y play important roles in the regulation of the TbetaR-II gene.
Eppley Institute for Research in Cancer and Allied Diseases, University of Nebraska Medical Center, Omaha, Nebraska 68198, USA.
D
D
Kelly
S J
SJ
Kim
A
A
Rizzino
D051697:Activating Transcription Factor 1; D000818:Animals; D001483:Base Sequence; D022762:CCAAT-Enhancer-Binding Proteins; D018236:Carcinoma, Embryonal / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D002454:Cell Differentiation; D002460:Cell Line; D004247:DNA; D004268:DNA-Binding Proteins / Q000378:metabolism; D015972:Gene Expression Regulation, Neoplastic; D051379:Mice; D009687:Nuclear Proteins / Q000378:metabolism; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D017346:Protein Serine-Threonine Kinases; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D000077294:Receptor, Transforming Growth Factor-beta Type II; D018125:Receptors, Transforming Growth Factor beta / Q000235:genetics* / Q000378:metabolism; D012045:Regulatory Sequences, Nucleic Acid; D014157:Transcription Factors / Q000378:metabolism; D015533:Transcriptional Activation; D016212:Transforming Growth Factor beta / Q000378:metabolism; D014407:Tumor Cells, Cultured
D051697:Activating Transcription Factor 1; C497100:Atf1 protein, mouse; D022762:CCAAT-Enhancer-Binding Proteins; D004268:DNA-Binding Proteins; D009687:Nuclear Proteins; D012333:RNA, Messenger; D018125:Receptors, Transforming Growth Factor beta; D014157:Transcription Factors; D016212:Transforming Growth Factor beta; D004247:DNA; D017346:Protein Serine-Threonine Kinases; D000077294:Receptor, Transforming Growth Factor-beta Type II