The cellular protein hnRNP A2/B1 enhances HIV-1 transcription by unfolding LTR promoter G-quadruplexes

Abstract

G-quadruplexes are four-stranded conformations of nucleic acids that act as cellular epigenetic regulators. A dynamic G-quadruplex forming region in the HIV-1 LTR promoter represses HIV-1 transcription when in the folded conformation. This activity is enhanced by nucleolin, which induces and stabilizes the HIV-1 LTR G-quadruplexes. In this work by a combined pull-down/mass spectrometry approach, we consistently found hnRNP A2/B1 as an additional LTR-G-quadruplex interacting protein. Surface plasmon resonance confirmed G-quadruplex specificity over linear sequences and fluorescence resonance energy transfer analysis indicated that hnRNP A2/B1 is able to efficiently unfold the LTR G-quadruplexes. Evaluation of the thermal stability of the LTR G-quadruplexes in different-length oligonucleotides showed that the protein is fit to be most active in the LTR full-length environment. When hnRNP A2/B1 was silenced in cells, LTR activity decreased, indicating that the protein acts as a HIV-1 transcription activator. Our data highlight a tightly regulated control of transcription based on G-quadruplex folding/unfolding, which depends on interacting cellular proteins. These findings provide a deeper understanding of the viral transcription mechanism and may pave the way to the development of drugs effective against the integrated HIV-1, present both in actively and latently infected cells.

X-Linked RNA-Binding Motif Protein Modulates HIV-1 Infection of CD4 + T Cells by Maintaining the Trimethylation of Histone H3 Lysine 9 at the Downstream Region of the 5′ Long Terminal Repeat of HIV Proviral DNA

“…Repression of gene transcriptional represents a main mechanism by which HIV-1 latency is maintained (6). Multiply nuclear factors, including hnRNPs, such as hnRNPA2/ B1, hnRNP U, have been shown to modulate the transcription of latently infected HIV-1 (39)(40)(41). We have previously reported that one hnRNP family member, SAFB1 (scaffold attachment factor B), is capable of inhibiting HIV-1 transcription by preventing the recruitment of RNA pol II to HIV-1 LTR (29).…”

G-quadruplex forming sequences in the genome of all known human viruses: A comprehensive guide

“…A G4 was also detected in hepatitis B virus (HBV) genome, the only member of dsDNA viruses with RT activity [27]. Finally, functionally significant G4s were identified both in the RNA and DNA proviral genome of the human immunodeficiency virus (HIV), a retrovirus belonging to group 6 ( Table 1) [28][29][30][31][32][33][34][35], and [33,34]in the LTR region of lentiviruses in general (ssRNA RT) [36]. Given this amount of scattered data, we here aimed at analyzing the presence of PQSs in the genome of all known viruses that can cause infections in humans.…”

G-Quadruplexes: More Than Just a Kink in Microbial Genomes

“…The identification of G4s in microbial genomes has opened up new avenues for therapeutics; additional studies on the specificity of G4-binding ligands and their undesired effects may help us to better understand the therapeutic potential of this novel group of antimicrobial agents. Host protein-microbial G4 interaction or the host G4-microbial protein interactions at the molecular interface of the host and microbe during infection are fascinating and merit further investigation [55,56,[66][67][68][69]. It would be interesting to understand if such interactions defend the host or demonstrate yet another mechanism of microbial pathogenesis.…”

“…Nucleolin stabilizes the quadruplex and represses the transcriptional activity of the LTR promoter, allowing the virus to enter latency. Interestingly, heterogeneous nuclear ribonucleoprotein A2/B1 (hnRNPA2/B1), a host protein, binds and unfolds the quadruplex in the LTR promoter of HIV-1 provirus, leading to enhanced transcription [69]. Taken together, these results suggest that G4s in virus genomes may interact with host proteins not only to facilitate virus latency but also to revoke viruses from latency.…”