In vitro functional assessment of natural HIV-1 group M Vpu sequences using a universal priming approach

Asa Rahimi, Gursev Anmole, Maribel Soto-Nava, Tania Escamilla-Gomez, Tristan Markle, Steven W. Jin, Guinevere Q. Lee, P. Richard Harrigan, David Bangsberg, Jeffrey Martin, Santiago Avila-Rios, Gustavo Reyes-Teran, Mark A. Brockman, Zabrina L. Brumme

Research output: Contribution to journalArticle

2 Citations (Scopus)

Abstract

The HIV-1 accessory protein Vpu exhibits high inter- and intra- subtype genetic diversity that may influence Vpu function and possibly contribute to HIV-1 pathogenesis. However, scalable methods to evaluate genotype/phenotype relationships in natural Vpu sequences are limited, particularly those expressing the protein in CD4+ T-cells, the natural target of HIV-1 infection. A major impediment to assay scalability is the extensive genetic diversity within, and immediately upstream of, Vpu's initial 5′ coding region, which has necessitated the design of oligonucleotide primers specific for each individual HIV-1 isolate (or subtype). To address this, we developed two universal forward primers, located in relatively conserved regions 38 and 90 bases upstream of Vpu, and a single universal reverse primer downstream of Vpu, which are predicted to cover the vast majority of global HIV-1 group M sequence diversity. We show that inclusion of up to 90 upstream bases of HIV-1 genomic sequence does not significantly influence in vitro Vpu expression or function when a Rev/Rev Response Element (RRE)-dependent expression system is used. We further assess the function of four diverse HIV-1 Vpu sequences, revealing reproducible and significant differences between them. Our approach represents a scalable option to measure the in vitro function of genetically diverse natural Vpu isolates in a CD4+ T-cell line.

Original languageEnglish (US)
Pages (from-to)32-41
Number of pages10
JournalJournal of Virological Methods
Volume240
DOIs
StatePublished - Feb 1 2017
Externally publishedYes

Fingerprint

HIV-1
T-Lymphocytes
DNA Primers
Response Elements
In Vitro Techniques
HIV Infections
Proteins
Genotype
Phenotype
Cell Line

Keywords

  • BST2/tetherin
  • CD4
  • Downregulation
  • HIV-1 Vpu
  • Rev/RRE
  • Sequence diversity

ASJC Scopus subject areas

  • Virology

Cite this

Rahimi, A., Anmole, G., Soto-Nava, M., Escamilla-Gomez, T., Markle, T., Jin, S. W., ... Brumme, Z. L. (2017). In vitro functional assessment of natural HIV-1 group M Vpu sequences using a universal priming approach. Journal of Virological Methods, 240, 32-41. https://doi.org/10.1016/j.jviromet.2016.11.004

In vitro functional assessment of natural HIV-1 group M Vpu sequences using a universal priming approach. / Rahimi, Asa; Anmole, Gursev; Soto-Nava, Maribel; Escamilla-Gomez, Tania; Markle, Tristan; Jin, Steven W.; Lee, Guinevere Q.; Harrigan, P. Richard; Bangsberg, David; Martin, Jeffrey; Avila-Rios, Santiago; Reyes-Teran, Gustavo; Brockman, Mark A.; Brumme, Zabrina L.

In: Journal of Virological Methods, Vol. 240, 01.02.2017, p. 32-41.

Research output: Contribution to journalArticle

Rahimi, A, Anmole, G, Soto-Nava, M, Escamilla-Gomez, T, Markle, T, Jin, SW, Lee, GQ, Harrigan, PR, Bangsberg, D, Martin, J, Avila-Rios, S, Reyes-Teran, G, Brockman, MA & Brumme, ZL 2017, 'In vitro functional assessment of natural HIV-1 group M Vpu sequences using a universal priming approach', Journal of Virological Methods, vol. 240, pp. 32-41. https://doi.org/10.1016/j.jviromet.2016.11.004
Rahimi, Asa ; Anmole, Gursev ; Soto-Nava, Maribel ; Escamilla-Gomez, Tania ; Markle, Tristan ; Jin, Steven W. ; Lee, Guinevere Q. ; Harrigan, P. Richard ; Bangsberg, David ; Martin, Jeffrey ; Avila-Rios, Santiago ; Reyes-Teran, Gustavo ; Brockman, Mark A. ; Brumme, Zabrina L. / In vitro functional assessment of natural HIV-1 group M Vpu sequences using a universal priming approach. In: Journal of Virological Methods. 2017 ; Vol. 240. pp. 32-41.
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