Mapping and characterization of G‐quadruplexes in monkeypox genomes

Monkeypox virus (MPXV) is a double‐stranded DNA virus from the family Poxviridae, which is endemic in West and Central Africa. Various human outbreaks occurred in the 1980s, resulting from a cessation of smallpox vaccination. Recently, MPXV cases have reemerged in non‐endemic nations, and the 2022 outbreak has been declared a public health emergency. Treatment optionsare limited, and many countries lack the infrastructure to provide symptomatic treatments. The development of cost‐effective antivirals could ease severe health outcomes. G‐quadruplexes have been a target of interest in treating viral infections with different chemicals. In the present work, a genomic‐scale mapping of different MPXV isolates highlighted two conserved putative quadruplex‐forming sequences MPXV‐exclusive in 590 isolates. Subsequently, we assessed the G‐quadruplex formation using circular dichroism spectroscopy and solution small‐angle X‐ray scattering. Furthermore, biochemical assays indicated the ability of MPXV quadruplexes to be recognized by two specific G4‐binding partners—Thioflavin T and DHX36. Additionally, our work also suggests that a quadruplex binding small‐molecule with previously reported antiviral activity, TMPyP4, interacts with MPXV G‐quadruplexes with nanomolar affinity in the presence and absence of DHX36. Finally, cell biology experiments suggests that TMPyP4 treatment substantially reduced gene expression of MPXV proteins. In summary, our work provides insights into the G‐quadruplexes from the MPXV genome that can be further exploited to develop therapeutics.