In vivo analysis of mechanotransduction

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In vivo analysis of mechanotransduction Cells in biological tubes must integrate biochemical and mechanical cues in order to expand or contract in a coordinated manner. Inappropriate responses to changing states underlie conditions such as heart disease, hypertension and asthma. Despite insights from biophysics and from cell biology on engineered substrates, many important questions remain regarding how mechanical information is sensed by cells, translated into biochemical signals, and integrated to produce a coordinated tissue-level response. For example, how is multicellular contractility regulated in space and time? How are the induction and propagation of biochemical signals regulated by mechanical cues? How do different cell types within a tissue coordinate their actions? To address these questions, we have developed an in vivo model, the C. elegans spermatheca, which is a tubular tissue in the nematode reproductive system comprised of 24 smooth-muscle-like cells that connect to the uterus via a toroidal valve. The major advantages of this system are that the cells are naturally stretched and contract as oocytes enter, and are amenable to quantitative live imaging and targeted genetic manipulation, enabling observation and manipulation of individual cells in the context of an intact tissue. We have discovered that oocyte entry induces Ca2+ pulses that sweep across the tissue, culminating in a coordinated contraction that pushes the fertilized embryo into the uterus. Ca2+ release and contractility in the spermatheca and valve are coordinated such that while the spermathecal bag contracts, the valve dilates to allow exit of the fertilized embryo. Well-conserved gene networks regulate these processes, suggesting broad applicability of our findings to other contractile systems. Here, we propose a combination of 4D imaging of genetically-encoded biosensors, proteomics, molecular genetics, and modeling to elucidate the mechanisms which coordinate Ca2+ signaling in response to stretch. Specifically, we will 1) test the hypothesis that the heterotrimeric G protein, Gαs, signals through PKA to regulate spermathecal contractility; 2) model the mechanisms by which stretch triggers calcium release and signal propagation; and 3) determine how valve contractility is regulated, both autonomously and via communication from the spermathecal bag. This research will lead to important advances in our understanding of the fundamental mechanisms by which cells convert mechanical information into biochemical signals, and how this signaling is integrated to regulate tissue function.