Physical Signaling Mechanisms That Regulate Intestinal Architecture

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PROJECT SUMMARY Physical signals are increasingly recognized as playing an important role in modulating cell behavior. The goal of this proposal is to characterize the cellular response to force-mediated signaling in the intestine and esophagus using zebrafish and mouse models. We have shown that an activating mutation in smooth muscle myosin heavy chain gene myh11 disrupts intestinal architecture in zebrafish meltdown mutants. Physical signals arising from the mutant myosin activate a conserved redox signaling pathway in the intestinal epithelium that drives the formation of plasma membrane protrusions known as invadopodia that degrade matrix proteins. The invadopodia drive the invasive transformation and cystic expansion of the epithelium. Animals that are heterozygous for the meltdown mutation develop normally but are sensitized to form the homozygous cell invasion phenotype when oncogenic signaling pathways activated. The proposal consists of three aims designed to understand how physical signals from unregulated myosin are processed by digestive epithelia and how they may be risk factors for digestive disease. The goal of the first aim are to understand how the mutant smooth muscle myosin initiates invadopodia in the epithelium of meltdown mutants and to compare this to mechanisms that regulate invadopodia formation in mammalian cells. Invadopodia have rarely if ever been observed in vivo, thus this aim offers the opportunity to understand their regulation in a live animal model. The goal of the second aim is to understand how co- activation of KRas and Wnt signaling sensitize heterozygous mutants to invasive triggers. The experiments proposed for this aim have both basic and clinical relevance, as both pathways are activated in human digestive cancers. The goal of the third aim is to characterize a recently engineered mouse model of the meltdown mutation. This includes characterization of esophageal and intestinal phenotypes in homozygous mutants, and comparison with mice newly engineered to carry knock-in mutations that are identical to human MYH11 mutations associated with heritable motility syndromes. Collectively, the proposed experiments will define novel factors and signaling mechanisms that establish and maintain digestive organ architecture and function.