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The CDK9-associated cyclins T1 and T2 exert opposite effects on HIV-1 Tat activity

Napolitano, Giuliana; Licciardo, Paolo; Gallo, Pasquale; Majello, Barbara; Giordano, Antonioa; Lania, Luigi

Basic Science: Original Papers

Objectives: To examine the functional interaction between HIV-1 Tat protein and the cyclin T1 and T2 proteins which, in association with cyclin dependent kinase (CDK)9, are the regulatory subunits of the TAK/P-TEFb cellular complex strictly required for Tat transactivation.

Design: HIV-1 long terminal repeat (LTR) reporter plasmid was co-transfected into human and rodent cells with expression vectors encoding Tat and vectors encoding the cyclins T1, T2a and T2b, respectively.

Methods: Tat-mediated transactivation of HIV-1 LTR-driven transcription was compared in the presence or absence of different cyclins T (T1, T2a and T2b), upon co-transfections into human and rodent cell lines. Protein interactions were analysed by in vitro binding assays.

Results: It was found that Tat function in rodent cells is enhanced by co-expression of cyclin T1 but not cyclin T2. The N-terminal region (amino acids 1-290) of cyclin T1 is sufficient for this function and for binding to Tat and CDK9. Cyclin T2 binds to CDK9 but not to Tat. Moreover, enforced expression of cyclin T2 inhibits cyclin T1-mediated enhancement of Tat in rodent cells and it represses Tat activity in human cells.

Conclusion: Efficient Tat transactivation in rodent cells occurs in the presence of human cyclin T1 but not in the presence of cyclin T2; overexpression of cyclin T2 inhibits Tat function in both rodent and human cells.

From the Department of Genetics, Molecular and General Biology, University of Naples ‚Federico II‚, and International Institute of Genetics and Biophysics, CNR, Naples, Italy and the aDepartment of Pathology, Anatomy and Cell Biology, Sbarro Institute for Cancer Research and Molecular Medicine, Jefferson Medical College, Philadelphia, USA.

Sponsorship: Supported by grants from the Istituto Superiore di Sanità, Programma Nazionale di Ricerca AIDS (Grant 40A.0.57), MURST (Cofin) and Italian Association for Cancer Research (AIRC) to L.L., and in part from the Sbarro Institute for Cancer Research and Molecular Medicine, NIH grants R01 CA 60999-01A1, PO1 NS 36466, PO1CA 56309-06 to A.G.

Requests for reprints to: L. Lania, Dipartimento di Genetica, Biologia Generale e Molecolare, University of Naples ‚Federico II‚, Via Mezzocannone 8, 80134 Naples, Italy.

Received: 4 March 1999; revised: 26 April 1999; accepted: 4 May 1999.

© 1999 Lippincott Williams & Wilkins, Inc.