The long-term goal of this research is to elucidate the mechanisms of toxicant/hypoxia-induced cell injury and death. Our laboratory and others have shown that a mitochondrial calpain-like activity exists that mediates mitochondrial dysfunction under conditions of Ca2+ overload. This calpain-like activity is activated by Ca2+, induces the membrane permeability transition (MPT), decreases mitochondrial state 3 respiration, and can be inhibited by calpain inhibitors. However, studies to date have not conclusively identified a mitochondrial calpain. Previous reports have shown calpain-like activity in the intermembrane space and/or matrix fractions of isolated mitochondria. Some labs have even identified mitochondrial proteins cleaved by a calpain-like protease. This proposal will address the question of whether or not a mitochondrial calpain exists and if so, how it regulates mitochondrial dysfunction.
Specific Aim 1 will conclusively identify the calpain isoform present in renal cortical mitochondria (RCM).
Specific Aim 2 will generate RCM calpain clones and """"""""knockouts"""""""" that will be used to evaluate the effects of calpain over- and under-expression on mitochondrial function. Generation of an RCM calpain clone will allow us to isolate and purify the protein for purposes of antibody production. Further, tissue and sub-cellular localization will be determined by RT-PCR and confocal microscopy, respectively.
Specific Aim 3 will address the mechanism of RCM calpain-induced mitochondrial dysfunction. Specifically, we will focus on altered electron transport chain function and the formation of MPT after Ca2+ stimulation. Completion of these studies will increase our understanding of the events leading to hypoxic cell death, and aid in the development of therapeutic agents for the treatment of acute renal failure.
| Rasbach, Kyle A; Arrington, David D; Odejinmi, Sina et al. (2009) Identification and optimization of a novel inhibitor of mitochondrial calpain 10. J Med Chem 52:181-8 |
| Arrington, David D; Schnellmann, Rick G (2008) Targeting of the molecular chaperone oxygen-regulated protein 150 (ORP150) to mitochondria and its induction by cellular stress. Am J Physiol Cell Physiol 294:C641-50 |
| Arrington, David D; Van Vleet, Terry R; Schnellmann, Rick G (2006) Calpain 10: a mitochondrial calpain and its role in calcium-induced mitochondrial dysfunction. Am J Physiol Cell Physiol 291:C1159-71 |
