Basic patches on the E2 glycoprotein of eastern equine encephalitis virus influence viral vascular clearance and dissemination
Description
Abstract: Previously we found SINV-EEEV particles can be removed from the murine blood circulation in a phagocyte-dependent manner which can be disrupted by either transient depletion of vascular heparan sulfate glycosaminoglycans (GAG) or mutation of the viral E2 glycoprotein (K71/74/77A) associated with decreased GAG binding in vitro. Here, we further investigate the viral determinants of EEEV vascular clearance and evaluate their role in viremia development. We identified two large basic patches on the EEEV E2 glycoprotein which contain two known GAG-binding sites (K71/74/77 and K156/R157) and six additional basic residues (K10, R13, K56, R152, K231, and K232). We find that disruption of either basic patch by single alanine substitutions promotes prolonged retention of SINV-EEEV particles in the murine blood circulation in an experimental viremia model. Furthermore, like the attenuated K71/74/77A mutant, we observed the K156/R157A, K10A, and K231A mutations are also associated with similar enhanced viral dissemination in a mouse infection model as the attenuated K71/74/77A mutant. Surprisingly, despite known differences in GAG-binding and potential alteration in receptor interactions, we find the initial dispersal of WT and mutant SINV-EEEV virions from the inoculation site to the draining lymph node to be equivalent at 1 hour post-subcutaneous inoculation. Moreover, our data suggests the higher viremia associated with mutation of the E2 basic patches may be attributed to evasion of viremic control by blood-filtering phagocytes. Overall, this study defines viral features of the EEEV E2 glycoprotein that influence tissue-specific viral dissemination and highlights the capacity of blood-filtering phagocytes to modulate EEEV viremia. Data description: Datasets are qPCR results quantifying viral genomes in each sample. For each sample, RNA was extracted and used to generated cDNA; the cDNA was analyzed by TaqMan qPCR against a standard curve for quantification.