A cross-species approach using an in vivo evaluation platform in mice demonstrates that sequence variation in the human RABEP2 gene modulates ischemic stroke outcomes

Abstract

Ischemic stroke, caused by vessel blockage, results in cerebral infarction; the death of brain tissue. Previously, quantitative trait locus mapping (QTL) of cerebral infarct volume and collateral vessel number identified a single, strong genetic locus regulating both phenotypes. Additional studies identified the causative gene, encoding RAB GTPase Binding Effector Protein 2 (Rabep2). However, there is yet no evidence that variation in the human ortholog of this gene plays any role in ischemic stroke outcomes. We established an in vivo evaluation platform in mice using adeno-associated virus (AAV) gene replacement and verified that both mouse and human RABEP2 rescue the mouse Rabep2 KO ischemic stroke volume and collateral vessel phenotypes. Importantly, this cross-species complementation enabled us to experimentally investigate the functional effects of coding sequence variation in the human RABEP2 gene. We chose four coding variants from the human population that are predicted by multiple in silico algorithms to be damaging to RABEP2 function. In vitro and in vivo analyses verify that all four led to decreased collateral vessel connections and increased infarct volume. Thus, there are naturally occurring loss-of-function alleles. This cross-species approach will expand the number of targets for therapeutics development for ischemic stroke.