Posttranscriptional Modification of Retroviral Primers is Required for Late Stages of DNA Replication

During reverse transcription of retroviral RNA, synthesis of (-) strand DNA is primed by a cellular tRNA that anneals to an 18-nt primer binding site within the 5′long terminal repeat. For (+) strand synthesis using a (-) strand DNA template linked to the tRNA primer, only the first 18 nt of tRNA ar...

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Bibliographic Details
Published in:Proceedings of the National Academy of Sciences - PNAS 1997-07, Vol.94 (14), p.7210-7215
Main Authors: Burnett, Bruce P., McHenry, Charles S.
Format: Article
Language:English
Subjects:
Binding sites
Biochemistry
Biological Sciences
Deoxyribonucleic acid
DNA
DNA Replication
DNA, Viral - genetics
HIV
human immunodeficiency virus
Oligonucleotides
Retroviridae - genetics
Reverse transcription
Ribonucleic acid
RNA
RNA - genetics
RNA Processing, Post-Transcriptional
RNA, Viral - genetics
Terminal repeat sequences
Transfer RNA
Viruses
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Summary:During reverse transcription of retroviral RNA, synthesis of (-) strand DNA is primed by a cellular tRNA that anneals to an 18-nt primer binding site within the 5′long terminal repeat. For (+) strand synthesis using a (-) strand DNA template linked to the tRNA primer, only the first 18 nt of tRNA are replicated to regenerate the primer binding site, creating the (+) strand strong stop DNA intermediate and providing a 3′terminus capable of strand transfer and further elongation. On model HIV templates that approximate the (-) strand linked to natural modified or synthetic unmodified tRNA3 Lys, we find that a (+) strand strong stop intermediate of the proper length is generated only on templates containing the natural, modified tRNA3 Lys, suggesting that a posttranscriptional modification provides the termination signal. In the presence of a recipient template, synthesis after strand transfer occurs only from intermediates generated from templates containing modified tRNA3 Lys. Reverse transcriptase from Moloney murine leukemia virus and avian myoblastosis virus shows the same requirement for a modified tRNA3 Lystemplate. Because all retroviral tRNA primers contain the same 1-methyl-A58modification, our results suggest that 1-methyl-A58is generally required for termination of replication 18 nt into the tRNA sequence, generating the (+) strand intermediate, strand transfer, and subsequent synthesis of the entire (+) strand. The possibility that the host methyl transferase responsible for methylating A58may provide a target for HIV chemotherapy is discussed.
ISSN:0027-8424
1091-6490
DOI:10.1073/pnas.94.14.7210