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J Biol Chem, Vol. 274, Issue 15, 10339-10348, April 9, 1999

Two E2F Sites Control Growth-regulated and Cell Cycle-regulated Transcription of the Htf9-a/RanBP1 Gene through Functionally Distinct Mechanisms

Barbara Di Fiore, Giulia Guarguaglini, Antonella Palena, Ron M. Kerkhoven, René Bernards, and Patrizia Lavia

From the  Consiglio Nazionale delle Ricerche Centre of Evolutionary Genetics, c/o University La Sapienza, Via degli Apuli 4, 00185 Rome, Italy and the  Division of Molecular Carcinogenesis, The Netherlands Cancer Institute, Plesmanlaan 121, 1066 CX Amsterdam, The Netherlands

The gene encoding Ran-binding protein 1 (RanBP1) is transcribed in a cell cycle-dependent manner. The RanBP1 promoter contains two binding sites for E2F factors, named E2F-c, located proximal to the transcription start, and E2F-b, falling in a more distal promoter region. We have now induced site-directed mutagenesis in both sites. We have found that the distal E2F-b site, together with a neighboring Sp1 element, actively controls up-regulation of transcription in S phase. The proximal E2F-c site plays no apparent role in cycling cells yet is required for transcriptional repression upon growth arrest. Protein binding studies suggest that each E2F site mediates specific interactions with individual E2F family members. In addition, transient expression assays with mutagenized promoter constructs indicate that the functional role of each site is also dependent on its position relative to other regulatory elements in the promoter context. Thus, the two E2F sites play opposite genetic functions and control RanBP1 transcription through distinct molecular mechanisms.

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