Cross-linking localization of a HIV-1 reverse transcriptase peptide involved in the binding of primer tRNALys3

Dufour, Emmanuelle; Reinbolt, Joseph; Castroviejo, Michel; Ehresmann, Bernard; Litvak, Simón; Tarragó-Litvak, Laura; Andréola, Marie‐Line

doi:10.1006/jmbi.1998.2430

Cited by 24 publications

(22 citation statements)

References 41 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…One group found a 127-amino acid cyanogen bromide fragment of RT (amino acids 230-357, containing all of the thumb subdomain flanked by small regions from the palm and connection subdomains) was cross-linked to a synthetic tRNA Lys3 thiolated at base 36 in the anticodon loop (16). Another group found an interaction between the 3 terminus of purified bovine liver tRNA Lys3 and an RT peptide whose amino terminus contains the conserved amino acid sequence 241VQPI244, which is found at the junction of the palm and thumb subdomains of RT (17 ). These in vitro experiments support the in vivo findings that the thumb region of RT within Pr160 gag-pol is important during tRNA Lys packaging, but whether it interacts with the anticodon arm or the 3 terminus of tRNA Lys3 is not clear, and the different conclusions obtained could be due to the use of a synthetic tRNA Lys3 versus fully modified beef liver tRNA Lys3 .…”

Section: Viral Proteins Involved In Trna Lys Packagingmentioning

confidence: 99%

tRNALys3: The Primer tRNA for Reverse Transcription in HIV‐1

Kleiman

2002

IUBMB Life

View full text Add to dashboard Cite

show abstract

Section: Viral Proteins Involved In Trna Lys Packagingmentioning

confidence: 99%

tRNALys3: The Primer tRNA for Reverse Transcription in HIV‐1

Kleiman

2002

IUBMB Life

View full text Add to dashboard Cite

show abstract

“…One obvious candidate is the reverse transcriptase (RT) polymerase, and a functional analysis of mutant HIV-1 virion particles revealed that RT is indeed involved in the selection and PBS annealing of the tRNA 3 Lys primer (45,57). Biochemical studies have provided additional information on the RT-tRNA 3 Lys complex and its involvement with the RNase H domain (17,54,56,58), but no high-resolution picture has emerged from these studies.…”

mentioning

confidence: 99%

Forced Selection of a Human Immunodeficiency Virus Type 1 Variant That Uses a Non-Self tRNA Primer for Reverse Transcription: Involvement of Viral RNA Sequences and the Reverse Transcriptase Enzyme

Abbink

Beerens

Berkhout

2004

J Virol

View full text Add to dashboard Cite

Human immunodeficiency virus type 1 uses the tRNA 3Lys molecule as a selective primer for reverse transcription. This primer specificity is imposed by sequence complementarity between the tRNA primer and two motifs in the viral RNA genome: the primer-binding site (PBS) and the primer activation signal (PAS). In addition, there may be specific interactions between the tRNA primer and viral proteins, such as the reverse transcriptase (RT) enzyme. We constructed viruses with mutations in the PAS and PBS that were designed to employ the nonself primer tRNA Pro or tRNA 1,2 Lys . These mutants exhibited a severe replication defect, indicating that additional adaptation of the mutant virus is required to accommodate the new tRNA primer. Multiple independent virus evolution experiments were performed to select for fast-replicating variants. Reversion to the wild-type PBS-lys3 sequence was the most frequent escape route. However, we identified one culture in which the virus gained replication capacity without reversion of the PBS. This revertant virus eventually optimized the PAS motif for interaction with the nonself primer. Interestingly, earlier evolution samples revealed a single amino acid change of an otherwise well-conserved residue in the RNase H domain of the RT enzyme, implicating this domain in selective primer usage. We demonstrate that both the PAS and RT mutations improve the replication capacity of the tRNA 1,2 Lys -using virus.

show abstract

“…In HIV-1, primer placement on the viral genome and primer packaging may depend on destabilization of the secondary structure of the tRNA and the viral RNA template by the p55 gag precursor (1,5,8,12,17,18,26,32,39,40), which also seems to be the case for avian leukosis virus, but not for MLV (15). Primer and RT interaction may also be necessary for initiation of reverse transcription (1,2,11,42,50), although RT may recognize the overall L-shaped tRNA rather than specific features of its cognate tRNA primer (45). Thus, we cannot exclude that one or several of the possible non-PBS interactions may contribute to the observed higher titers in SIV for the tRNA 3 Lys backbone in this study; however, the primary determinant for primer specificity is the complementarity between the viral PBS and the tRNA primer in SIVmac239.…”

mentioning

confidence: 99%

Transfer of Primer Binding Site-Mutated Simian Immunodeficiency Virus Vectors by Genetically Engineered Artificial and Hybrid tRNA-Like Primers

Hansen

Grünwald

Lund

et al. 2001

J Virol

View full text Add to dashboard Cite

Simian immunodeficiency viruses (SIV) harbor primer binding sites (PBS) matching tRNA 3Lys or tRNA 5 Lys . To study determinants of primer usage in SIV, a SIVmac239-based vector was impaired by mutating the PBS to a sequence (PBS-X2) with no match to any tRNA. By cotransfection of a synthetic gene encoding a tRNA Pro -like RNA with a match to PBS-X2, the activity of this vector could be restored to a transduction efficiency slightly lower than that of the wild-type vector. A vector with a PBS matching tRNA Pro was functional at a level slightly below that of the wild-type vector, but higher transduction efficiency could be obtained by cotransfection of a gene for an engineered tRNA Pro -tRNA 3 Lys hybrid with a match to PBS-Pro. The importance of tRNA backbone identity was further analyzed by complementing the PBS-X2 vector with a gene for a matching x2 primer with a tRNA 3 Lys backbone, which led to three-to fourfold-higher titers than those observed for the x2 primer with the tRNA Pro backbone. In summary, our results demonstrate flexibility in PBS and primer usage for SIVmac239, with PBS-primer complementarity being the major determinant, in analogy with previous findings for murine leukemia viruses and human immunodeficiency virus type 1.Retroviruses replicate through reverse transcription of the viral RNA genome into a double-stranded DNA form that is integrated into the genome of the host. The virus-encoded reverse transcriptase (RT) enzyme mediates this process through the use of a host-encoded tRNA. The 3Ј-terminal 18-nucleotide (nt) segment of the tRNA primer molecule anneals to the RNA genome at a complementary sequence known as the primer binding site (PBS). In order to synthesize a double-stranded DNA copy, two template shifts are involved. In plus-strand synthesis, the 3Ј 18-nt segment of the tRNA primer is reverse transcribed into a DNA copy, which mediates the second template shift by annealing to the minus-strand

show abstract

Cross-linking localization of a HIV-1 reverse transcriptase peptide involved in the binding of primer tRNALys3

Cited by 24 publications

References 41 publications

tRNALys3: The Primer tRNA for Reverse Transcription in HIV‐1

tRNALys3: The Primer tRNA for Reverse Transcription in HIV‐1

Forced Selection of a Human Immunodeficiency Virus Type 1 Variant That Uses a Non-Self tRNA Primer for Reverse Transcription: Involvement of Viral RNA Sequences and the Reverse Transcriptase Enzyme

Transfer of Primer Binding Site-Mutated Simian Immunodeficiency Virus Vectors by Genetically Engineered Artificial and Hybrid tRNA-Like Primers

Contact Info

Product

Resources

About