Vpu of a Simian Immunodeficiency Virus Isolated from Greater Spot-Nosed Monkey Antagonizes Human BST-2 via Two AxxxxxxxW Motifs

Viral protein U (Vpu) is an accessory protein encoded by human immunodeficiency virus type 1 (HIV-1) and certain simian immunodeficiency virus (SIV) strains. Some of these viruses were reported to use Vpu to overcome restriction by BST-2 of their natural hosts. Our own recent report revealed that Vpu of SIVgsn-99CM71 (SIVgsn71) antagonizes human BST-2 through two AxxxxxxxW motifs (A 22 W 30 and A 25 W 33 ) whereas antagonizing BST-2 of its natural host, greater spot-nosed monkey (GSN), involved only A 22 W 30 motif. Here we show that residues A 22 , A 25 , W 30 , and W 33 of SIVgsn71 Vpu are all essential to antagonize human BST-2, while, neither single mutation of A 22 nor W 30 affected the ability to antagonize GSN BST-2. Similar to A 18 , which is located in the middle of the A 14 xxxxxxxW 22 motif in HIV-1 NL4-3 Vpu and is essential to antagonize human BST-2, A 29 , located in the middle of the A 25 W 33 motif of SIVgsn71 Vpu was found to be necessary for antagonizing human but not GSN BST-2. Further mutational analyses revealed that residues L 21 and K 32 of SIVgsn71 Vpu were also essential for antagonizing human BST-2. On the other hand, the ability of SIVgsn71 Vpu to target GSN BST-2 was unaffected by single amino acid substitutions but required multiple mutations to render SIVgsn71 Vpu inactive against GSN BST-2. These results suggest additional requirements for SIVgsn71 Vpu antagonizing human BST-2, implying evolution of the bst-2 gene under strong selective pressure. Importance Genes related to survival against life-threating pathogens are important determinants of natural selection in animal evolution. For instance, BST-2, a protein showing broad-spectrum antiviral activity, shows polymorphisms entailing different phenotypes even among primate species, suggesting that the bst-2 gene of primates has been subject to strong selective pressure during evolution. At the same time, viruses readily adapt to these evolutionary changes. Thus, we found that Vpu of an SIVgsn isolate (SIVgsn-99CM71) can target BST-2 from humans as well as from its natural host thus potentially facilitating zoonosis. Here we mapped residues in SIVgsn71 Vpu potentially contributing to cross-species transmission. We found that the requirements for targeting human BST-2 are distinct from and more complex than those for targeting GSN BST-2. Our results suggest that the human bst-2 gene might have evolved to acquire more restrictive phenotype than GSN bst-2 against viral proteins after being derived from their common ancestor.

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HIV-1 RNAs whose transcription initiates from the third deoxyguanosine of GGG tract in the 5′ long terminal repeat serve as a dominant genome for efficient provirus DNA formation

Yoshida,

Kasuya,

Yamamoto

et al. 2024

J Virol

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Unspliced HIV-1 RNAs function as messenger RNAs for Gag or Gag-Pol polyproteins and progeny genomes packaged into virus particles. Recently, it has been reported that fate of the RNAs might be primarily determined, depending on transcriptional initiation sites among three consecutive deoxyguanosine residues (GGG tract) downstream of TATA-box in the 5′ long terminal repeat (LTR). Although HIV-1 RNA transcription starts mostly from the first deoxyguanosine of the GGG tract and often from the second or third deoxyguanosine, RNAs beginning with one guanosine (G1-form RNAs), whose transcription initiates from the third deoxyguanosine, were predominant in HIV-1 particles. Despite selective packaging of G1-form RNAs into virus particles, its biological impact during viral replication remains to be determined. In this study, we revealed that G1-form RNAs are primarily selected as a template for provirus DNA rather than other RNAs. In competitions between HIV-1 and lentiviral vector transcripts in virus-producing cells, approximately 80% of infectious particles were found to generate provirus using HIV-1 transcripts, while lentiviral vector transcripts were conversely selected when we used HIV-1 mutants in which the third deoxyguanosine in the GGG tract was replaced with deoxythymidine or deoxycytidine (GGT or GGC mutants, respectively). In the other analyses of proviral sequences after infection with an HIV-1 mutant in which the GGG tract in 3′ LTR was replaced with TTT, most proviral sequences of the GGG-tract region in 5′ LTR were found to be TTG, which is reasonably generated using the G1-form transcripts. Our results indicate that the G1-form RNAs serve as a dominant genome to establish provirus DNA. IMPORTANCE Since the promoter for transcribing HIV-1 RNA is unique, all viral elements including genomic RNA and viral proteins have to be generated by the unique transcripts through ingenious mechanisms including RNA splicing and frameshifting during protein translation. Previous studies suggested a new mechanism for diversification of HIV-1 RNA functions by heterogeneous transcriptional initiation site usage; HIV-1 RNAs whose transcription initiates from a certain nucleotide were predominant in virus particles. In this study, we established two methods to analyze heterogenous transcriptional initiation site usage by HIV-1 during viral infection and showed that RNAs beginning with one guanosine (G1-form RNAs), whose transcription initiates from the third deoxyguanosine of the GGG tract in 5′ LTR, were primarily selected as viral genome in infectious particles and thus are used as a template to generate provirus for continuous replication. This study provides insights into the mechanism for diversification of unspliced RNA functions and requisites of lentivirus infectivity.

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Vpu of a Simian Immunodeficiency Virus Isolated from Greater Spot-Nosed Monkey Antagonizes Human BST-2 via Two AxxxxxxxW Motifs

Cited by 3 publications

References 58 publications

A Novel Vpu Adaptive Mutation of HIV-1 Degrades Tetherin in Northern Pig-Tailed Macaques (Macaca leonina) Mainly via the Ubiquitin-Proteasome Pathway and Increases Viral Release

A Novel Vpu Adaptive Mutation of HIV-1 Degrades Tetherin in Northern Pig-Tailed Macaques (Macaca leonina) Mainly via the Ubiquitin-Proteasome Pathway and Increases Viral Release

Simian Immunodeficiency Virus SIVgsn-99CM71 Vpu Employs Different Amino Acids To Antagonize Human and Greater Spot-Nosed Monkey BST-2

HIV-1 RNAs whose transcription initiates from the third deoxyguanosine of GGG tract in the 5′ long terminal repeat serve as a dominant genome for efficient provirus DNA formation

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