RNase H activity is required for high-frequency repeat deletion during Moloney murine leukemia virus replication

J. L. Brincat, J. K. Pfeiffer, A. Telesnitsky

Research output: Contribution to journalArticle

26 Citations (Scopus)

Abstract

It has been postulated that retroviral recombination, like strong stop template switching, requires the RNase H activity of reverse transcriptase. To address this hypothesis, Moloney murine leukemia virus-based vectors, which were designed to test the recombination-related property of direct repeat deletion, were encapsidated in virions engineered to contain phenotypic mixtures of wild-type and RNase H catalytic site point mutant reverse transcriptase. Integrated provirus titers per milliliter were determined for these phenotypically mixed virions, and vector proviruses were screened to determine what percentage contained repeat deletions. The results revealed a steady decline in direct repeat deletion frequency that correlated with decreases in functional RNase H, with greater than fourfold decreases in repeat deletion frequency observed when 95% of virion reverse transcriptase was RNase H defective. Parallel experiments were performed to address effects of molar excesses of RNase H relative to functional DNA polymerase. These experiments demonstrated that increasing the stoichiometry of RNase H relative to the amount of functional DNA polymerase had minimal effects on direct repeat deletion frequency. DNA synthesis was error prone when directed principally by RNase H mutant reverse transcriptase, suggesting a role for RNase H catalytic integrity in the fidelity of intracellular reverse transcription.

Original languageEnglish (US)
Pages (from-to)88-95
Number of pages8
JournalJournal of Virology
Volume76
Issue number1
DOIs
StatePublished - 2002

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Murine leukemia virus
Moloney murine leukemia virus
Ribonuclease H
RNA-directed DNA polymerase
Virus Replication
virus replication
virion
proviruses
RNA-Directed DNA Polymerase
DNA-directed DNA polymerase
Nucleic Acid Repetitive Sequences
Virion
Proviruses
mutants
reverse transcription
stoichiometry
DNA-Directed DNA Polymerase
active sites
Genetic Recombination
synthesis

ASJC Scopus subject areas

  • Immunology

Cite this

RNase H activity is required for high-frequency repeat deletion during Moloney murine leukemia virus replication. / Brincat, J. L.; Pfeiffer, J. K.; Telesnitsky, A.

In: Journal of Virology, Vol. 76, No. 1, 2002, p. 88-95.

Research output: Contribution to journalArticle

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