Isolating human transcription factor targets by coupling chromatin immunoprecipitation and CpG island microarray analysis.

Previously, identification of promoters regulated by mammalian transcription factors has relied upon overexpression studies. Here we present the identification of a large set of promoters that are bound by E2F in physiological conditions. Probing a human CpG microarray with chromatin immunoprecipitated using an antibody to E2F4, we have identified 68 unique target loci; 15% are bidirectional promoters and 25% recruit E2F via a mechanism distinct from the defined consensus site. Interestingly, although E2F has been shown previously to regulate genes involved in cell cycle progression, many of the new E2F target genes encode proteins involved in DNA repair or recombination. We suggest that human CpG microarrays, in combination with chromatin immunoprecipitation, will allow rapid identification of target promoters for many mammalian transcription factors.

[1]  J. Nevins,et al.  Expression of transcription factor E2F1 induces quiescent cells to enter S phase , 1993, Nature.

[2]  P. Farnham,et al.  Mre11 Complex and DNA Replication: Linkage to E2F and Sites of DNA Synthesis , 2001, Molecular and Cellular Biology.

[3]  Michael Ruogu Zhang,et al.  Computer-assisted identification of cell cycle-related genes: new targets for E2F transcription factors. , 2001, Journal of molecular biology.

[4]  M. P. Holloway,et al.  Aberrant Regulation of Survivin by the RB/E2F Family of Proteins* , 2004, Journal of Biological Chemistry.

[5]  W. Lee,et al.  The transcription factor E2F-1 mediates the autoregulation of RB gene expression , 1994, Molecular and cellular biology.

[6]  John J. Wyrick,et al.  Genome-wide location and function of DNA binding proteins. , 2000, Science.

[7]  T. Huang,et al.  Methylation profiling of CpG islands in human breast cancer cells. , 1999, Human molecular genetics.

[8]  A. Levine,et al.  p53 and E2F-1 cooperate to mediate apoptosis. , 1994, Proceedings of the National Academy of Sciences of the United States of America.

[9]  W. Kaelin,et al.  Deregulated transcription factor E2F-1 expression leads to S-phase entry and p53-mediated apoptosis. , 1994, Proceedings of the National Academy of Sciences of the United States of America.

[10]  David Botstein,et al.  Promoter-specific binding of Rap1 revealed by genome-wide maps of protein–DNA association , 2001, Nature Genetics.

[11]  D. Botstein,et al.  Genomic binding sites of the yeast cell-cycle transcription factors SBF and MBF , 2001, Nature.

[12]  W. Lee,et al.  Deregulated expression of E2F-1 induces S-phase entry and leads to apoptosis , 1994, Molecular and cellular biology.

[13]  C. M. Chen,et al.  Dissecting complex epigenetic alterations in breast cancer using CpG island microarrays. , 2001, Cancer research.

[14]  Andrew J. Bannister,et al.  Rb targets histone H3 methylation and HP1 to promoters , 2001, Nature.

[15]  R. Young,et al.  A common set of gene regulatory networks links metabolism and growth inhibition. , 2004, Molecular cell.

[16]  S. Cross,et al.  Purification of CpG islands using a methylated DNA binding column , 1994, Nature Genetics.

[17]  P. Farnham,et al.  Identification of target genes of oncogenic transcription factors. , 1999, Proceedings of the Society for Experimental Biology and Medicine. Society for Experimental Biology and Medicine.

[18]  R. Spang,et al.  Role for E2F in Control of Both DNA Replication and Mitotic Functions as Revealed from DNA Microarray Analysis , 2001, Molecular and Cellular Biology.

[19]  S. Asai,et al.  Thymidylate Synthase Expression Correlates Closely with E 2 F 1 Expression in Colon Cancer 1 , 2000 .

[20]  J. Nevins,et al.  E2Fs link the control of G1/S and G2/M transcription , 2004, The EMBO journal.

[21]  P. Farnham,et al.  Exogenous E2F expression is growth inhibitory before, during, and after cellular transformation , 2000, Oncogene.

[22]  J. Nevins,et al.  E2F1 overexpression in quiescent fibroblasts leads to induction of cellular DNA synthesis and apoptosis , 1995, Journal of virology.

[23]  T. Volkert,et al.  E2F integrates cell cycle progression with DNA repair, replication, and G(2)/M checkpoints. , 2002, Genes & development.

[24]  A. Bird,et al.  Number of CpG islands and genes in human and mouse. , 1993, Proceedings of the National Academy of Sciences of the United States of America.

[25]  F. Christians,et al.  E2Fs regulate the expression of genes involved in differentiation, development, proliferation, and apoptosis. , 2001, Genes & development.

[26]  F. Oswald,et al.  The E2F transcription factor activates a replication-dependent human H2A gene in early S phase of the cell cycle , 1996, Molecular and cellular biology.

[27]  Michael Q. Zhang,et al.  Large-scale human promoter mapping using CpG islands , 2000, Nature Genetics.

[28]  J. Nevins,et al.  Cellular targets for activation by the E2F1 transcription factor include DNA synthesis- and G1/S-regulatory genes , 1995, Molecular and cellular biology.

[29]  E. Lam,et al.  An E2F‐binding site mediates cell‐cycle regulated repression of mouse B‐myb transcription. , 1993, The EMBO journal.

[30]  Michael Q. Zhang,et al.  Use of Chromatin Immunoprecipitation To Clone Novel E2F Target Promoters , 2001, Molecular and Cellular Biology.

[31]  Sridhar Hannenhalli,et al.  Promoter prediction in the human genome , 2001, ISMB.