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Complementarity between 3' terminal nucleotides of tRNA and primer binding site is a major determinant for selection of the tRNA primer used for initiation of HIV-1 reverse transcription.

Abstract
The initiation of reverse transcription of human immunodeficiency virus type 1 (HIV-1) exclusively utilizes tRNALys,3 as a primer. Previous studies have shown that HIV-1 could use alternative tRNAs, such as tRNAIle or tRNAHis, to initiate reverse transcription only if the primer binding site (PBS) was made complementary to the 3' terminal 18 nucleotides of the cognate tRNA. However, upon in vitro culture, the viruses with a PBS complementary to the alternative tRNAs rapidly reverted to generate a PBS complementary to tRNALys,3. To investigate the process of reversion, we have constructed defective proviral genomes that contain a PBS complementary to tRNAIle or tRNAHis. The genomes contain the gene for xanthine-guanosine phosphoribosyl transferase (gpt) in place of env. Cotransfection of these proviral genomes with a plasmid-encoding vesicular stomatitis virus G protein (VSV-G) results in viruses that undergo a single round of HIV-1 infection; successful infections are scored as cells resistant to the drug mycophenolic acid. Using this single-round infection system, we demonstrated that HIV-1 with a PBS complementary to tRNAIle or tRNAHis is three- to fivefold less efficient in replication as measured by production of drug-resistant cell colonies compared to the wild-type virus. These viruses predominantly used the cognate tRNA as primer in their initial round of replication, although we did obtain a single cell colony in which the PBS was complementary to tRNALys,3. Using an HIV-1 provirus with a PBS complementary to yeast tRNAPhe, we established a single-round infection system in which the infectivity of this mutant HIV-1 relies on transfected yeast tRNAPhe. The results of our studies suggest that the mechanism for selection of the tRNA primer for initiation of reverse transcription relies primarily on the complementarity between the tRNA primerthe PBS.
AuthorsQ Yu, C D Morrow
JournalVirology (Virology) Vol. 254 Issue 1 Pg. 160-8 (Feb 01 1999) ISSN: 0042-6822 [Print] United States
PMID9927583 (Publication Type: Journal Article, Research Support, U.S. Gov't, P.H.S.)
CopyrightCopyright 1999 Academic Press.
Chemical References
  • RNA primers
  • RNA, Fungal
  • RNA, Transfer, His
  • RNA, Transfer, Ile
  • RNA, Transfer, Phe
  • RNA, Viral
  • RNA
  • RNA, Transfer
Topics
  • Binding Sites
  • Cell Line, Transformed
  • Genetic Complementation Test
  • Genome, Viral
  • HIV-1 (genetics)
  • HeLa Cells
  • Humans
  • Proviruses (genetics)
  • RNA
  • RNA, Fungal
  • RNA, Transfer
  • RNA, Transfer, His
  • RNA, Transfer, Ile
  • RNA, Transfer, Phe
  • RNA, Viral
  • Saccharomyces cerevisiae
  • Transcription, Genetic

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