G-Quadruplexes Formation at the Upstream Region of Replication Origin (OriL) of the Pseudorabies Virus: Implications for Antiviral Targets

Abstract

Pseudorabies virus (PRV) is the causative agent of Aujeszky’s disease, which still causes large economic losses for the swine industry. Therefore, it is urgent to find a new strategy to prevent and control PRV infection. Previous studies have proven that guanine (G)-rich DNA or RNA sequences in some other viruses’ genomes have the potential to form G-quadruplex (G4), which serve as promising antivirus targets. In this study, we identified two novel G4-forming sequences, OriL-A and OriL-S, which are located at the upstream origin of replication (OriL) in the PRV genome and conserved across 32 PRV strains. Circular dichroism (CD) spectroscopy and a gel electrophoresis assay showed that the two G-rich sequences can fold into parallel G4 structures in vitro. Moreover, fluorescence resonance energy transfer (FRET) melting and a Taq polymerase stop assay indicated that the G4 ligand PhenDC3 has the capacity to bind and stabilize the G4. Notably, the treatment of PRV-infected cells with G4-stabilizer PhenDC3 significantly inhibited PRV DNA replication in host cells but did not affect PRV’s attachment and entry. These results not only expand our knowledge about the G4 characteristics in the PRV genome but also suggest that G4 may serve as an innovative therapeutic target against PRV.

G-quadruplexes of KSHV oriLyt play important roles in promoting lytic DNA replication

“…We used TMPyP4 as well as PhenDC-3, another G4 stabilizing agent, to confirm the impact of G4 stabilization on lytic replication. PhenDC3 has also been extensively studied as a G-quadruplex stabilizing agent in altering cellular and viral genome processes ( 71 – 73 ). We analyzed the effect of G-quadruplex stabilization through the detection of lytic genes as a reduction of lytic DNA replication impacts the transcription of lytic genes.…”

Identification of a conserved G-quadruplex within the E165R of African swine fever virus (ASFV) as a potential antiviral target

“…Previous studies have reported G4 formation in DNA viruses such as HSV, an alphaherpesvirus with a double-stranded DNA genome, which exhibits peak G4 formation during the virus replication stage ( 61 ). Previously, we reported G4 formation in pseudorabies virus, an HSV homolog, where G-rich sequences fold into G4s during replication ( 20 , 21 ). In addition to DNA G4s, it has been shown that G-rich RNA sequences are prone to forming RNA G4s as there is no complementary strand to complete base pairing.…”

“…The G4 structure consists of stacking tetrads composed of four guanine bases connected by Hoogsteen hydrogen bonds ( 18 , 19 ). Growing evidence indicates the presence of G4s in viral genomes, including pseudorabies virus ( 20 , 21 , 22 , 23 ), HIV ( 24 ), Epstein-Barr virus ( 25 ), human papillomavirus ( 26 ), hepatitis B virus ( 27 ), HSV ( 28 ), Nipah virus ( 29 ), hepatitis C virus (HCV) ( 30 ), Zika virus ( 31 , 32 ), Ebola virus ( 33 ), chikungunya virus ( 34 ), enterovirus A71 ( 35 ), Kaposi’s sarcoma-associated herpes virus ( 36 ), varicella-Zoster virus ( 37 ), severe acute respiratory syndrome (SARS)-CoV ( 38 ), and SARS-CoV-2 ( 39 , 40 ). These G4 structures play vital roles in regulating virus replication, transcription, and translation.…”