Martins, I. C. | Search bioRxiv

bioRxiv2025

Dengue virus capsid protein interaction with nucleic acids

Dengue virus (DENV) transmission has greatly increased in the last decade, in part due to the geographical expansion of its vector, the Aedes spp. mosquitoes. These arthropods are now found even in temperate climates, including Europe, where outbreaks have already occurred. A better understanding of the virus life cycle is essential, as it may enable the development of specific treatments or therapies which are currently lacking. However, recent breakthroughs concerning the biologically relevant viral capsid (C) protein structure and function are encouraging. It is now clear the C protein binds both to host lipid droplets and to the viral genome, a set of interactions that is crucial for viral encapsidation and replication. Previously, we studied DENV C protein interaction with lipid droplets, at the molecular level. Here, we examine how the association of DENV C protein with the viral genome occurs. Using DENV C protein and single-stranded DNA (ssDNA) sequences analogous to previously identified important regions of the viral genome, we biophysically characterize their interactions. A decrease in fluorescence intensity and lifetime, an increase in both anisotropy and hydrodynamic diameter, as well as changes in the proteins secondary structure, were observed upon binding to ssDNA. These changes are consistent with molecular condensation and raise the possibility of liquid-liquid phase separation involvement in DENV C-nucleic acid complex formation. These findings may be useful for the development of future therapeutic strategies based on nucleic acids, not only for DENV but also for other related viruses.

biophysics↗

bioRxiv2019

Genome-wide Associations of Flavivirus Capsid Proteins

Dengue virus (DENV) and Zika virus (ZIKV) are both positive sense single-stranded RNA viruses. They are packaged within the virion with a capsid (C) protein to form the nucleocapsid. Based on cryo-electron microscopy imaging, the nucleocapsid has been described as lacking symmetry, whilst there is distinguishable separation of the C proteins from the viral RNA (vRNA) genome. Here, to elucidate the architecture of the nucleocapsid of DENV serotype 2 and ZIKV, we used a nuclease digestion assay and next-generation sequencing to map the respective vRNA genome wide association with the C protein in vitro. The C protein exhibited non-uniform binding along the vRNA, and as C protein concentration increased, the normalized read counts also increased. A saturation point of 1:100 (vRNA:C protein monomers) was found, and binding regions showed variable saturation patterns. We also observed that C protein had a preference for G-rich sequences for both viruses. Taken together, we demonstrate that the DENV 2 and ZIKV C proteins bind vRNA in a non-uniform manner with distinct patterns of association.\n\nSingificance StatementOur study demonstrates that flavivirus capsid proteins associate with the viral genome at specific sites rather than in a uniform manner as commonly expected. We estimate the number of capsid proteins binding to a single genomic RNA. We proceed to locate the capsid binding sites along the viral genomes of Dengue and Zika viruses. We characterize the binding sites in terms of affinity and analyze the nucleotide composition and sequence motifs at binding sites. We cross-reference binding sites against SHAPE reactivity data corresponding to local RNA secondary structure, which allows us to identify structural motifs of capsid binding sites. As capsid proteins are essential for viral packaging, these interactions may form attractive targets for therapeutic intervention.

genomics↗