Biophysical Society Thematic Meeting | Singapore

Mechanobiology of Disease

Wednesday Speaker Abstracts

Calpains Influence both Cytoskeletal Remodeling and Ca 2+ -Triggered Vesicle Fusion in the Emergency Response to Repair a Membrane Injury Ann-Katrin Piper 1,2 , Angela Lek 3,4 , Gregory Redpath 1,2 , Frances Lemckert 1,2 , Natalie Woolger 1,2 , Sandra T. Cooper 1,2 . 1 The University of Sydney, Sydney, NSW, Australia, 2 Institute for Neuroscience and Muscle Research, Sydney, NSW, Australia, 3 Division of Genetics and Genomics, Boston Children's Hospital, Boston, MA, USA, 4 Australian Regenerative Medicine Institute, Monash University, Clayton, VIC, Australia. Repairing membrane lesions is an evolutionary conserved process vital for eukaryotic cells to survive injury from osmotic stress, bacterial infection and parasites as well as mechanical and ischemic insults. Wounded cells survive by mounting an emergency repair response utilizing vesicle fusion to ‘patch’ membrane tears. The muscular dystrophy protein dysferlin is a Ca 2+ - regulated vesicle fusion protein that plays a key role in membrane repair. Our research reveals that cells ‘sense’ and repair membrane injuries through regulated interplay between calpains and dysferlin. We show that in the unique setting of membrane injury, rapid Ca 2+ -influx activates calpains that specifically cleave dysferlin, releasing a C-terminal effector fragment termed mini-dysferlin C72 . 3D structured illumination microscopy (3D-SIM) of primary human myotubes subject to ballistics injury resolved the rapid recruitment of mini-dysferlin C72 -containing cytoplasmic vesicles to sites of membrane injury. These dysferlin vesicles undergo Ca 2+ -dependent integration into the plasma membrane; intensely labeling the periphery of the lesion, then form a repair lattice that is eventually ‘zippered’ together by cytoskeletal motors to repair the injury. We propose calpains are central regulators of the membrane repair response, acting both to functionally modify the vesicle fusion protein dysferlin and sever plasma membrane tethers facilitating rapid remodeling of cortical actin and microtubule networks for the rapid transport of vesicles and subsequent acto-myosin contraction of the wound site. We are using murine and cell biology models of dysferlin- and calpain-deficiency to elucidate the respective roles and hierarchy of dysferlin and calpain for the emergency cell survival mechanism of membrane repair. Our molecular understanding of membrane repair will directly inform best practice for emerging calpain-modulatory therapies for recovery from cardiac and brain ischemia-reperfusion injury, and evaluate their application to muscular dystrophy and bacterial infection.

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