Hossain, Mohammed Iqbal, Roulston, Carli L, Kamaruddin, Mohd Aizuddin, Chu, Percy WY, Ng, Dominic CH ORCID: 0000-0003-4657-159X, Dusting, Gregory James, Bjorge, Jeffrey D, Williamson, Nicholas A ORCID: 0000-0002-2173-3452, Fujita, Donald J, Cheung, Steve N, Chan, Tung On ORCID: 0000-0001-6574-9555, Hill, Andrew F ORCID: 0000-0001-5581-2354 and Cheng, Heung-Chin ORCID: 0000-0002-4965-7148 (2013) A truncated fragment of Src protein kinase generated by calpain-mediated cleavage is a mediator of neuronal death in excitotoxicity. Journal of Biological Chemistry, 288 (14). pp. 9696-9709. ISSN 0021-9258

Abstract

Excitotoxicity resulting from overstimulation of glutamate receptors is a major cause of neuronal death in cerebral ischemic stroke. The overstimulated ionotropic glutamate receptors exert their neurotoxic effects in part by overactivation of calpains, which induce neuronal death by catalyzing limited proteolysis of specific cellular proteins. Here, we report that in cultured cortical neurons and in vivo in a rat model of focal ischemic stroke, the tyrosine kinase Src is cleaved by calpains at a site in the N-terminal unique domain. This generates a truncated Src fragment of ∼52 kDa, which we localized predominantly to the cytosol. A cell membrane-permeable fusion peptide derived from the unique domain of Src prevents calpain from cleaving Src in neurons and protects against excitotoxic neuronal death. To explore the role of the truncated Src fragment in neuronal death, we expressed a recombinant truncated Src fragment in cultured neurons and examined how it affects neuronal survival. Expression of this fragment, which lacks the myristoylation motif and unique domain, was sufficient to induce neuronal death. Furthermore, inactivation of the prosurvival kinase Akt is a key step in its neurotoxic signaling pathway. Because Src maintains neuronal survival, our results implicate calpain cleavage as a molecular switch converting Src from a promoter of cell survival to a mediator of neuronal death in excitotoxicity. Besides unveiling a new pathological action of Src, our discovery of the neurotoxic action of the truncated Src fragment suggests new therapeutic strategies with the potential to minimize brain damage in ischemic stroke.

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