G-quadruplex in the TMV Genome Regulates Viral Proliferation and Acts as Antiviral Target of Photodynamic Therapy

Plant viruses seriously disrupt crop growth and development, and classic protein-targeted antiviral drugs could not provide complete protection against them. It is urgent to develop antiviral compounds with novel targets. Photodynamic therapy shows potential in controlling agricultural pests, but nonselective damage from reactive oxygen species (ROS) unexpectedly affects healthy tissues. A G-quadruplex (G4)-forming sequence in the tobacco mosaic virus (TMV) genome was identified to interfere the RNA replication in vitro, and affect the proliferation of TMV in tobacco. N-methyl mesoporphyrin IX stabilizing the G4 structure exhibited inhibition against viral proliferation, which was comparable to the inhibition effect of ribavirin. This indicated that G4 could work as an antiviral target. The large conjugate planes shared by G4 ligands and photosensitizers (PSs) remind us that the PSs could work as antiviral agents by targeting G4 in the genome of TMV. Chlorin e6 (Ce6) was identified to stabilize the G4 structure in the dark and selectively cleave the G4 sequence by producing ROS upon LED-light irradiation, leading to 92.2% inhibition against TMV in vivo, which is higher than that of commercial ningnanmycin. The inhibition of Ce6 was lost against the mutant variants lacking the G4-forming sequence. These findings indicated that the G-quadruplex in the TMV genome worked as an important structural element regulating viral proliferation, and could act as the antiviral target of photodynamic therapy. Plant viruses cause significant losses in crops worldwide, and there is an urgent need to develop antiviral compounds with novel targets. Drugs targeting RNA have attracted much attention, but RNA structures as the antiviral targets remain to be explored. In this study, a photosensitizer, chlorin e6, was identified to cleave a G4 forming sequence in the TMV genome by producing ROS upon LED-light irradiation and to exhibit inhibition against TMV higher than that of commercial Ningnanmycin. This is the first time to show that the folding of G4 regulates the proliferation of plant viruses, which illustrates the viral replication mechanism from the RNA structural perspective. Compounds stabilizing G4 structures or breaking G4 forming sequences are expected to be the novel antivirals against the plant viruses.