Nuclear localization of DP and E2F transcription factors by heterodimeric partners and retinoblastoma protein family members.

E2F is a family of transcription factors implicated in the regulation of genes required for progression through G1 and entry into the S phase. The transcriptionally active forms of E2F are heterodimers composed of one polypeptide encoded by the E2F gene family and one polypeptide encoded by the DP gene family. The transcriptional activity of E2F/DP heterodimers is influenced by association with the members of the retinoblastoma tumor suppressor protein family (pRb, p107, and p130). Here the intracellular distribution of E2F and DP proteins was investigated in transiently transfected Chinese hamster and human cells. In transfected cells, DP-1 did not accumulate in the nucleus unless it was coexpressed with the heterodimeric partners E2F-1, E2F-2, or E2F-3. Domain mapping experiments showed that regions of E2F-1 and DP-1 that are required for stable association of the two proteins were also required for nuclear localization of DP-1. Unlike E2F-1, -2, and -3, E2F-4 did not accumulate in the nucleus unless it was coexpressed with DP-2, p107 and p130, but not pRb, stimulated nuclear localization of E2F-4, either alone or in combination with DP-2. These results indicate that DP proteins preferentially associate with specific E2F partners, and suggest that the ability of specific E2F/DP heterodimers to localize in the nucleus contributes to the regulation of E2F activity.

[1]  N. L. La Thangue.  DP and E2F proteins: components of a heterodimeric transcription factor implicated in cell cycle control. , 1994, Current opinion in cell biology.

[2]  L. Johnston,et al.  Functional synergy between DP‐1 and E2F‐1 in the cell cycle‐regulating transcription factor DRTF1/E2F. , 1993, The EMBO journal.

[3]  M. Pagano,et al.  Association of cdk2 kinase with the transcription factor E2F during S phase. , 1992, Science.

[4]  M. Vidal,et al.  The retinoblastoma protein binds to a family of E2F transcription factors , 1993, Molecular and cellular biology.

[5]  J. Nevins,et al.  Interactions of the p107 and Rb proteins with E2F during the cell proliferation response. , 1993, The EMBO journal.

[6]  E. Moran.  DNA tumor virus transforming proteins and the cell cycle. , 1993, Current opinion in genetics & development.

[7]  P. Farnham,et al.  The role of E2F in the mammalian cell cycle. , 1993, Biochimica et biophysica acta.

[8]  L. J. Veer,et al.  E2F-5, a new E2F family member that interacts with p130 in vivo , 1995, Molecular and cellular biology.

[9]  W. Hong,et al.  Overexpression of E2F‐1 in rat embryo fibroblasts leads to neoplastic transformation. , 1994, The EMBO journal.

[10]  J. Nevins,et al.  The Rb-related p107 protein can suppress E2F function independently of binding to cyclin A/cdk2 , 1995, Molecular and cellular biology.

[11]  J. Hsuan,et al.  A new component of the transcription factor DRTF1/E2F , 1993, Nature.

[12]  J. Horowitz.  Regulation of transcription by the retinoblastoma protein , 1993, Genes, chromosomes & cancer.

[13]  S. Hiebert.  Regions of the retinoblastoma gene product required for its interaction with the E2F transcription factor are necessary for E2 promoter repression and pRb-mediated growth suppression , 1993, Molecular and cellular biology.

[14]  Chin-Lee Wu,et al.  Copyright � 1995, American Society for Microbiology In Vivo Association of E2F and DP Family Proteins , 1995 .

[15]  L. Zhu,et al.  E2F-4, a new member of the E2F gene family, has oncogenic activity and associates with p107 in vivo. , 1994, Genes & development.

[16]  W. Kaelin,et al.  E2F-1-mediated transactivation is inhibited by complex formation with the retinoblastoma susceptibility gene product. , 1993, Proceedings of the National Academy of Sciences of the United States of America.

[17]  S. Chellappan,et al.  Cloning and characterization of human DP2, a novel dimerization partner of E2F. , 1995, Oncogene.

[18]  Yang Xh,et al.  Overexpression of the E2F-1 transcription factor gene mediates cell transformation. , 1995 .

[19]  N. L. La Thangue,et al.  Transcriptional repression by the Rb-related protein p107. , 1993, Molecular biology of the cell.

[20]  Marc Vidal,et al.  A cDNA encoding a pRB-binding protein with properties of the transcription factor E2F , 1992, Cell.

[21]  D. Heimbrook,et al.  Cloning and characterization of E2F-2, a novel protein with the biochemical properties of transcription factor E2F , 1993, Molecular and cellular biology.

[22]  Nic Jones,et al.  Phosphorylation of E2F-1 modulates its interaction with the retinoblastoma gene product and the adenoviral E4 19 kDa protein , 1994, Cell.

[23]  J. Lawrence,et al.  E2F-4, a new member of the E2F transcription factor family, interacts with p107. , 1994, Genes & development.

[24]  W. White,et al.  Methotrexate-resistant Chinese hamster ovary cells have amplified a 135-kilobase-pair region that includes the dihydrofolate reductase gene. , 1981, Proceedings of the National Academy of Sciences of the United States of America.

[25]  W. Sellers,et al.  Transcription of the E2F-1 gene is rendered cell cycle dependent by E2F DNA-binding sites within its promoter , 1994, Molecular and cellular biology.

[26]  K. Helin,et al.  Heterodimerization of the transcription factors E2F-1 and DP-1 leads to cooperative trans-activation. , 1993, Genes & development.

[27]  D. Hall,et al.  Altered shape and cell cycle characteristics of fibroblasts expressing the E2F1 transcription factor. , 1994, Molecular biology of the cell.

[28]  J. Nevins,et al.  E2F: a link between the Rb tumor suppressor protein and viral oncoproteins. , 1992, Science.

[29]  M. Ewen,et al.  The transcription factor E2F-1 is a downstream target of RB action , 1995, Molecular and cellular biology.

[30]  R. Weinberg,et al.  Cell cycle-specific association of E2F with the p130 E1A-binding protein. , 1993, Genes & development.

[31]  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.

[32]  M. Ewen,et al.  Analysis of p107-associated proteins: p107 associates with a form of E2F that differs from pRB-associated E2F-1 , 1993, Journal of virology.

[33]  W. Lee,et al.  Molecular cloning of cellular genes encoding retinoblastoma-associated proteins: identification of a gene with properties of the transcription factor E2F , 1992, Molecular and cellular biology.

[34]  R. Weinberg,et al.  E2F-4 and E2F-5, two members of the E2F family, are expressed in the early phases of the cell cycle. , 1995, Proceedings of the National Academy of Sciences of the United States of America.

[35]  J. Nevins,et al.  The interaction of RB with E2F coincides with an inhibition of the transcriptional activity of E2F. , 1992, Genes & development.

[36]  Joseph R. Nevins,et al.  The E2F transcription factor is a cellular target for the RB protein , 1991, Cell.

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

[38]  M. Ewen,et al.  Inhibition of cell proliferation by p107, a relative of the retinoblastoma protein. , 1993, Genes & development.

[39]  P. Farnham,et al.  Multiple DNA elements are required for the growth regulation of the mouse E2F1 promoter. , 1994, Genes & development.

[40]  K. Wiman.  The retinoblastoma gene: role in cell cycle control and cell differentiation , 1993, FASEB journal : official publication of the Federation of American Societies for Experimental Biology.

[41]  N. Heintz,et al.  Protein-DNA interactions at the major and minor promoters of the divergently transcribed dhfr and rep3 genes during the Chinese hamster ovary cell cycle , 1996, Molecular and cellular biology.

[42]  J. Nevins,et al.  A cyclin A-protein kinase complex possesses sequence-specific DNA binding activity: p33cdk2 is a component of the E2F-cyclin A complex , 1992, Cell.

[43]  G. Wahl,et al.  Deficiency of retinoblastoma protein leads to inappropriate S-phase entry, activation of E2F-responsive genes, and apoptosis. , 1995, Proceedings of the National Academy of Sciences of the United States of America.

[44]  R. Müller,et al.  Transcriptional regulation during the mammalian cell cycle. , 1995, Trends in genetics : TIG.

[45]  V. Garsky,et al.  Protein domains governing interactions between E2F, the retinoblastoma gene product, and human papillomavirus type 16 E7 protein , 1993, Molecular and cellular biology.

[46]  L. Tsai,et al.  Independent binding of the retinoblastoma protein and p107 to the transcription factor E2F , 1992, Nature.

[47]  D. Livingston,et al.  Functional interaction between E2F-4 and p130: evidence for distinct mechanisms underlying growth suppression by different retinoblastoma protein family members. , 1995, Genes & development.

[48]  E. Lam,et al.  DP and E2F proteins: coordinating transcription with cell cycle progression. , 1994, Current opinion in cell biology.

[49]  David M. Livingston,et al.  Functional interactions of the retinoblastoma protein with mammalian D-type cyclins , 1993, Cell.

[50]  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.

[51]  J. Nevins,et al.  Oncogenic capacity of the E2F1 gene. , 1994, Proceedings of the National Academy of Sciences of the United States of America.

[52]  D. Heimbrook,et al.  Transcription factor E2F binds DNA as a heterodimer. , 1993, Proceedings of the National Academy of Sciences of the United States of America.

[53]  M. Ewen,et al.  Direct binding of cyclin D to the retinoblastoma gene product (pRb) and pRb phosphorylation by the cyclin D-dependent kinase CDK4. , 1993, Genes & development.

[54]  E. Harlow,et al.  The pRB‐related protein p107 contains two growth suppression domains: independent interactions with E2F and cyclin/cdk complexes. , 1995, The EMBO journal.

[55]  D. Livingston,et al.  Cell cycle analysis of E2F in primary human T cells reveals novel E2F complexes and biochemically distinct forms of free E2F , 1993, Molecular and cellular biology.

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

[57]  B. Dynlacht,et al.  Differential regulation of E2F transactivation by cyclin/cdk2 complexes. , 1994, Genes & development.

[58]  P. Farnham,et al.  Cloning, chromosomal location, and characterization of mouse E2F1 , 1994, Molecular and cellular biology.

[59]  C. Peng,et al.  Cyclin A/CDK2 binds directly to E2F-1 and inhibits the DNA-binding activity of E2F-1/DP-1 by phosphorylation , 1994, Molecular and cellular biology.

[60]  D. Livingston,et al.  Binding to DNA and the retinoblastoma gene product promoted by complex formation of different E2F family members. , 1993, Science.

[61]  W. Kaelin,et al.  RB associates with an E2F-like, sequence-specific DNA-binding protein. , 1991, Cold Spring Harbor symposia on quantitative biology.

[62]  S. Reed,et al.  Cyclin E/cdk2 and cyclin A/cdk2 kinases associate with p107 and E2F in a temporally distinct manner. , 1992, Genes & development.

[63]  T. Sladek,et al.  Overexpression of the E2F-1 transcription factor gene mediates cell transformation. , 1995, Gene expression.

[64]  F. Kaye,et al.  Differential specificity for binding of retinoblastoma binding protein 2 to RB, p107, and TATA-binding protein , 1994, Molecular and cellular biology.

[65]  D. Livingston,et al.  Multiple members of the E2F transcription factor family are the products of oncogenes. , 1995, Proceedings of the National Academy of Sciences of the United States of America.

[66]  M. Ewen,et al.  The transcription factor E2F interacts with the retinoblastoma product and a p107-cyclin A complex in a cell cycle-regulated manner , 1992, Cell.

[67]  N. Dyson.  pRB, p107 and the regulation of the E2F transcription factor , 1994, Journal of Cell Science.

[68]  Lingyun Zhu,et al.  Differential roles of two tandem E2F sites in repression of the human p107 promoter by retinoblastoma and p107 proteins , 1995, Molecular and cellular biology.

[69]  Wilhelm Krek,et al.  Negative regulation of the growth-promoting transcription factor E2F-1 by a stably bound cyclin A-dependent protein kinase , 1994, Cell.

[70]  J. Nevins,et al.  A genetic analysis of the E2F1 gene distinguishes regulation by Rb, p107, and adenovirus E4. , 1993, Molecular and cellular biology.

[71]  Peggy J. Farnham,et al.  Expression cloning of a cDNA encoding a retinoblastoma-binding protein with E2F-like properties , 1992, Cell.

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

[73]  J. Nevins,et al.  Cell cycle regulation of the E2F transcription factor involves an interaction with cyclin A , 1991, Cell.

[74]  J. Nevins,et al.  Autoregulatory control of E2F1 expression in response to positive and negative regulators of cell cycle progression. , 1994, Genes & development.