TRF2 associates with DREF and directs promoter-selective gene expression in Drosophila

Abstract

Drosophila TATA-box-binding protein (TBP)-related factor 2 (TRF2) is a member of a family of TBP-related factors present in metazoan organisms. Recent evidence suggests that TRF2s are required for proper embryonic development and differentiation1,2,3,4,5. However, true target promoters and the mechanisms by which TRF2 operates to control transcription remain elusive. Here we report the antibody affinity purification of a Drosophila TRF2-containing complex that contains components of the nucleosome remodelling factor (NURF) chromatin remodelling complex as well as the DNA replication-related element (DRE)-binding factor DREF. This latter finding led us to potential target genes containing TRF2-responsive promoters. We have used a combination of in vitro and in vivo assays to show that the DREF-containing TRF2 complex directs core promoter recognition of the proliferating cell nuclear antigen (PCNA) gene. We also identified additional TRF2-responsive target genes involved in DNA replication and cell proliferation. These data suggest that TRF2 functions as a core promoter-selectivity factor responsible for coordinating transcription of a subset of genes in Drosophila.

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Figure 1: Purification of a Drosophila TRF2 complex by co-immunoprecipitation polyacrylamide gradient gel electrophoresis (PAGE) a, Co-immunoprecipitation using anti-TRF2 and anti-DREF monoclonal antibodies.
Figure 2: The PCNA tandem promoters use two distinct transcription start sites designated start site 1 (+1) and start site 2 (-63).
Figure 3: The PCNA tandem promoters are synergistically activated by TRF2, DREF and E2F in SL2 cells.
Figure 4: Microarray analysis of TRF2-responsive target genes.
Figure 5: Double-stranded RNA interference in SL2 cells and promoter occupancy of TRF2 and DREF.

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Acknowledgements

We thank M. Haggart and R. Bell for technical assistance; D. Fyodorov and J. Kadonaga for providing antibodies raised against Acf, ISWI and p55/NURF-55; F. Hirose for generously providing Hybridoma cell lines producing anti-DREF antibody; T. Orr-Weaver for generously providing antibody raised against E2F prior to publication; E. Morris and N. Dyson for providing E2F and DP plasmids; H. Xiao and C. Wu for analysing NURF-301 for peptide sequence matches before publication; and Y. Isogai for providing affinity purified antibodies for ChIPs. We also thank Y. Berghöfer-Hochheimer, R. Freiman, B. Glover, Y. Isogai, T. Kutzkey and B. Lemon for comments on the manuscript, and all members in our laboratory for discussions and technical advice. A.H. was in part supported by a research fellowship by the Deutsche Forschungsgemeinschaft (DFG).

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Correspondence to Robert Tjian.

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Hochheimer, A., Zhou, S., Zheng, S. et al. TRF2 associates with DREF and directs promoter-selective gene expression in Drosophila. Nature 420, 439–445 (2002). https://doi.org/10.1038/nature01167

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