Comprehensive Analysis of G-Quadruplexes in African Swine Fever Virus Genome Reveals Potential Antiviral Targets by G-Quadruplex Stabilizers.

Elishiba Muturi,Hang Yang,Hongping Wei,Fei Meng,Mengwei Jiang,Huan Liu

doi:10.3389/fmicb.2021.798431

Abstract

African Swine Fever Virus (ASFV), a lethal hemorrhagic fever of the swine, poses a major threat to the world’s swine population and has so far resulted in devastating socio-economic consequences. The situation is further compounded by the lack of an approved vaccine or antiviral drug. Herein, we investigated a novel anti-ASFV approach by targeting G-Quadruplexes (G4s) in the viral genome. Bioinformatics analysis of putative G-quadruplex-forming sequences (PQSs) in the genome of ASFV BA71V strain revealed 317 PQSs on the forward strand and 322 PQSs on the reverse strand of the viral genome, translating to a density of 3.82 PQSs/kb covering 9.52% of the entire genome, which means that 85% of genes in the ASFV genome have at least 1 PQS on either strand. Biochemical characterization showed that 8 out of 13 conserved PQSs could form stable G4s in the presence of K+, and 4 of them could be stabilized by G4 ligands, N-Methyl Mesoporphyrin (NMM), and pyridostatin (PDS) in vitro. An enhanced green fluorescent protein (EGFP)-based reporter system revealed that the expression of two G4-containing genes, i.e., P1192R and D117L, could be significantly suppressed by NMM and PDS in 293T cells. In addition, a virus infection model showed that NMM could inhibit the replication of ASFV in Porcine Alveolar Macrophages (PAM) cells with an EC50 value of 1.16 μM. Altogether, the present study showed that functional PQSs existent in the promoters, CDS, 3′ and 5′ UTRs of the ASFV genome could be stabilized by G4 ligands, such as NMM and PDS, and could serve as potential targets for antivirals.

Highlights

African Swine Fever Virus (ASFV) is an arbovirus that causes African Swine Fever (ASF), a highly contagious and fatal hemorrhagic fever affecting domestic pig and wild boars (Arias and Sánchez-Vizcaíno, 2002)
We broadened our search to putative G-quadruplex-forming sequences (PQSs) comprising of two G-tetrads and a longer loop length of up to 12 nucleotides since it has been demonstrated that two G-tetrads are sufficient to form a G4 and that despite the low stability observed in PQSs with long loop lengths, they are capable of folding into G4s at physiological temperatures (Guédin et al, 2010; Qin et al, 2015)
We analyzed the distribution of PQSs in the ASFV genome and investigated the possibility of a G4-targeted antiviral approach in vitro

Summary

Introduction

African Swine Fever Virus (ASFV) is an arbovirus that causes African Swine Fever (ASF), a highly contagious and fatal hemorrhagic fever affecting domestic pig and wild boars (both species of Sus scrofa) (Arias and Sánchez-Vizcaíno, 2002). ASFV infection induces varying clinical forms of the disease ranging from acute to chronic infections, depending on the host’s characteristics and the virus strain (Tulman et al, 2009). ASFV has severely impacted the global swine industry, as reflected by a hike in pork prices, shut down of export markets, and shortage of swine-derived heparin (SánchezCordón et al, 2018; Vilanova et al, 2019). This situation is further aggravated by the lack of an approved vaccine or antiviral drug for the prevention or treatment of the disease. Countermeasures to contain ASF outbreaks are limited to strict animal quarantine and implementation of stamping-out policies (Gaudreault et al, 2020)

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