Congenital hydrocephalus (CH), the primary enlargement of the cerebrospinal (CSF)-filled brain ventricles, affects 1/1000 births and is a major cause of morbidity and mortality. Although ~60% of all CH cases are predicted to have a genetic etiology, known genes account for <5% of CH cases. Significant gaps in our understanding of the molecular pathogenesis of CH impede the development of preventive, diagnostic, and therapeutic measures. Fundamental obstacles to CH gene discovery using traditional genetic approaches include locus heterogeneity, phenotypic complexity, and the sporadic nature of the majority of CH cases. Whole exome sequencing (WES) has the potential to overcome these obstacles and has led to unprecedented opportunities for gene discovery in autism and structural brain disorders. We recently used WES to identify four novel CH genes, accounting for ~10% of studied CH cases (Furey et al., Neuron, 2018). All four genes are required for neural tube development and regulate neural stem cell (NSC) fate. These results implicate impaired neurogenesis, rather than CSF over-accumulation, in the pathogenesis of a significant subset of CH patients, with potentially paradigm-changing diagnostic and therapeutic implications. As many causal CH genes remain undiscovered, our objective here is to utilize a functional genomics approach to discover, validate, and gain mechanistic insight into newly identified CH-causing mutations. Our hypothesis is that WES will identify multiple novel CH genes, many of which will converge on pathways that regulate the NSC development. Based on our experience that has been successful in identifying several CH and structural brain disorder genes over the past several years, we now propose to ascertain additional sporadic CH case-parent trios and Turkish consanguineous familial CH forms and perform WES on our large, well-phenotyped CH cohort to discover novel de novo and transmitted CH gene mutations. This will be followed by analyses to determine the expression patterns of newly identified prioritized genes during mammalian brain development. We will then rapidly and inexpensively functionally screen prioritized CH candidate gene mutations for their ability to recapitulate hydrocephalus using our novel, validated platform that utilizes live Xenopus embryos, CRISPR/Cas9 gene editing, quantitative optical coherence tomography, and real-time CSF particle tracking (Date et al., Sci Rep, 2019, Accepted). For select validated genes, we will establish Xenopus lines to elucidate the biological consequences of human CH mutations on cilia-regulated CSF dynamics and NSC growth/differentiation and patterning. This functional genomics approach will elucidate the genetic architecture of CH, and set the stage for more detailed future biological studies in mouse models beyond the scope of this proposal. Ultimately, such knowledge has the potential to improve clinical management, prognostication, surveillance, and genetic advice; stimulate research into new non-surgical therapies; and improve the quality of our support for CH patients and their families.
Hydrocephalus affects 1/1,000 live births and is a major cause of morbidity and mortality. We propose to use a functional genomics approach that combines next-generation DNA sequencing and a high-throughout functional screening platform to discover, validate, and gain mechanistic insight into novel gene mutations associated with human congenital hydrocephalus. Results could lead to fundamental new insights into human brain development and serve as the basis for improved clinical decision-making, prognostication, and novel biology-based treatments for congenital hydrocephalus patients.