Author(s): Caroline Aragon, University of Minnesota; Divya Alley, University of Minnesota; Lihsia Chen, University of Minnesota //

ABSTRACT: Primary congenital hydrocephalus (CH) is a progressive, life-threatening condition resulting from an imbalance in production, flow, and absorption of cerebrospinal fluid (CSF) in the brain. If untreated, subsequent ventricular dilation and increased intracranial pressure resulting from CSF buildup can cause brain malformations, progressive macrocephaly, neurocognitive deficits, and death. While the pathophysiology underlying CH is undefined, the most common cause is loss or impaired function of L1, a transmembrane immunoglobulin cell adhesion molecule with well-defined roles in neuronal migration, synaptic regulation, and axon guidance. While these roles suggest neurodevelopmental defects as the underlying cause, the precise mechanism for how loss of L1 causes CH remains largely unknown. A major factor contributing to our lack of knowledge is the incomplete penetrance and variable expressivity seen within families harboring L1 mutations. This variability in phenotypic severity, also observed in L1 knockout mice, points to the influence of genetic background on L1-associated CH. Uncovering L1 mechanisms of action is key to developing novel therapies and preventative measures for CH.   To facilitate the identification of genes interacting with L1, we use C. elegans, which has been a premier model  system to investigate clinically significant human genetic variation. The C. elegans model provides simplicity and ease for genetic manipulation while preserving access to complex genetic interactions. Importantly, C. elegans has a single canonical L1 gene, sax-7, with conserved neurodevelopmental roles including axon branching and dendrite morphogenesis and maintenance of the neural architecture. Recently, we identified a genetic interaction between SAX-7 and the Ras/Erk signaling pathway, uncovering novel roles for SAX-7 in fluid regulation and vulval development. sax-7 null animals with hyperactive Ras/LET-60(gf) display a synthetic phenotype of progressive fluid buildup that often results in mid-to-late larval lethality. Dying animals often show ruptures at the excretory pore, suggestive of increased internal pressure. Consistent with internal fluid build-up, sax-7 let-60(gf) animals also show hypersensitivity to hypotonic solutions, their bodies tending to burst shortly after immersion as compared to sax-7 and let-60(gf) single mutants. Conditional knockout experiments using a Cre/loxP system revealed a neuronal requirement for SAX-7 in this process.  While sax-7 null animals do not exhibit vulva abnormalities, loss of SAX-7 synergistically enhances the multiple vulva phenotype (Muv) exhibited by let-60(gf) animals, revealing a previously uncharacterized role for SAX-7 in vulva development. Both the sax-7 let-60(gf) fluid buildup and Muv phenotypes can be suppressed by knocking out KSR-1, a key protein scaffold that promotes Erk signaling, consistent with elevated Erk activity as underlying the sax-7 let-60(gf) synergistic phenotypes. Interestingly, CH is sometimes presented in a cluster of diseases known as “Rasopathies,” characterized by elevated Ras/Erk signaling. Based on these findings, we hypothesize that SAX-7 and Ras/Erk signaling act synergistically in fluid homeostasis and vulval development and are currently testing this hypothesis. Improving our mechanistic understanding will shed light on the role of L1 in fluid regulation in mammals and has the potential to provide novel insight into the phenotypic variability of CH patients.

Source of Funding: NIH NINDS