DOI: 10.1126/science.275.5299.523. Regulation of NF-κB by Cyclin-Dependent Kinases Associated with the p300 Coactivator

Regulation of NF-κB by Cyclin-Dependent Kinases Associated with the p300 Coactivator

10.1126/science.275.5299.523

Crossref journal-article

American Association for the Advancement of Science (AAAS)

Science (221)

Abstract

The nuclear factor κB (NF-κB) transcription factor is responsive to specific cytokines and stress and is often activated in association with cell damage and growth arrest in eukaryotes. NF-κB is a heterodimeric protein, typically composed of 50- and 65-kilodalton subunits of the Rel family, of which RelA(p65) stimulates transcription of diverse genes. Specific cyclin-dependent kinases (CDKs) were found to regulate transcriptional activation by NF-κB through interactions with the coactivator p300. The transcriptional activation domain of RelA(p65) interacted with an amino-terminal region of p300 distinct from a carboxyl-terminal region of p300 required for binding to the cyclin E-Cdk2 complex. The CDK inhibitor p21 or a dominant negative Cdk2, which inhibited p300-associated cyclin E-Cdk2 activity, stimulated κB-dependent gene expression, which was also enhanced by expression of p300 in the presence of p21. The interaction of NF-κB and CDKs through the p300 and CBP coactivators provides a mechanism for the coordination of transcriptional activation with cell cycle progression.

Bibliography

Perkins, N. D., Felzien, L. K., Betts, J. C., Leung, K., Beach, D. H., & Nabel, G. J. (1997). Regulation of NF-ÎºB by Cyclin-Dependent Kinases Associated with the p300 Coactivator. Science, 275(5299), 523â527.

Authors 6

Neil D. Perkins (first)
Lisa K. Felzien (additional)
Jonathan C. Betts (additional)
Kwanyee Leung (additional)
David H. Beach (additional)
Gary J. Nabel (additional)

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We thank D. Gschwend for secretarial assistance; N. Barrett and K. Carter for preparation of figures; G. Hannon for p21 and p16 plasmids; P. Hobart (Vical Inc.) for the VCL-1012 CMV expression plasmid; G. Enders J. LaBaer and E. Harlow for the dominant negative Cdk2 plasmid; and W. Kong Z. Yang S. Parker and other members of the Nabel lab for their advice and comments. N.D.P. is the recipient of a Scholar Award from the American Foundation for AIDS Research (AmFAR) in memory of James Hoese. Supported in part by NIH grant RO1 AI29179.

Dates

Type	When
Created	23 years, 1 month ago (July 27, 2002, 5:50 a.m.)
Deposited	1 year, 7 months ago (Jan. 12, 2024, 9:56 p.m.)
Indexed	57 minutes ago (Aug. 28, 2025, 1:03 a.m.)
Issued	28 years, 7 months ago (Jan. 24, 1997)
Published	28 years, 7 months ago (Jan. 24, 1997)
Published Print	28 years, 7 months ago (Jan. 24, 1997)

Funders 0

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BibTeX

@article{Perkins_1997, title={Regulation of NF-κB by Cyclin-Dependent Kinases Associated with the p300 Coactivator}, volume={275}, ISSN={1095-9203}, url={http://dx.doi.org/10.1126/science.275.5299.523}, DOI={10.1126/science.275.5299.523}, number={5299}, journal={Science}, publisher={American Association for the Advancement of Science (AAAS)}, author={Perkins, Neil D. and Felzien, Lisa K. and Betts, Jonathan C. and Leung, Kwanyee and Beach, David H. and Nabel, Gary J.}, year={1997}, month=jan, pages={523–527} }

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  "abstract": "<jats:p>The nuclear factor \u03baB (NF-\u03baB) transcription factor is responsive to specific cytokines and stress and is often activated in association with cell damage and growth arrest in eukaryotes. NF-\u03baB is a heterodimeric protein, typically composed of 50- and 65-kilodalton subunits of the Rel family, of which RelA(p65) stimulates transcription of diverse genes. Specific cyclin-dependent kinases (CDKs) were found to regulate transcriptional activation by NF-\u03baB through interactions with the coactivator p300. The transcriptional activation domain of RelA(p65) interacted with an amino-terminal region of p300 distinct from a carboxyl-terminal region of p300 required for binding to the cyclin E-Cdk2 complex. The CDK inhibitor p21 or a dominant negative Cdk2, which inhibited p300-associated cyclin E-Cdk2 activity, stimulated \u03baB-dependent gene expression, which was also enhanced by expression of p300 in the presence of p21. The interaction of NF-\u03baB and CDKs through the p300 and CBP coactivators provides a mechanism for the coordination of transcriptional activation with cell cycle progression.</jats:p>",
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      "first-page": "3389",
      "author": "Steinman R. A.",
      "unstructured": "Steinman R. A., et al., ibid., p. 3389.",
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    {
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      "first-page": "3391",
      "volume": "54",
      "author": "Michieli P.",
      "year": "1994",
      "unstructured": "Michieli P., et al., Cancer Res. 54, 3391 (1994).",
      "journal-title": "Cancer Res."
    },
    {
      "key": "e_1_3_1_10_2",
      "doi-asserted-by": "publisher",
      "DOI": "10.1016/0092-8674(86)90807-X"
    },
    {
      "key": "e_1_3_1_10_3",
      "first-page": "793",
      "volume": "2",
      "author": "Thevenin C.",
      "year": "1990",
      "unstructured": "Thevenin C., et al., New Biol. 2, 793 (1990);",
      "journal-title": "New Biol."
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    {
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      "first-page": "4943",
      "volume": "11",
      "author": "Baldwin A. S.",
      "year": "1991",
      "unstructured": "Baldwin A. S. , Azizkhan J. C. , Jensen D. E. , Beg A. A. , Coodly L. R., Mol. Cell. Biol. 11, 4943 (1991);",
      "journal-title": "Mol. Cell. Biol."
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    {
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      "first-page": "1442",
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      "volume": "261",
      "author": "Devary Y.",
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      "unstructured": "Devary Y. , Rosette C. , DiDonato J. A. , Karin M., Science 261, 1442 (1993).",
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    {
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      "doi-asserted-by": "crossref",
      "first-page": "211",
      "DOI": "10.1016/S0959-437X(05)80276-X",
      "volume": "2",
      "author": "Nolan G. P.",
      "year": "1992",
      "unstructured": "Nolan G. P., Baltimore D., Curr. Opin. Genet. Dev. 2, 211 (1992);",
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    {
      "key": "e_1_3_1_11_3",
      "doi-asserted-by": "crossref",
      "first-page": "1",
      "DOI": "10.1016/S0074-7696(08)61873-2",
      "volume": "143",
      "author": "Grilli M.",
      "year": "1993",
      "unstructured": "Grilli M. , Chiu J. J. , Lenardo M. J., Int. Rev. Cytol. 143, 1 (1993);",
      "journal-title": "Int. Rev. Cytol."
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    {
      "key": "e_1_3_1_11_4",
      "doi-asserted-by": "crossref",
      "first-page": "405",
      "DOI": "10.1146/annurev.cb.10.110194.002201",
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      "author": "Siebenlist U.",
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      "unstructured": "Siebenlist U. , Franzoso G. , Brown K., Annu. Rev. Cell Biol. 10, 405 (1994).",
      "journal-title": "Annu. Rev. Cell Biol."
    },
    {
      "key": "e_1_3_1_12_2",
      "doi-asserted-by": "crossref",
      "first-page": "70",
      "DOI": "10.1038/339070a0",
      "volume": "339",
      "author": "Griffin G. E.",
      "year": "1989",
      "unstructured": "Griffin G. E., Leung K., Folks T. M., Kunkel S., Nabel G. J., Nature 339, 70 (1989).",
      "journal-title": "Nature"
    },
    {
      "key": "e_1_3_1_13_2",
      "first-page": "6530",
      "volume": "13",
      "author": "Eck S. L.",
      "year": "1993",
      "unstructured": "Eck S. L., Perkins N. D., Carr D. P., Nabel G. J., Mol. Cell. Biol. 13, 6530 (1993).",
      "journal-title": "Mol. Cell. Biol."
    },
    {
      "key": "e_1_3_1_14_2",
      "doi-asserted-by": "crossref",
      "first-page": "711",
      "DOI": "10.1038/326711a0",
      "volume": "326",
      "author": "Nabel G.",
      "year": "1987",
      "unstructured": "Nabel G., Baltimore D., Nature 326, 711 (1987).",
      "journal-title": "Nature"
    },
    {
      "key": "e_1_3_1_15_2",
      "first-page": "1315",
      "volume": "13",
      "author": "Duckett C. S.",
      "year": "1993",
      "unstructured": "Duckett C. S., et al., Mol. Cell. Biol. 13, 1315 (1993).",
      "journal-title": "Mol. Cell. Biol."
    },
    {
      "key": "e_1_3_1_16_2",
      "doi-asserted-by": "crossref",
      "first-page": "704",
      "DOI": "10.1038/366704a0",
      "volume": "366",
      "author": "Serrano M.",
      "year": "1993",
      "unstructured": "Serrano M., Hannon G. J., Beach D., Nature 366, 704 (1993).",
      "journal-title": "Nature"
    },
    {
      "key": "e_1_3_1_17_2",
      "first-page": "257",
      "volume": "371",
      "author": "Hannon G. J.",
      "year": "1994",
      "unstructured": "Hannon G. J., Beach D., ibid. 371, 257 (1994);",
      "journal-title": "ibid."
    },
    {
      "key": "e_1_3_1_17_3",
      "doi-asserted-by": "crossref",
      "first-page": "1607",
      "DOI": "10.1002/j.1460-2075.1996.tb00505.x",
      "volume": "15",
      "author": "Suzuki T.",
      "year": "1996",
      "unstructured": "Suzuki T. , Kitao S. , Matsushime H. , Yoshida M., EMBO J. 15, 1607 (1996).",
      "journal-title": "EMBO J."
    },
    {
      "key": "e_1_3_1_18_2",
      "unstructured": "Perkins N. D. Felzien L. K. Nabel G. J. unpublished data."
    },
    {
      "key": "e_1_3_1_19_2",
      "doi-asserted-by": "crossref",
      "first-page": "776",
      "DOI": "10.1038/333776a0",
      "volume": "333",
      "author": "Leung K.",
      "year": "1988",
      "unstructured": "Leung K., Nabel G. J., Nature 333, 776 (1988).",
      "journal-title": "Nature"
    },
    {
      "key": "e_1_3_1_20_2",
      "doi-asserted-by": "crossref",
      "first-page": "2336",
      "DOI": "10.1073/pnas.86.7.2336",
      "volume": "86",
      "author": "Osborn L.",
      "year": "1989",
      "unstructured": "Osborn L., Kunkel S., Nabel G. J., Proc. Natl. Acad. Sci. U.S.A. 86, 2336 (1989).",
      "journal-title": "Proc. Natl. Acad. Sci. U.S.A."
    },
    {
      "key": "e_1_3_1_21_2",
      "first-page": "344",
      "volume": "11",
      "author": "Lin W.",
      "year": "1991",
      "unstructured": "Lin W., Culp L. A., Biotechniques 11, 344 (1991);",
      "journal-title": "Biotechniques"
    },
    {
      "key": "e_1_3_1_21_3",
      "doi-asserted-by": "crossref",
      "first-page": "6820",
      "DOI": "10.1128/jvi.68.10.6820-6823.1994",
      "volume": "68",
      "author": "Perkins N. D.",
      "year": "1994",
      "unstructured": "Perkins N. D. , Agranoff A. B. , Duckett C. S. , Nabel G. J., J. Virol. 68, 6820 (1994).",
      "journal-title": "J. Virol."
    },
    {
      "key": "e_1_3_1_22_2",
      "unstructured": "Nuclear protein extracts (\u223c125 \u03bcg) were incubated at 4\u00b0C for 2 hours with anti-RelA (2 \u03bcg) (sc-109X; Santa Cruz Biotechnology Santa Cruz CA) or control rabbit immunoglobulin G (IgG) in Dignam D buffer [J. D. Dignam R. M. Lebovitz R. G. Roeder Nucleic Acids Res. 11 1475 (1983)] containing 0.1% NP-40 (adjusted to a final NaCl concentration of 75 mM). Antibody-protein complexes were recovered by incubation for 1 hour with protein G-agarose (10 \u03bcl) and low-speed centrifugation. Sedimental material was washed three times with 0.5 ml of incubation buffer [20 mM Hepes (pH 7.9) 75 mM KCl 2.5 mM MgCl 2 1 mM dithiothreitol (DTT) 0.1% NP-40 0.5 mM phenylmethylsulfonyl fluoride (PMSF) aprotinin (1 \u03bcg/ml) pepstatin A (1 \u03bcg/ml) leupeptin (2 \u03bcg/ml) and 1 mM sodium vanadate]. For histone H1 kinase assays protein G-bound complexes were resuspended in 50 \u03bcl of kinase buffer [50 mM tris (pH 7.5) 10 mM MgCl 2 1 mM DTT and 0.5 mM PMSF]. Purified glutathione-S-transferase fusion protein GST-p21 (\u223c50 ng) or GST proteins eluted into kinase buffer containing 10 mM glutathione were added as indicated. The samples were incubated with 133 \u03bcCi of [\u03b3- 32 P]adenosine triphosphate (ATP) and 2 \u03bcg of purified histone H1 (Boehringer Mannheim) at 37\u00b0C for 15 min with frequent agitation. The reaction was terminated by the addition of SDS gel loading buffer and a sample was resolved on an SDS-polyacrylamide (15%) gel which was then dried and subjected to autoradiography. Cyclin-Cdk immunoprecipitations were done as described above for RelA using 100 or 600 \u03bcg of nuclear extract and 10 \u03bcg of agarose-conjugated antibodies from Santa Cruz [sc-163AC (Cdk2) sc-601AC (Cdk4) sc-054AC (Cdc2) sc-239AC (cyclin A) sc-245AC (cyclin B1) sc-246AC (cyclin D1) and sc-248AC (cyclin E)]. All antibodies to cyclins were mouse monoclonals (IgG1) the antibodies to Cdk2 and Cdk4 were rabbit polyclonal antibodies to Cdk peptides and the antibody to Cdc2 was a mouse monoclonal (IgG2a). Protein immunoblotting of RelA was done with a rabbit polyclonal antibody (sc-109 Santa Cruz)."
    },
    {
      "key": "e_1_3_1_23_2",
      "doi-asserted-by": "publisher",
      "DOI": "10.1126/science.8266103"
    },
    {
      "key": "e_1_3_1_24_2",
      "doi-asserted-by": "crossref",
      "first-page": "799",
      "DOI": "10.1016/0092-8674(94)90127-9",
      "volume": "77",
      "author": "Arany Z.",
      "year": "1994",
      "unstructured": "Arany Z., Sellers W. R., Livingston D. M., Eckner R., Cell 77, 799 (1994).",
      "journal-title": "Cell"
    },
    {
      "key": "e_1_3_1_25_2",
      "unstructured": "Immunoprecipitations and transfections of 293 cells were done as described (Figs. 1 and 2) with antibodies from Pharmingen (14991A for p300 in Fig. 3 A B and D) or Santa Cruz [sc-109 (RelA) sc-369 (CBP) and sc-584 (p300 in Fig. 3C)]. Proteins were eluted from RelA (Fig. 3B) by incubation at room temperature for 15 min in wash buffer (19) that contained 300 mM KCl and bovine serum albumin (1 mg/ml). Either purified rabbit IgG or mouse IgG2b was used as the control where appropriate. In Fig. 3D the p300 protein was immunoprecipitated from 200 \u03bcg of Jurkat nuclear extract and incubated with the indicated in vitro translated 35 S-labeled proteins (TNT rabbit coupled reticulocyte lysate system Promega). Purified mouse IgG2b was used as the control. Protein association experiments Bluescript RelA RelA/VP16 Sp1 NF-\u03baB1(p50) and pET RelA(1\u2013306) plasmids used for in vitro transcription-translation have been described [N. D. Perkins A. B. Agranoff E. Pascal G. J. Nabel Mol. Cell. Biol. 14 6570 (1994)]."
    },
    {
      "key": "e_1_3_1_26_2",
      "doi-asserted-by": "crossref",
      "first-page": "85",
      "DOI": "10.1038/374085a0",
      "volume": "374",
      "author": "Lundblad J. R.",
      "year": "1995",
      "unstructured": "Lundblad J. R., Kwok R. P. S., Laurance M. E., Harter M. L., Goodman R. H., Nature 374, 85 (1995);",
      "journal-title": "Nature"
    },
    {
      "key": "e_1_3_1_26_3",
      "doi-asserted-by": "crossref",
      "first-page": "4758",
      "DOI": "10.1002/j.1460-2075.1995.tb00157.x",
      "volume": "14",
      "author": "Bannister A. J.",
      "year": "1995",
      "unstructured": "Bannister A. J. and , Kouzarides T., EMBO J. 14, 4758 (1995);",
      "journal-title": "EMBO J."
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    {
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      "first-page": "1961",
      "volume": "12",
      "author": "Janknect R.",
      "year": "1996",
      "unstructured": "Janknect R. and , Nordheim A., Oncogene 12, 1961 (1996);",
      "journal-title": "Oncogene"
    },
    {
      "key": "e_1_3_1_26_5",
      "doi-asserted-by": "crossref",
      "first-page": "528",
      "DOI": "10.1101/gad.10.5.528",
      "volume": "10",
      "author": "Dai P.",
      "year": "1996",
      "unstructured": "Dai P., et al., Genes Dev. 10, 528 (1996).",
      "journal-title": "Genes Dev."
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    {
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      "first-page": "5389",
      "volume": "11",
      "author": "Yaciuk P.",
      "year": "1991",
      "unstructured": "Yaciuk P., Moran E., Mol. Cell. Biol. 11, 5389 (1991);",
      "journal-title": "Mol. Cell. Biol."
    },
    {
      "key": "e_1_3_1_27_3",
      "first-page": "1639",
      "volume": "8",
      "author": "Abraham S. E.",
      "year": "1993",
      "unstructured": "Abraham S. E. , Lobo S. , Yaciuk P. , Wang H. H. , Moran E., Oncogene 8, 1639 (1993);",
      "journal-title": "Oncogene"
    },
    {
      "key": "e_1_3_1_27_4",
      "doi-asserted-by": "crossref",
      "first-page": "869",
      "DOI": "10.1101/gad.8.8.869",
      "volume": "8",
      "author": "Eckner R.",
      "year": "1994",
      "unstructured": "Eckner R., et al., Genes Dev. 8, 869 (1994);",
      "journal-title": "Genes Dev."
    },
    {
      "key": "e_1_3_1_27_5",
      "first-page": "1733",
      "volume": "9",
      "author": "Banerjee A. C.",
      "year": "1994",
      "unstructured": "Banerjee A. C., et al., Oncogene 9, 1733 (1994);",
      "journal-title": "Oncogene"
    },
    {
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      "doi-asserted-by": "crossref",
      "first-page": "5451",
      "DOI": "10.1073/pnas.92.12.5451",
      "volume": "92",
      "author": "Missero C.",
      "year": "1995",
      "unstructured": "Missero C., et al., Proc. Natl. Acad. Sci. U.S.A. 92, 5451 (1995).",
      "journal-title": "Proc. Natl. Acad. Sci. U.S.A."
    },
    {
      "key": "e_1_3_1_28_2",
      "doi-asserted-by": "crossref",
      "first-page": "1310",
      "DOI": "10.1073/pnas.87.4.1310",
      "volume": "87",
      "author": "Ventura A. M.",
      "year": "1990",
      "unstructured": "Ventura A. M., Arens M. Q., Srinivasan A., Chinnadurai G., Proc. Natl. Acad. Sci. U.S.A. 87, 1310 (1990);",
      "journal-title": "Proc. Natl. Acad. Sci. U.S.A."
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    {
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      "doi-asserted-by": "crossref",
      "first-page": "476",
      "DOI": "10.1128/jvi.67.1.476-488.1993",
      "volume": "67",
      "author": "Wang H. G.",
      "year": "1993",
      "unstructured": "Wang H. G., et al., J. Virol. 67, 476 (1993);",
      "journal-title": "J. Virol."
    },
    {
      "key": "e_1_3_1_28_4",
      "first-page": "2907",
      "volume": "69",
      "author": "Song C.",
      "year": "1995",
      "unstructured": "Song C. , Loewenstein P. M. , Green M., ibid. 69, 2907 (1995);",
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    {
      "key": "e_1_3_1_28_5",
      "author": "Parker S. F.",
      "unstructured": "Parker S. F. , Felzien L. K. , Perkins N. D. , Imperiale M. J. , Nabel G. J., J. Virol., in press.",
      "journal-title": "J. Virol."
    },
    {
      "key": "e_1_3_1_29_2",
      "doi-asserted-by": "crossref",
      "first-page": "1093",
      "DOI": "10.1016/0092-8674(94)90039-6",
      "volume": "79",
      "author": "Roy R.",
      "year": "1994",
      "unstructured": "Roy R., et al., Cell 79, 1093 (1994);",
      "journal-title": "Cell"
    },
    {
      "key": "e_1_3_1_29_3",
      "doi-asserted-by": "crossref",
      "first-page": "283",
      "DOI": "10.1038/374283a0",
      "volume": "374",
      "author": "Shiekhattar R.",
      "year": "1995",
      "unstructured": "Shiekhattar R., et al., Nature 374, 283 (1995);",
      "journal-title": "Nature"
    },
    {
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      "first-page": "193",
      "author": "Liao S.-M.",
      "unstructured": "Liao S.-M., et al., ibid., p. 193;",
      "journal-title": "ibid."
    },
    {
      "key": "e_1_3_1_29_5",
      "first-page": "660",
      "author": "Gerber H.-P.",
      "unstructured": "Gerber H.-P., et al., ibid., p. 660.",
      "journal-title": "ibid."
    },
    {
      "key": "e_1_3_1_30_2",
      "doi-asserted-by": "crossref",
      "first-page": "791",
      "DOI": "10.1016/0092-8674(86)90845-7",
      "volume": "44",
      "author": "Gilmore T. D.",
      "year": "1986",
      "unstructured": "Gilmore T. D., Temin H. M., Cell 44, 791 (1986);",
      "journal-title": "Cell"
    },
    {
      "key": "e_1_3_1_30_3",
      "first-page": "1075",
      "volume": "67",
      "author": "Neri A.",
      "year": "1991",
      "unstructured": "Neri A., et al., ibid. 67, 1075 (1991).",
      "journal-title": "ibid."
    },
    {
      "key": "e_1_3_1_31_2",
      "doi-asserted-by": "publisher",
      "DOI": "10.1038/376348a0"
    },
    {
      "key": "e_1_3_1_31_3",
      "first-page": "1565",
      "volume": "12",
      "author": "Muraoka M.",
      "year": "1996",
      "unstructured": "Muraoka M., et al., Oncogene 12, 1565 (1996).",
      "journal-title": "Oncogene"
    },
    {
      "key": "e_1_3_1_32_2",
      "doi-asserted-by": "crossref",
      "first-page": "6705",
      "DOI": "10.1128/jvi.69.11.6705-6711.1995",
      "volume": "69",
      "author": "He J.",
      "year": "1995",
      "unstructured": "He J., et al., J. Virol. 69, 6705 (1995);",
      "journal-title": "J. Virol."
    },
    {
      "key": "e_1_3_1_32_3",
      "first-page": "6304",
      "author": "Jowett J. B. M.",
      "unstructured": "Jowett J. B. M., et al., ibid., p. 6304;",
      "journal-title": "ibid."
    },
    {
      "key": "e_1_3_1_32_4",
      "first-page": "2324",
      "volume": "70",
      "author": "Bartz S. R.",
      "year": "1996",
      "unstructured": "Bartz S. R. , Rogel M. E. , Emerman M., ibid. 70, 2324 (1996).",
      "journal-title": "ibid."
    },
    {
      "key": "e_1_3_1_33_2",
      "unstructured": "The antibodies to p300 used were 14991A (Pharmingen) and sc-584 (Santa Cruz); the antibodies to CBP TBP TAF II 250 TAF II 130 TFIIH p89 subunit Cdk2 cyclin A cyclin B1 and cyclin E were from Santa Cruz (sc-369 sc-204 sc-735 sc-736 sc-293 sc-163 sc-239AC sc-245AC and sc-248AC respectively). The p300 cDNAs were obtained by polymerase chain reaction (PCR) amplified with the EXPAND PCR kit (Boehringer) from polyadenylated RNA (0.5 \u03bcg) from human spleen (Clontech) with oligo(dT) as a primer and reverse transcriptase (Stratagene). A 5\u2032 fragment (sense 5\u2032-GCTAAGCTTCACCATGGCCGAGAATG- TGGTGGAACCGGGGCCG-3\u2032; antisense 5\u2032-CACAGATCTGATGCATCTTTCTTCCGCACTCTGTAC-3\u2032) as well as a 3\u2032 p300 fragment overlapping the internal Bgl II site (sense 5\u2032-ATCAGATCTGTGTCCTTCACCATGAGATCATCTGGC-3\u2032; antisense 5\u2032-GC- TAGATCTCTAGTGTATGTCTAGTGTACTCTGTG- AGAGG-3\u2032) were isolated and subcloned into pBluescript downstream of the T7 promoter."
    },
    {
      "key": "e_1_3_1_34_2",
      "unstructured": "We thank D. Gschwend for secretarial assistance; N. Barrett and K. Carter for preparation of figures; G. Hannon for p21 and p16 plasmids; P. Hobart (Vical Inc.) for the VCL-1012 CMV expression plasmid; G. Enders J. LaBaer and E. Harlow for the dominant negative Cdk2 plasmid; and W. Kong Z. Yang S. Parker and other members of the Nabel lab for their advice and comments. N.D.P. is the recipient of a Scholar Award from the American Foundation for AIDS Research (AmFAR) in memory of James Hoese. Supported in part by NIH grant RO1 AI29179."
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