Determining the Role of Annexin A6 in Cell Membrane Repair

All cells are susceptible to stress or contraction‐induced membrane damage; any disruption to the plasma membrane, if left unrepaired, leads to cytoplasmic efflux, ion influx, and ultimately cell death. To maintain homeostasis, cells utilize a highly‐conserved, calcium‐dependent repair system that comprises multiple players that dispatch two fundamental operations: resealing of the plasma membrane and repair of the cortical cytoskeleton. Without an efficient repair system, human diseases like muscular dystrophy, diabetic myopathy, and acute lung damage result. To develop therapies to treat repair‐deficient pathologies, we are interested in identifying promoters of cell membrane repair. Due to their calcium‐dependent, lipid‐binding activity, the annexins have long attracted attention as potential participants in the repair response. Because the annexin isoform A6 has been genetically linked to muscular dystrophy, we predicted that annexin A6 plays an active role in the repair response and is a viable therapeutic target. To test whether annexin A6 promotes membrane repair, we investigated A6 function in a Xenopus laevis oocyte model of membrane repair. Specifically, we injected recombinant annexin A6 proteins into oocytes, generated a laser‐induced wound, and either measured protein efflux as a readout on repair or monitored A6 dynamic localization and behavior microscopically. Consistent with a potential role in repair for A6, GFP A6 was intensely and rapidly recruited to wound sites. Its recruitment was significantly faster than the accumulation of f‐actin around wounds and annexin A6 concentrated interior to the bulk of f‐actin around wounds. Further, the biochemical assay demonstrated addition of exogenous annexin A6 promotes the wounding response. That is, annexin A6 addition resulted in significantly less leakage of cytoplasmic protein out of the cell during the repair process. To confirm the GFP‐A6 concentration used for imaging was comparable to the annexin A6 used in the biochemical assay, we tested the relative activity of the two constructs and found that the two indeed produced equivalent results. Next steps involve examining potential binding partners of annexin A6, such as alpha actinin and s100 proteins, as well as continuing to compare localization patterns to established wound repair proteins. Since the repair mechanism is highly conserved across kingdoms, we expect results from Xenopus oocytes to readily translate to our understanding of mammalian repair response. This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal .