1: Genes Dev. 2004 Dec 1;18(23):2929-40. Epub 2004 Nov 15. Identification of a Drosophila Myb-E2F2/RBF transcriptional repressor complex. Lewis PW, Beall EL, Fleischer TC, Georlette D, Link AJ, Botchan MR. Department of Molecular and Cell Biology, University of California, Berkeley, CA 94720-3204, USA. The Drosophila Myb complex has roles in both activating and repressing developmentally regulated DNA replication. To further understand biochemically the functions of the Myb complex, we fractionated Drosophila embryo extracts relying upon affinity chromatography. We found that E2F2, DP, RBF1, RBF2, and the Drosophila homolog of LIN-52, a class B synthetic multivulva (synMuv) protein, copurify with the Myb complex components to form the Myb-MuvB complex. In addition, we found that the transcriptional repressor protein, lethal (3) malignant brain tumor protein, L(3)MBT, and the histone deacetylase, Rpd3, associated with the Myb-MuvB complex. Members of the Myb-MuvB complex were localized to promoters and were shown to corepress transcription of developmentally regulated genes. These and other data now link together the Myb and E2F2 complexes in higher-order assembly to specific chromosomal sites for the regulation of transcription. PMID: 15545624 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 2: EMBO J. 2004 Nov 24;23(23):4615-26. Epub 2004 Oct 28. E2Fs link the control of G1/S and G2/M transcription. Zhu W, Giangrande PH, Nevins JR. Department of Molecular Genetics and Microbiology, Duke Institute for Genome Sciences and Policy, Duke University Medical Center, Durham, NC 27710, USA. Previous work has provided evidence for E2F-dependent transcription control of both G1/S- and G2/M-regulated genes. Analysis of the G2-regulated cdc2 and cyclin B1 genes reveals the presence of both positive- and negative-acting E2F promoter elements. Additional elements provide both positive (CCAAT and Myb) and negative (CHR) control. Chromatin immunoprecipitation assays identify multiple interactions of E2F proteins that include those previously shown to activate and repress transcription. We find that E2F1, E2F2, and E2F3 bind to the positive-acting E2F site in the cdc2 promoter, whereas E2F4 binds to the negative-acting site. We also find that binding of an activator E2F is dependent on an adjacent CCAAT site that is bound by the NF-Y transcription factor and binding of a repressor E2F is dependent on an adjacent CHR element, suggesting a role for cooperative interactions in determining both activation and repression. Finally, the kinetics of B-Myb interaction with the G2-regulated promoters coincides with the activation of the genes, and RNAi-mediated reduction of B-Myb inhibits expression of cyclin B1 and cdc2. The ability of B-Myb to interact with the cdc2 promoter is dependent on an intact E2F binding site. These results thus point to a role for E2Fs, together with B-Myb, which is an E2F-regulated gene expressed at G1/S, in linking the regulation of genes at G1/S and G2/M. PMID: 15510213 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 3: Proc Natl Acad Sci U S A. 2000 Jun 6;97(12):6481-6. CpG methylation as a mechanism for the regulation of E2F activity. Campanero MR, Armstrong MI, Flemington EK. Department of Cancer Immunology and AIDS, Harvard Medical School and Dana-Farber Cancer Institute, 44 Binney Street, Boston, MA, 02115, USA. Regulation of gene expression in mammals through methylation of cytosine residues at CpG dinucleotides is involved in the development and progression of tumors. Because many genes that are involved in the control of cell proliferation are regulated by members of the E2F family of transcription factors and because some E2F DNA-binding sites are methylated in vivo, we have investigated whether CpG methylation can regulate E2F functions. We show here that methylation of E2F elements derived from the dihydrofolate reductase, E2F1, and cdc2 promoters prevents the binding of all E2F family members tested (E2F1 through E2F5). In contrast, methylation of the E2F elements derived from the c-myc and c-myb promoters minimally affects the binding of E2F2, E2F3, E2F4, and E2F5 but significantly inhibits the binding of E2F1. Consistent with these studies, E2F3, but not E2F1, activates transcription through methylated E2F sites derived from the c-myb and c-myc genes whereas both E2F1 and E2F3 fail to transactivate a reporter gene that is under the control of a methylated dihydrofolate reductase E2F site. Together, these data illustrate a means through which E2F activity can be controlled. PMID: 10823896 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 4: Kokubyo Gakkai Zasshi. 1998 Jun;65(2):172-88. [Interaction of E2F/Rb family members with factor binding to co-repressor element on B-myb and E2F1 promoters] [Article in Japanese] Nakajima Y. Second Department of Oral and Maxillofacial Surgery, Faculty of Dentistry, Tokyo Medical and Dental University. The E2F transcription factor plays an important role in controlling the expression of genes required for cell cycle progression. The transcription of a number of these genes, including E2F1 and B-myb, is repressed in G0/early G1 at E2F DNA binding sites mediated by interaction of E2F with the Rb family member proteins. It was shown that a corepressor element CHR, which was originally identified in the B-myb promoter, is also responsible for the repression of the E2F1 promoter. The mutation of the CHR element adjacent to E2F sites leads to a derepression of the E2F1 promoter in quiescent cells. The CHR-mutated promoter is activated by the E2F family of proteins (E2F1, E2F2, E2F3, and E2F4) but unable to be repressed by any of the Rb family members (Rb, p107, and p130) to the level of the wild-type promoter activity in G0, indicating that the repression by the Rb family members is required for the corepressor element. Moreover, it was shown that a factor specifically bound to the CHR element is co-purified with E2F by DNA affinity purification and co-immunoprecipitated with E2F4 and the Rb family members. These results seggested that E2F and the Rb family member proteins regulate the transcription of the E2F1 and B-myb genes by associating with an additional corepressor protein. PMID: 9711036 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 5: Cell Growth Differ. 1994 Aug;5(8):789-99. Inhibition of cell growth by TGF beta 1 is associated with inhibition of B-myb and cyclin A in both BALB/MK and Mv1Lu cells. Satterwhite DJ, Aakre ME, Gorska AE, Moses HL. Department of Cell Biology, Vanderbilt University School of Medicine, Nashville, Tennessee 37232. The concept of positive and negative regulation of normal cellular growth by diffusible factors is well illustrated by the effects of epidermal growth factor and transforming growth factor beta 1 (TGF beta 1) on mouse keratinocytes (MK) and mink lung epithelial cells (Mv1Lu). MK and Mv1Lu are nontransformed cell lines that reversibly arrest at a point in late G1 in response to TGF beta 1. Previously, we have shown that expression of the protooncogene c-myc is induced upon epidermal growth factor stimulation of quiescent MK and Mv1Lu cells and that transcriptional suppression of c-myc by TGF beta 1 treatment is important in the TGF beta 1 growth inhibition pathway. Using epidermal growth factor-stimulated synchronized MK and Mv1Lu cells, we have investigated the mRNA expression of a large number of growth factor-inducible genes that are critical regulators of growth in G1 and at the G1/S transition. These genes, often found to be dysregulated in cancer, include transcription factors as well as cyclins and their associated kinases, that promote growth, and tumor suppressor genes, that inhibit growth. As reported here, TGF beta 1 significantly inhibited mRNA expression of B-myb and cyclin A in both cell lines, suggesting that these may be important common downstream targets in the growth inhibition pathway. In contrast, the expression patterns of cyclins D1 and D2 and the transcription factors E2F1 and E2F2 were unaffected in MK cells treated with TGF beta 1 but were significantly inhibited in TGF beta 1-treated Mv1Lu cells. We cite the evidence suggesting that the inhibition of B-myb and cyclin A may contribute to the late G1 arrest caused by TGF beta 1 and that these events may be linked through the actions of the product of the retinoblastoma susceptibility gene (Rb) or an Rb family member. PMID: 7986745 [PubMed - indexed for MEDLINE] ---------------------------------------------------------------