Natural Single-Nucleotide Polymorphisms in the 3′ Region of the HIV-1
            <i>pol</i>
            Gene Modulate Viral Replication Ability

Nomaguchi, Masako; Miyake, Ariko; Doi, Naoya; Fujiwara, Sachi; Miyazaki, Yasuaki; Tsunetsugu-Yokota, Yasuko; Yokoyama, Masahiro; Sato, Hironori; Masuda, Tohru; Adachi, Akio

doi:10.1128/jvi.01859-13

Cited by 25 publications

(52 citation statements)

References 56 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Subsequent virological studies of these mutations showed that in human lymphocyte cell lines, growth phenotypes of not only prototype macaque cell-tropic HIV-1s but also human cell-tropic HIV-1 (NL4-3 clone) were markedly altered by the introduction of naturally occurring single-nucleotide variations (Y226tac, D229gat, P233cct, and V234gtg) into SA1prox ( Fig. 2A) (6). Since the common and unique characteristics of these variants relative to the parental clones were their altered vif mRNA levels (6), we determined the expression levels of the Vif protein for each NL4-3 derivative.…”

Section: Resultsmentioning

confidence: 99%

“…1A) modulated viral replication ability. Furthermore, we found that these variations in SA1prox affected HIV-1 mRNA expression patterns, especially its vif mRNA levels, thereby suggesting a role of this region in viral gene expression (6).…”

mentioning

confidence: 81%

“…The expression levels of viral and cellular A3G proteins were determined as described previously (6). Briefly, HEK 293T cells were cotransfected with proviral clones (3.5 g) and the pGL3 luciferase reporter vector (0.5 g) (Promega Corporation, Madison, WI) by using Lipofectamine 2000 (Life Technologies Corporation).…”

Section: Proviral Clonesmentioning

confidence: 99%

“…Northern blot analysis was performed to determine vif mRNA expression levels as previously described (6). Briefly, HEK 293T cells were transfected with proviral clones (2.5 g), and total RNA was extracted at 20 h posttransfection by using the RNeasy Plus minikit (Qiagen GmbH).…”

Section: Proviral Clonesmentioning

confidence: 99%

“…For qRT-PCR analysis, samples used for semiquantitative RT-PCR analysis were subjected to qRT-PCR analysis by using the QuantiTect Probe PCR kit (Qiagen GmbH), using the primer pairs for qRT-PCR analysis of viral transcripts described above. Tat trans-activation assays were performed as described previously (6). Briefly, Proviral clones and an LTR-driven luciferase reporter clone, 5RLTR-Luc (6), were cotransfected into HEK 293T cells with Lipofectamine 2000 (Life Technologies Corporation).…”

Section: Proviral Clonesmentioning

confidence: 99%

See 4 more Smart Citations

Natural Single-Nucleotide Variations in the HIV-1 Genomic SA1prox Region Can Alter Viral Replication Ability by Regulating Vif Expression Levels

Nomaguchi

Doi

Sakai

et al. 2016

J Virol

Self Cite

View full text Add to dashboard Cite

We previously found that natural single-nucleotide variations located within a proximal region of splicing acceptor 1 (SA1prox) in the HIV-1 genome could alter the viral replication potential and mRNA expression pattern, especially the vif mRNA level. Here, we studied the virological and molecular basis of nucleotide sequence variations in SA1prox for alterations of viral replication ability. Consistent with our previous findings, variant clones indeed expressed Vif at different levels and grew distinctively in cells with various APOBEC3G expression levels. Similar effects were observed for natural variations found in HIV-2 SA1prox, suggesting the importance of the SA1prox sequence. To define nucleotides critical for the regulation of HIV-1 Vif expression, effects of natural SA1prox variations newly found in the HIV Sequence Compendium database on vif mRNA/Vif protein levels were examined. Seven out of nine variations were found to produce Vif at lower, higher, or more excessive levels than wild-type NL4-3. Combination experiments of variations giving distinct Vif levels suggested that the variations mutually affected vif transcript production. While low and high producers of Vif grew in an APOBEC3G-dependent manner, excessive expressers always showed an impeded growth phenotype due to defects in single-cycle infectivity and/or virion production levels. The phenotype of excessive expressers was not due primarily to inadequate expression of Tat or Rev, although SA1prox variations altered the overall HIV-1 mRNA expression pattern. Collectively, our results demonstrate that HIV SA1prox regulates Vif expression levels and suggest a relationship between SA1prox and viral adaptation/evolution given that variations occurred naturally. IMPORTANCEWhile human cells possess restriction factors to inhibit HIV-1 replication, HIV-1 encodes antagonists to overcome these barriers. Conflicts between host restriction factors and viral counterparts are critical driving forces behind mutual evolution. The interplay of cellular APOBEC3G and viral Vif proteins is a typical example. Here, we demonstrate that naturally occurring single-nucleotide variations in the proximal region of splicing acceptor 1 (SA1prox) of the HIV-1 genome frequently alter Vif expression levels, thereby modulating viral replication potential in cells with various ABOBEC3G levels. The results of the present study reveal a previously unidentified and important way for HIV-1 to compete with APOBEC3G restriction by regulating its Vif expression levels. We propose that SA1prox plays a regulatory role in Vif counteraction against APOBEC3G in order to contribute to HIV-1 replication and evolution, and this may be applicable to other primate lentiviruses.

show abstract

Section: Resultsmentioning

confidence: 99%

mentioning

confidence: 81%

Section: Proviral Clonesmentioning

confidence: 99%

Section: Proviral Clonesmentioning

confidence: 99%

Section: Proviral Clonesmentioning

confidence: 99%

See 3 more Smart Citations