CNBP Binds and Unfolds In Vitro G-Quadruplexes Formed in the SARS-CoV-2 Positive and Negative Genome Strands

BEZZI, G.; DIEDRICH, L.; PIGA, E.J.; BINOLFI, A.; ARMAS, P.

Marienbad (Marianské Lázne)

Congreso; G4thering 2022. 8th International Meeting on Quadruplex Nucleic Acids.; 2022

G4thering 2022. 8th International Meeting on Quadruplex Nucleic Acids.

The Coronavirus Disease 2019 (COVID-19) pandemic is caused by an RNA-genome virus called severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). G-quadruplexes (G4s) are secondary nucleic acid structures formed by nucleic acids (DNA or RNA). These secondary structures have been proposed as new transcriptional and translational regulatory elements that were originally and mainly described in oncogenes. As well, G4s have been involved in the control of a variety of biological processes, including viral replication. Using several G4 prediction tools, we found putative G4 sequences (PQS) within the RNA genome of SARS-CoV-2 (positive sense or +gRNA) and the negative sense RNA intermediate during viral replication (-gRNA) , some of which were conserved in related betacoronaviruses and others where exclusive of SARS-CoV-2 (and to some of the betacoronaviruses most related to SARS-CoV-2). Interestingly, several variations reported for SARS-CoV-2 genome may alter G4 formation, in some cases without affecting protein codification. By using multiple spectroscopic and biophysical techniques, we confirmed the formation of two G4s in the +gRNA and provide the first evidence of G4 formation by five PQSs in the −gRNA of SARS-CoV-2. Finally, spectroscopic and biochemical approaches were used to demonstrate for the first time that CNBP, the main human cellular protein bound to the SARS-CoV-2 RNA genome, binds and promotes the unfolding of some of the G4s formed by both strands of SARS-CoV-2 RNA genome. Our results suggest that G4s found in SARS-CoV-2 RNA genome and its negative-sense replicative intermediates, as well as the cellular proteins that interact with them, are relevant factors for viral genes expression and replication cycle, and may constitute interesting targets for antiviral drugs development.References:1. Bezzi, G.; Piga, E.J.; Binolfi, A.; Armas, P. CNBP Binds and Unfolds In Vitro G-Quadruplexes Formed in the SARS-CoV-2 Positive and Negative Genome Strands. Int. J. Mol. Sci. 2021, 22, 2614.