MilliporeSigma
  • The allosteric HIV-1 integrase inhibitor BI-D affects virion maturation but does not influence packaging of a functional RNA genome.

The allosteric HIV-1 integrase inhibitor BI-D affects virion maturation but does not influence packaging of a functional RNA genome.

PloS one (2014-07-30)
Nikki van Bel, Yme van der Velden, Damien Bonnard, Erwann Le Rouzic, Atze T Das, Richard Benarous, Ben Berkhout
ABSTRACT

The viral integrase (IN) is an essential protein for HIV-1 replication. IN inserts the viral dsDNA into the host chromosome, thereby aided by the cellular co-factor LEDGF/p75. Recently a new class of integrase inhibitors was described: allosteric IN inhibitors (ALLINIs). Although designed to interfere with the IN-LEDGF/p75 interaction to block HIV DNA integration during the early phase of HIV-1 replication, the major impact was surprisingly found on the process of virus maturation during the late phase, causing a reverse transcription defect upon infection of target cells. Virus particles produced in the presence of an ALLINI are misformed with the ribonucleoprotein located outside the virus core. Virus assembly and maturation are highly orchestrated and regulated processes in which several viral proteins and RNA molecules closely interact. It is therefore of interest to study whether ALLINIs have unpredicted pleiotropic effects on these RNA-related processes. We confirm that the ALLINI BI-D inhibits virus replication and that the produced virus is non-infectious. Furthermore, we show that the wild-type level of HIV-1 genomic RNA is packaged in virions and these genomes are in a dimeric state. The tRNAlys3 primer for reverse transcription was properly placed on this genomic RNA and could be extended ex vivo. In addition, the packaged reverse transcriptase enzyme was fully active when extracted from virions. As the RNA and enzyme components for reverse transcription are properly present in virions produced in the presence of BI-D, the inhibition of reverse transcription is likely to reflect the mislocalization of the components in the aberrant virus particle.

MATERIALS
Product Number
Brand
Product Description

Supelco
Dehydrated Alcohol, Pharmaceutical Secondary Standard; Certified Reference Material
USP
Dehydrated Alcohol, United States Pharmacopeia (USP) Reference Standard
Supelco
Ethanol solution, certified reference material, 2000 μg/mL in methanol
Sigma-Aldrich
Magnesium chloride solution, BioUltra, for molecular biology, ~1 M in H2O
Sigma-Aldrich
Urea-12C, 99.9 atom % 12C
Sigma-Aldrich
Magnesium chloride solution, BioUltra, for molecular biology, 2 M in H2O
Sigma-Aldrich
Urea solution, BioUltra, ~8 M in H2O
Sigma-Aldrich
Magnesium chloride solution, BioUltra, for molecular biology, ~0.025 M in H2O
Sigma-Aldrich
Magnesium chloride solution, PCR Reagent, 25 mM MgCI2 solution for PCR
Sigma-Aldrich
Ethanol Fixative 80% v/v, suitable for fixing solution (blood films)
Supelco
Ethanol standards 10% (v/v), 10 % (v/v) in H2O, analytical standard
Sigma-Aldrich
Urea solution, 40 % (w/v) in H2O
Sigma-Aldrich
Magnesium chloride solution, for molecular biology, 1.00 M±0.01 M
Millipore
Urea solution, suitable for microbiology, 40% in H2O
Sigma-Aldrich
Magnesium chloride solution, 0.1 M
Sigma-Aldrich
Ethylenediaminetetraacetic acid, BioUltra, ≥99.0% (KT)
Sigma-Aldrich
Ethylenediaminetetraacetic acid disodium salt solution, BioUltra, for molecular biology, pH 8.0, ~0.5 M in H2O
Sigma-Aldrich
L-Glutamine, meets USP testing specifications, suitable for cell culture, 99.0-101.0%, from non-animal source
Sigma-Aldrich
L-Glutamine, ReagentPlus®, ≥99% (HPLC)
Sigma-Aldrich
Magnesium chloride, suitable for insect cell culture, BioReagent, ≥97.0%
Sigma-Aldrich
Magnesium chloride, anhydrous, ≥98%
Sigma-Aldrich
Ethylenediaminetetraacetic acid solution, 0.02% in DPBS (0.5 mM), sterile-filtered, BioReagent, suitable for cell culture
Sigma-Aldrich
Urea, ReagentPlus®, ≥99.5%, pellets
Sigma-Aldrich
Urea, BioXtra, pH 7.5-9.5 (20 °C, 5 M in H2O)
Sigma-Aldrich
Urea, meets USP testing specifications
Sigma-Aldrich
Urea, powder, BioReagent, for molecular biology, suitable for cell culture
Sigma-Aldrich
Ethylenediaminetetraacetic acid, BioUltra, anhydrous, ≥99% (titration)
Sigma-Aldrich
Ethylenediaminetetraacetic acid, ACS reagent, 99.4-100.6%, powder
Sigma-Aldrich
Ethylenediaminetetraacetic acid, purified grade, ≥98.5%, powder
Sigma-Aldrich
Ethylenediaminetetraacetic acid, anhydrous, crystalline, BioReagent, suitable for cell culture