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Human immunodeficiency virus uses tRNA Lys,3 as primer for reverse transcription in HeLa‐CD4 + cells
Author(s) -
Das Atze T.,
Koken Sabine E.C.,
Oude Essink Belinda B.,
van Wamel Jeroen L.B.,
Berkhout Ben
Publication year - 1994
Publication title -
febs letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.593
H-Index - 257
eISSN - 1873-3468
pISSN - 0014-5793
DOI - 10.1016/0014-5793(94)80238-6
Subject(s) - primer binding site , primer (cosmetics) , transfer rna , reverse transcriptase , biology , microbiology and biotechnology , mutant , hela , transcription (linguistics) , rna , virus , virology , chemistry , genetics , cell culture , gene , linguistics , philosophy , organic chemistry
Significant amounts of different tRNA molecules are present in retroviral particles, but one specific tRNA species functions as primer in reverse transcription. It is generally believed that the HIV‐1 virus uses the tRNA Lys,3 molecule as primer. This is based on sequence complementarity between the 3' end of tRNA Lys,3 and the primer‐binding site (PBS) on HIV‐1 genomic RNA. Recent biochemical analyses indicated that tRNA Lys,3 is indeed incorporated into viral particles. Interestingly, tRNA Lys,3 could not be detected in virions produced by HeLa‐CD4 + cells [(1992) Biochem. Biophys. Res. Commun. 185, 1105‐1115]. In order to test whether alternative tRNA molecules can function as primer in HIV replication, we performed a series of experiments based on the observation that tRNA primer sequences are inherited by the viral progeny. We cultured HIV‐1 for prolonged periods of time in HeLa‐CD4 + cells, but did not detect sequence changes in the PBS region. Furthermore, we found PBS‐mutants to be replication incompetent, again suggesting that HIV‐1 solely uses tRNA Lys,3 as primer. Most importantly, we obtained revertants of one such PBS‐mutant, which had restored a wild‐type PBS sequence. This tRNA Lys,3 ‐mediated repair demonstrates a general requirement for this primer in HIV‐1 reverse transcription.