Cellular Mechanisms of Periventricular Heterotopia

About This Grant

ABSTRACT Cellular polarity is a fundamental characteristic of all epithelial cells. Conserved polarity complexes include the apical domain of Crumbs/Pals1/Patj and Pard3/Par6/aPKC, and the basolateral domain of Scribble/Dlg/Lgl. The functional antagonism between these complexes is critical. Specifically, within the neuroepithelium, polarity complexes help orchestrate the appropriate lineage progression and subsequent neuron production of the cerebral cortex. Disruption to normal polarity processes or mutations in polarity genes contribute to a variety of human conditions. Mutations in Crumbs, Pals1, and Scribble, have been shown to cause ventriculomegaly, microcephaly, and seizures, respectively. Though many of the polarity complexes have been extensively studied in the context of neurodevelopment, the function of the basal polarity protein LGL1 (lethal giant larvae 1) is less understood. Loss of Lgl1 causes periventricular heterotopia (PH), or the abnormal accumulation of neurons along the ventricular surface. These current notions of LGL1 are derived from murine and drosophila studies, with no model of human cortical development to date. To address this, my work focuses on determine the function of Llgl1 in a human model of neurodevelopment using dorsal forebrain organoids (hCOs). We preliminarily have found that Llgl1 KO hCOs have abnormal development, including increased rosette size and reduction in rosette number. Ectopically located cells are also abundant within rosettes, suggestive of PH. Previous works have established a dependence on YAP signaling in the development of PH due to various genetic causes. Despite this association, no mechanistic studies exist describing how YAP is causing PH. We aim to address this gap in knowledge by elucidating the mechanism of YAP repression by Llgl1 in a human model. We hypothesize that LLGL1 directly represses YAP activation through modulation of kinase activity, and that the loss of this repression is what drives PH. To address this hypothesis we will start by determining the function of LLGL1 in hCOs through a KO hESC line. With this model we will perform immunostaining on markers of cerebral cortex development to characterize disruptions to neurodevelopment. I will also investigate the cause of increased rosette size, which we currently believe is due to an increase in proliferative division of neural progenitor cells. With single cell RNA sequencing I will also identify molecular and cellular changes through gene expression analysis. These studies will be followed up by a series of mechanistic studies that will identify the protein interactome of LLGL1 in neural progenitor cells, as well as elucidate the mechanism of YAP regulation by LLGL1. This work will provide novel insight into how apicobasal polarity regulates cortical development in a human model, as well as determine how YAP activation contributes to the development of PH.