Discontinuous plus-strand DNA synthesis in human immunodeficiency virus type 1-infected cells and in a partially reconstituted cell-free system.

Important note: Information in this article was accurate in 1998. The state of the art may have changed since the publication date.

Discontinuous plus-strand DNA synthesis in human immunodeficiency virus type 1-infected cells and in a partially reconstituted cell-free system.

J Virol. 1997 Dec;71(12):9259-69. Unique Identifier : AIDSLINE MED/98037635
Klarmann GJ; Yu H; Chen X; Dougherty JP; Preston BD; Department of Biochemistry, Eccles Institute of Human Genetics and the; Huntsman Cancer Institute, University of Utah, Salt Lake City; 84112-5330, USA.

Abstract: Human immunodeficiency virus type 1 (HIV-1) replication requires conversion of viral RNA to double-stranded DNA. To better understand the molecular mechanisms of this process, we examined viral DNA synthesis in a simple cell-free system that uses the activities of HIV-1 reverse transcriptase to convert regions of single-stranded HIV-1 RNA to double-stranded DNA in a single incubation. This system recapitulated several of the required intermediate steps of viral DNA synthesis: RNA-templated minus-strand polymerization, preferential plus-strand initiation at the central and 3' HIV-1 polypurine tracts, and DNA-templated plus-strand polymerization. Secondary sites of plus-strand initiation were also observed at low frequency both in the cell-free system and in cultured virus. Direct comparison of viral and cell-free products revealed differences in the precision and selectivity of plus-strand initiation, suggesting that the cell-free system lacks one or more essential replication components. These studies provide clues about mechanisms of plus-strand initiation and serve as a starting point for the development of more complex multicomponent cell-free systems.

Keywords: *DNA, Viral/BIOSYNTHESIS *HIV-1/GENETICS
980330
M9831184

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