LDLRAD3 Receptor Interaction with Venezuelan Equine Encephalitis Virus

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Project Summary Alphaviruses are mosquito-transmitted, positive-strand enveloped RNA viruses of the Togaviridae family that cause global disease in humans. At present, no antiviral agents or licensed vaccines exist for the treatment or prevention of any alphavirus infections. We recently used a genome-wide CRISPR/Cas9-based screen to identify the cell surface molecule LDLRAD3 as a novel, highly conserved entry receptor for Venezuelan equine encephalitis virus (VEEV), an emerging pathogen capable of causing fatal neuroinvasive disease in humans and other vertebrate animals. Gene editing of mouse or human LDLRAD3 resulted in reduced VEEV infection of neuronal cells, and reciprocally, ectopic expression of LDLRAD3 resulted in increased infection. LDLRAD3 bound directly to VEEV virions and enhanced virus attachment and internalization into cells. Genetic studies indicated that domain 1 (D1) of LDLRAD3 is necessary and sufficient to support VEEV infection. We hypothesize that engagement of LDLRAD3 by VEEV will explain how infection, tissue targeting, and disease pathogenesis occurs. The primary goals of this collaborative, interactive project between the Diamond, Fremont, and Whelan laboratories are to define the precise mechanism(s) by which LDLRAD3 facilitates alphavirus entry into cells, to gain high-resolution structural insight as to how LDLRAD3 engages the spike proteins on the virion, and to determine the cell-type specific role of LDLRAD3 in VEEV pathogenesis in vivo. The experiments in this proposal will define fundamental aspects of VEEV biology that enhance our understanding of infection and cell tropism. This information may facilitate the development of small molecules or biologicals that disrupt LDLRAD3 interaction with VEEV spike proteins, which could form the basis of future therapeutics that ameliorate disease of this emerging and highly pathogenic alphavirus.