Thursday, April 3, 2025
4/3/2025
Converting cytoskeletal forces into biochemical signals - PROJECT SUMMARY Cells perceive mechanical cues in their local environments, which must be converted into intracellular biochemical signals to modulate cellular physiology and control gene expression. There is increasing appreciation for mechanical signal transduction’s (“mechanotransduction”) critical role in development and its dysfunction in disease states such as cancer. However, in contrast to canonical signal transduction, cellular force sensing is poorly understood, hampering efforts to define mechanistically distinct mechanotransduction pathways, delineate their specific biological functions, and target them therapeutically. The actin cytoskeleton, a network of dynamic actin filaments, myosin motor proteins, and hundreds of associated factors, enables cells to mechanically interface with their surroundings. The cytoskeleton is classically understood to serve as a force generation and transmission apparatus that indirectly facilitates mechano- transduction through its physical linkages to membrane-anchored sites which mediate force signal conversion (e.g. cell-cell and cell-matrix adhesions). However, we and others have recently reported direct binding of soluble cytosolic proteins containing tandem arrays of LIM (LIN-11, Isl-1 & Mec-3) domains to tensed actin filaments, suggesting that the cytoskeleton itself may have the capacity to transduce forces into biochemical signals. Here I propose to test the hypothesis that force-activated actin binding by distinct LIM proteins is upstream of functionally discrete downstream mechanotransduction pathways. Through cellular assays and biophysical reconstitution, we will investigate how the representative force-activated actin binding LIM proteins zyxin (Aim 1) and FHL1/2 (Four-and-a-Half LIM domains 1/2, Aim 2) mediate distinct downstream functions in cytoplasmic cytoskeletal damage repair and nuclear gene expression regulation, respectively. We will then innovatively interface these approaches with cryo-electron microscopy (cryo-EM) to visualize force-activated actin binding by LIM proteins in structural detail (Aim 3). Our studies will establish how a conserved mechanism of force transduction through LIM domains is linked to distinct downstream signaling outcomes, which is likely to reveal general principles underlying the modular organization of cytoskeletal mechanical signaling networks. In the longer term, this work will enable precision dissection of context-specific biological functions of LIM proteins in vivo, facilitating rigorous evaluation of their potential as therapeutic targets.
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