The major goal of this proposal is to understand the role of mitochondria, and their channels early in cell death pathways. Apoptosis is a basic cell process and alterations in apoptotic pathways have been implicated in loss of susceptibility of cancer cells to chemotherapeutic drugs. The recent findings that mitochondrial depolarization occurs early in apoptosis and that factors (e.g., cytochrome c) released from mitochondria induce nuclear manifestations of apoptosis in vitro indicate a pivotal role for these organelles in some apoptotic pathways. Interestingly, laser irradiation of just ten of the hundreds of mitochondria in a cell always induced an apparent exit from the cell cycle; half the cells died in 6-10 hr while the remaining cells became """"""""static"""""""", i.e., immobile and did not enter mitosis for greater than 50 hours, a time longer than two cell cycles. After approximately 50 hours, all the mitochondria of the static cells were depolarized even though only a few were irradiated. Clearly, the """"""""signal"""""""" released by just a few mitochondria into the cytosol was amplified and sufficient to cause an apparent cell cycle exit and death. These findings indicate that mitochondria can control entrance into a cell death pathway. Laser disruption of mitochondria mimics the permeability transition that mitochondria undergo when a pore in the inner membrane opens. Evidence suggests that this transition is caused by opening in the inner membrane of the multiple conductance channel, MCC, that was discovered and characterized by the PI. It is the hypothesis of this proposal that MCC opening and the resulting permeability transition initiate a cascade of events that ends in cell death. Opening of MCC is expected to cause depolarization, uncoupling, and some swelling of mitochondria leading to disruption of outer membrane integrity. The ensuing spillage of mitochondrial factors into the cytosol would resemble that which occurred during laser irradiation. Using a unique combination of electrophysiology and molecular biological tools, it will be determined if opening of the MCC can account for mitochondrial depolarization and precedes the appearance of other early markers, e.g., caspase activation, after drug treatment to induce apoptosis. The effect of agonists and antagonists of MCC on the onset and progression of apoptosis will also be determined. Overexpression of anti-apoptotic bcl-2 in mitochondria is known to block the onset of this permeability transition and delay apoptosis. Our results indicate that bcl-2 alters the Ca+2-regulation of MCC. Thus, the probability that apoptotic effectors such as bcl-2 or bax may function as modulators of MCC will be explored. The alternative possibility that bcl-2 and bax may directly affect mitochondrial permeability by forming novel ion channels will also be investigated by applying patch-clamp techniques to native mitochondrial membranes. It is expected that the results of these studies will lead to a better understanding of the role of mitochondria in cell death.
| Peixoto, Pablo M; Dejean, Laurent M; Kinnally, Kathleen W (2012) The therapeutic potential of mitochondrial channels in cancer, ischemia-reperfusion injury, and neurodegeneration. Mitochondrion 12:14-23 |
| Peixoto, Pablo M; Lue, Jennifer K; Ryu, Shin-Young et al. (2011) Mitochondrial apoptosis-induced channel (MAC) function triggers a Bax/Bak-dependent bystander effect. Am J Pathol 178:48-54 |
| Kinnally, Kathleen W; Peixoto, Pablo M; Ryu, Shin-Young et al. (2011) Is mPTP the gatekeeper for necrosis, apoptosis, or both? Biochim Biophys Acta 1813:616-22 |
| Ryu, Shin-Young; Beutner, Gisela; Kinnally, Kathleen W et al. (2011) Single channel characterization of the mitochondrial ryanodine receptor in heart mitoplasts. J Biol Chem 286:21324-9 |
| Ryu, Shin-Young; Peixoto, Pablo M; Teijido, Oscar et al. (2010) Role of mitochondrial ion channels in cell death. Biofactors 36:255-63 |
| Peixoto, Pablo M; Ryu, Shin-Young; Kinnally, Kathleen W (2010) Mitochondrial ion channels as therapeutic targets. FEBS Lett 584:2142-52 |
| Ryu, Shin-Young; Beutner, Gisela; Dirksen, Robert T et al. (2010) Mitochondrial ryanodine receptors and other mitochondrial Ca2+ permeable channels. FEBS Lett 584:1948-55 |
| Ryu, Shin-Young; Peixoto, Pablo M; Won, Jong Hak et al. (2010) Extracellular ATP and P2Y2 receptors mediate intercellular Ca(2+) waves induced by mechanical stimulation in submandibular gland cells: Role of mitochondrial regulation of store operated Ca(2+) entry. Cell Calcium 47:65-76 |
| Dejean, Laurent M; Ryu, Shin-Young; Martinez-Caballero, Sonia et al. (2010) MAC and Bcl-2 family proteins conspire in a deadly plot. Biochim Biophys Acta 1797:1231-8 |
| Peixoto, Pablo M; Ryu, Shin-Young; Pruzansky, Dawn Pietkiewicz et al. (2009) Mitochondrial apoptosis is amplified through gap junctions. Biochem Biophys Res Commun 390:38-43 |
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