3rd Conference on Retroviruses and Opportunistic Infections Washington, DC - January 28-February 1, 1996

Conf Retroviruses Opportunistic Infect 1996 Jan 28-Feb 1; 3rd:57 (abstract no. 22)

Sire J¹, Bouhamdan M¹, Benichou S², Rey F¹, Navarro JM¹, Agostini I¹, Spire B¹, Slupphaug G³, Camonis J², Benarous R², Vigne R^1
1 INSERM-U372, BP178 13009 Marseille, France. ² INSERM-U372, ICGM 75014 Paris. ³ UNIGEN Trondheim-University N-7005 Trondheim, Norway

The Vpr gene product of HIV is a virion-associated regulatory protein that appears to be multifunctional. It is involved in the nuclear import of viral preintegration complexes, transactivation of viral and cellular genes, induction of cell differentiation and G2-arrest of the cell cycle. We have used the yeast two-hybrid system to identify cellular partner(s) interacting with Vpr. A Jurkat cDNA library constructed as a fusion with the Ga14-activation domain was screened with Vpr fused to the Ga14-DNA binding domain. A cDNA clone which specifically interacts with Vpr was isolated and sequence analysis revealed that it is homologous to the human uracil-DNA glycosylase (UNG). Confirmation of the interaction was carried out by in vitro studies using either GST-UNG fusion protein and ³⁵ S-labelled Vpr, or GST-Vpr and recombinant UNG. Co-immunoprecipitation experiments confirmed that Vpr and UNG are associated within cells expressing Vpr. We also demonstrate that the enzymatic activity of UNG is still retained upon interaction with Vpr. The UNG enzyme belongs to the DNA excision-repair enzyme family and removes uracil misincorporated during DNA synthesis or arisen by cytosine deamination. All non primate lentiviruses encode for a viral dUTPase to maintain a low dUTP:dTTP ratio in infected cells to minimize uracil incorporation into DNA. In the case of HIV-1, such a protein is not encoded by the viral genome. We have shown that the HIV-l Vpr protein could interact with the cellular UNG enzyme, and the question of its functional relevance remains to be investigated.

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