Cell Cycle Regulation of E2F Site Occupation in Vivo

Zwicker, Jörk; Liu, Ningshu; Engeland, Kurt; Lucibello, Frances C.; Müller, Rolf

doi:10.1126/science.271.5255.1595

Cited by 148 publications

(129 citation statements)

References 26 publications

(6 reference statements)

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“…Interestingly, in the tsa cell lines it also contains p130, E2F4 and DP1 (data not shown). Since dl1135 T antigen was able to maintain the growth of tsa14 cells at 39.58C, but lacks a functional J domain, we undertook experiments to determine if its interaction with p130 was productive by measuring the levels of B-myb mRNA because transcription of the B-myb gene is E2F-dependent and transcriptionally downregulated by both p107 and p130 (Lam et al, 1994;Zwicker et al, 1996). B-myb mRNA was detected at high levels in tsa14, as well as tsa8 and tsa129 cells at 338C (Figure 3a).…”

Section: Dl1135 T Antigen Interacts Productively With P130mentioning

confidence: 99%

Different functions are required for initiation and maintenance of immortalization of rat embryo fibroblasts by SV40 large T antigen

et al. 1999

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We have used two di erent, but complementary assays to characterize functions of SV40 T antigen that are necessary for its ability to immortalize rat embryo ®broblasts. In accordance with previous work, we found that several functions were required. These include activities that map to the p53 binding domain and the amino terminal 176 amino acids which contain the J domain as well as the CR1 and CR2 domain required for binding and sequestering the RB family of pocket proteins. Moreover, we found that even though activities dependent only upon the amino terminus were su cient for immortalization they were unable to maintain it. This suggests that immortalization by these amino terminal functions requires either additional events or immortalization of a subset of cells within the heterogeneous rat embryo ®broblast population. We further found that an activity dependent upon amino acids 17 ± 27 which remove a portion of the CR1 domain and the predicted a-1 helix of the J domain was not necessary to maintain growth but was required for direct immortalization suggesting that at least one of the functions required initially was not required to maintain the immortal state. This represents the ®rst demonstration that some of the functions required for maintenance of the immortal state di er from those required for initiation of immortalization.

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Section: Dl1135 T Antigen Interacts Productively With P130mentioning

confidence: 99%

Different functions are required for initiation and maintenance of immortalization of rat embryo fibroblasts by SV40 large T antigen

et al. 1999

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“…Genes thought to be E2F targets include protooncogenes (Heibert et al, 1989;Lam and Watson, 1993), G1 cyclins (Muller et al, 1994;Henglein et al, 1994;Ohtani et al, 1995), enzymes required for DNA synthesis (Pearson et al, 1991;Means et al, 1992;Fridovich-Keil et al, 1993), and the G2-speci®c cdk, cdc2 (Dalton, 1992;Tommasi and Pfeifer, 1995). Several recent studies suggest that E2F may be primarily a negative regulator of transcription, directing the binding of repressors, such as pRb, to speci®c target genes (Tommasi and Pfeifer, 1995;Field et al, 1996;Yamasaki et al, 1996;Zwicker et al, 1996). This view is supported by the observation that mice bearing homozygous deletions of the E2F-1 gene do not have defects in cell proliferation, but rather show an increased tendency to develop tumors (Field et al, 1996;Yamasaki et al, 1996;Zwicker et al, 1996).…”

mentioning

confidence: 98%

“…Several recent studies suggest that E2F may be primarily a negative regulator of transcription, directing the binding of repressors, such as pRb, to speci®c target genes (Tommasi and Pfeifer, 1995;Field et al, 1996;Yamasaki et al, 1996;Zwicker et al, 1996). This view is supported by the observation that mice bearing homozygous deletions of the E2F-1 gene do not have defects in cell proliferation, but rather show an increased tendency to develop tumors (Field et al, 1996;Yamasaki et al, 1996;Zwicker et al, 1996). Nevertheless, once its association with pRb family proteins is disrupted, E2F also seems to be a positive regulator of certain genes, such as dhfr (Wells et al, 1997) and b-myb (Lam et al, 1995).…”

mentioning

confidence: 99%

pRb and p107 regulate E2F activity during lens fiber cell differentiation

et al. 1998

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During growth arrest and di erentiation, activity of the E2F family of transcription factors is inhibited by interactions with pRb and the related proteins, p107 and p130. To determine which members of the E2F and pRb families may contribute to growth arrest as lens epithelial cells di erentiate into ®ber cells, we examined the expression of individual E2F species and characterized the E2F protein complexes formed in rat lens epithelia and ®bers. RT/PCR detected all ®ve known members of the E2F family in lens epithelial cells, but only E2F-1, E2F-3, and E2F-5 in ®ber cells. Proteins extracted from lens epithelia of newborn rats formed at least two speci®c complexes with an E2F consensus oligonucleotide. Proteins from lens ®ber cells formed three speci®c complexes, one of which comigrated with an epithelial cell complex. Incubation of epithelial and ®ber cell extracts with an antibody speci®c for p107 demonstrated that two ®ber cell complexes and one epithelial cell complex contained p107. Although the remaining ®ber cell complex did not react with antibodies to pRb or p130 in this assay, a strong reaction with pRb antibody was observed when the electromobility shifted complexes were subsequently immunoblotted (shift/Western assay). Immunocytochemistry con®rmed that pRb protein is present in the nuclei of both epithelial cells and ®ber cells. Immunoblotting of whole cell extracts with pRb antibody showed multiple, phosphorylated forms of pRb in the epithelial cells, but predominantly hypophosphorylated pRb in the ®ber cells. None of the complexes formed with E2F were recognized exclusively by the p130 antibody, although the previously identi®ed p107 complexes reacted weakly. The absence of p130/E2F complexes was correlated with the presence of multiple ubiquitinated forms of p130, especially in the ®ber cells. Thus, although p130/E2F complexes are implicated in the terminal di erentiation of many cell types, in di erentiating lens ®ber cells pRb and p107 seem to be the primary regulators of E2F activity.

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“…E2F-1, E2F-3 or E2F-6) can bind the promoter during G0/G1 as components of repressive complexes. In this respect, it is surprising that results from both in vivo footprinting and ChIP analyses concur that the B-myb promoter E2F site is unoccupied in S phase (Zwicker et al, 1996;Takahashi et al, 2000;Wells et al, 2000), and this singular feature suggests that E2F species are prevented from binding at this stage of the cell cycle. Our previous findings suggest that the DRS motif adjacent to the E2F site contributes to this blockade .…”

Section: Discussionmentioning

confidence: 99%

“…Consistent with this, chromatin immunoprecipitation (ChIP) assays showed that the B-myb E2F site is occupied by E2F-4/p107 or E2F-4/p130 complexes during G0/G1. However other cell cycle-regulated promoters do not bind E2F during S phase (Zwicker et al, 1996;Takahashi et al, 2000;Wells et al, 2000). Binding of repressive E2F complexes to the B-myb E2F site is strongly facilitated by the adjacent downstream repression site (DRS) (Bennett et al, 1996;Catchpole et al, 2002).…”

Section: Introductionmentioning

confidence: 99%

Targeting an E2F site in the mouse genome prevents promoter silencing in quiescent and post-mitotic cells

2006

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Previous studies have shown that the cell cycle-regulated B-myb promoter contains a conserved E2F binding site that is critical for repressing transcription in quiescent cells. To investigate its significance for permanent promoter silencing, we have inactivated this binding site in the mouse genome. Mice homozygous for the mutant B-myb mE2F allele were fully viable, however, B-myb transcription was derepressed during quiescence in mouse embryo fibroblasts (MEFs) derived from mutant animals. Moreover, it was found that mutation of the E2F site resulted in abnormal maintenance of B-myb expression in senescent MEFs and in differentiated brain tissue. These findings therefore reveal a direct and primary role for repressive E2F complexes in silencing gene expression in post-mitotic cells. Analysis of histone modifications at the promoter showed that histone H3 lysine 9 was constitutively acetylated throughout the cell cycle in homozygous mutant MEFs. This mouse system is the first description of an E2F site mutation in situ and will facilitate the study of E2F function in vivo.

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Cell Cycle Regulation of E2F Site Occupation in Vivo

Cited by 148 publications

References 26 publications

Different functions are required for initiation and maintenance of immortalization of rat embryo fibroblasts by SV40 large T antigen

Different functions are required for initiation and maintenance of immortalization of rat embryo fibroblasts by SV40 large T antigen

pRb and p107 regulate E2F activity during lens fiber cell differentiation

Targeting an E2F site in the mouse genome prevents promoter silencing in quiescent and post-mitotic cells

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