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J. Virol., Jul 1997, 5178-5188, Vol 71, No. 7
J Guo, LE Henderson, J Bess, B Kane and JG Levin
During the first strand transfer in reverse transcription, minus-strand
strong-stop DNA [(-) SSDNA] is annealed to the 3' end of the acceptor RNA
in a reaction mediated by base-pairing between terminal repeat sequences in
the RNA and their complement in the DNA. The large stem- loop structure in
the repeat region known as TAR could interfere with this annealing
reaction. We have developed an in vitro human immunodeficiency virus type 1
(HIV-1) system to investigate the effect of TAR on strand transfer.
Mutational analysis demonstrates that the presence of TAR in the donor and
acceptor templates inhibits strand transfer and is correlated with
extensive synthesis of heterogeneous DNAs formed by self-priming from (-)
SSDNA. These DNAs are not precursors to the transfer product.
Interestingly, products of self- priming are not detected in HIV-1
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 efficiency of strand transfer.
In addition, the data suggest that the ability to eliminate self-priming is
a general property of NC which is manifested during reverse transcriptase
pausing at sites of secondary structure in the template. We conclude that
this activity of NC is critical for achieving highly efficient and specific
viral DNA synthesis. Our findings raise the possibility that inactivation
of NC could provide a new approach for targeting reverse transcription in
anti-HIV therapy.
Copyright © 1997, American Society for Microbiology
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
Laboratory of Molecular Genetics, National Institute of Child Health and Human Development, Bethesda, Maryland 20892, USA.
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