Cis-acting elements required for strong stop acceptor template selection during Moloney murine leukemia virus reverse transcription.
Template switching is required during normal retroviral DNA synthesis and is involved in retroviral genetic recombination. The first strong stop template switch during Moloney murine leukemia virus reverse transcription was studied to examine how template switch acceptor templates are selected. Retroviral vectors with specific alterations in their template switch acceptor regions were constructed, and DNA products templated by these vectors during a single replication cycle were analyzed. The results indicated that maximizing complementarity between the primer strand 3' end and the acceptor template was not the most significant factor in determining a strong stop template switch site. Instead, preferential transfer to the U3/R junction was observed, with as little as one contiguous base-pair of complementarity between the primer terminus and the template strand sufficient to direct template switching to the U3/R junction. These findings suggest that, in contrast to prevailing dogma, a base-pairing-independent mechanism functions in the specific guidance of retroviral strong stop template switch to the U3/R junction. Certain template alterations 3' of the template switch site were at least as disruptive to acceptor template use as was primer-terminal mismatch, suggesting that template structure or primer strand-internal sequences are important determinants of acceptor template selection. We discuss the implications of these findings for the mechanisms of retroviral DNA synthesis and homologous recombination.
Topping
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Department of Microbiology and Immunology, University of Michigan Medical School, Ann Arbor, MI, 48109-0620, USA.
Demoitie
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MA
Shin
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NH
Telesnitsky
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A
D016475:3T3 Cells; D000818:Animals; D001483:Base Sequence; D004279:DNA, Viral / Q000096:biosynthesis / Q000235:genetics; D005816:Genetic Complementation Test; D005822:Genetic Vectors; D051379:Mice; D008969:Molecular Sequence Data; D008979:Moloney murine leukemia virus / Q000235:genetics* / Q000502:physiology; D009154:Mutation; D009690:Nucleic Acid Conformation; D012367:RNA, Viral / Q000737:chemistry / Q000235:genetics; D011995:Recombination, Genetic; D014158:Transcription, Genetic; D014779:Virus Replication
D004279:DNA, Viral; D012367:RNA, Viral