HUMAN-IMMUNODEFICIENCY-VIRUS TYPE-1 NUCLEOCAPSID PROTEIN PROMOTES EFFICIENT STRAND TRANSFER AND SPECIFIC VIRAL-DNA SYNTHESIS BY INHIBITING TAR-DEPENDENT SELF-PRIMING FROM MINUS-STRAND STRONG-STOP DNA

During the first strand transfer in reverse transcription, minus-stran d strong-stop DNA [(-) SSDNA] is annealed to the 3' end of the accepto r RNA in a reaction mediated by base-pairing between terminal repeat s equences in the RNA and their complement in the DNA, The large stem-lo op structure in the repeat region known as TAR could interfere with th is annealing reaction, We have developed an in vitro human immuno-defi ciency virus type 1 (HIV-1) system to investigate the effect of TAR on strand transfer, Mutational analysis demonstrates that the presence o f TAR in the donor and acceptor templates inhibits strand transfer and is correlated with extensive synthesis of heterogeneous DNAs formed b y self-priming from (-) SSDNA. These DNAs art not precursors to the tr ansfer product. Interestingly, products of self-priming are not detect ed in NN-I endogenous reactions; this suggests that virions contain a component which prevents self-priming. Our results show that the viral nucleocapsid protein (NC), which can destabilize secondary structures , drastically reduces self-priming and dramatically increases the effi ciency of strand transfer, In addition, the data suggest that the abil ity to eliminate self-priming is a general property of NC which is man ifested during reverse transcriptase pausing at sites of secondary str ucture in the template. We conclude that this activity of NC is critic al for achieving highly efficient and specific viral DNA synthesis. Oa r findings raise the possibility that inactivation of NC could provide a new approach for targeting reverse transcription in anti-HIV therap y.