The La Crosse virus class II fusion glycoprotein loop contributes to infectivity and cholesterol-dependent entry.

Unlabelled: Arthropod-borne viruses (arboviruses) are an emerging and evolving global public health threat with little to no antiviral treatments. La Crosse virus (LACV) from the order is responsible for pediatric encephalitis cases in the United States, yet little is known about the infectivity of LACV. Given the structural similarities between class II fusion glycoproteins of LACV and chikungunya virus (CHIKV), an alphavirus from the family, we hypothesized that LACV would share similar entry mechanisms to CHIKV. To test this hypothesis, we performed cholesterol-depletion and repletion assays and used cholesterol modulating compounds to study LACV entry and replication. We found that LACV entry was cholesterol-dependent while replication was less affected by cholesterol manipulation. In addition, we generated single point mutants in the LACV loop that corresponded to known CHIKV residues important for virus entry. We found that a conserved histidine and alanine residue in the Gc loop impaired virus infectivity and attenuate LACV and . Finally, we took an evolution-based approach to explore how the LACV glycoprotein evolution in mosquitoes and mice. We found multiple variants that cluster in the Gc glycoprotein head domain, supporting the Gc glycoprotein as a target for LACV adaptation. Together, these results begin to characterize the mechanisms of LACV infectivity and how the LACV glycoprotein contributes to infectivity and pathogenesis.

Importance: Vector-borne arboviruses are significant health threats leading to devastating disease worldwide. This emergence and the fact that there are little to no vaccines or antivirals targeting these viruses highlights the need to study how arboviruses replicate at the molecular level. One potential antiviral target is the class II fusion glycoprotein. Alphaviruses, flaviviruses, and bunyaviruses encode a class II fusion glycoprotein that contain strong structural similarities in the tip of domain II. Here we show that the bunyavirus La Crosse virus uses similar mechanisms to entry as the alphavirus chikungunya virus and residues in the loop are important for virus infectivity. These studies show that genetically diverse viruses use similar mechanisms through concerned structure domains, suggesting these may be a target for broad-spectrum antivirals to multiple arbovirus families.