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  • Investigating the role of the actin regulating complex ARP2/3 in rapid ischemic tolerance induced neuro-protection.

Investigating the role of the actin regulating complex ARP2/3 in rapid ischemic tolerance induced neuro-protection.

International journal of physiology, pathophysiology and pharmacology (2014-01-01)
Veronica J Jessick, Mian Xie, Andrea N Pearson, Dan J Torrey, Michelle D Ashley, Simon Thompson, Robert Meller
ABSTRACT

Neuronal morphology is highly sensitive to ischemia, although some re-organization may promote neuroprotection. In this study we investigate the role of actin regulating proteins (ARP2, ARP3 and WAVE-1) and their role in rapid ischemic tolerance. Using an established in vitro model of rapid ischemic tolerance, we show that WAVE-1 protein levels are stabilized following brief tolerance inducing ischemia (preconditioning). The stabilization appears to be due to a reduction in the ubiquitination of WAVE-1. Levels of ARP2, ARP3 and N-WASP were not affected by ischemic preconditioning. Immunocytochemical studies show a relocalization of ARP2 and ARP3 proteins in neurons following preconditioning ischemia, as well as a re-organization of actin. Blocking the protein kinase CK2 using emodin blocks ischemic tolerance, and our data suggests CK2 binds to WAVE-1 in neurons. We observe an increase in binding of the ARP2 subunit with WAVE-1. The neuroprotection observed following preconditioning is inhibited when cells are transduced with an N-WASP CA domain that blocks the activation of ARP2/3. Together these data show that ischemia affects actin regulating enzymes, and that the ARP2/3 pathway plays a role in rapid ischemic tolerance induced neuroprotection.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
Anti-N-WASP Antibody, Chemicon®, from rabbit
Millipore
Protein G Immunoprecipitation Kit, sufficient for 50 assays
Sigma-Aldrich
Anti-PIR121-1/Sra-1 Antibody, Upstate®, from rabbit