ABSTRACT
The role of the mitochondrial protein receptor Tom22p in the interaction of pro-apoptotic protein Bax with yeast mitochondria was investigated. Co-immunoprecipitation assays showed that human Bax interacted with different TOM subunits, including Tom22p. Expression of the cytosolic receptor domain of human Tom22 increased Bax mitochondrial localization, but decreased the proportion of active Bax. BN-PAGE showed that the cytosolic domain of Tom22 interfered with the oligomerization of Bax. These data suggest that the interaction with the cytosolic domain of Tom22 helps Bax to acquire a conformation able to interact with the outer mitochondrial membrane.
Subject(s)
Mitochondrial Membrane Transport Proteins/chemistry , Mitochondrial Membrane Transport Proteins/physiology , Protein Interaction Domains and Motifs/physiology , Yeasts/metabolism , bcl-2-Associated X Protein/chemistry , bcl-2-Associated X Protein/metabolism , Amino Acid Sequence , Cells, Cultured , Cytosol/metabolism , Humans , Mitochondrial Membrane Transport Proteins/genetics , Mitochondrial Membrane Transport Proteins/metabolism , Mitochondrial Membranes/metabolism , Mitochondrial Precursor Protein Import Complex Proteins , Molecular Sequence Data , Organisms, Genetically Modified , Protein Binding/genetics , Protein Conformation , Protein Interaction Domains and Motifs/genetics , Protein Transport , Sequence Homology, Amino Acid , Yeasts/geneticsABSTRACT
Bax insertion into the mitochondrial outer membrane is essential for the implementation of apoptosis. However, little is known about the first stage of Bax integration into the mitochondrial outer membrane. We have recently shown that TOM22, a mitochondrial outer membrane receptor, is important for insertion, although other reports have suggested that only mitochondrial lipids are involved in this process. Here, we show that monomers, but not dimers, of Bax require the presence of TOM22 and TOM40 to integrate into mitochondria. In addition we show that once inserted into the membrane, Bax can act as a receptor for cytosolic Bax.
Subject(s)
Apoptosis , Mitochondrial Membranes/metabolism , bcl-2-Associated X Protein/metabolism , Animals , Dimerization , Membrane Transport Proteins/metabolism , Mitochondria, Liver/metabolism , Mitochondrial Membrane Transport Proteins , Mitochondrial Precursor Protein Import Complex Proteins , Mitochondrial Proteins/antagonists & inhibitors , Mitochondrial Proteins/metabolism , RatsABSTRACT
Keeping a cytosolic redox balance is a prerequisite for living cells in order to maintain a metabolic activity and enable growth. During growth of Saccharomyces cerevisiae, an excess of NADH is generated in the cytosol. Aerobically, it has been shown that the external NADH dehydrogenase, Nde1p and Nde2p, as well as the glycerol-3-phosphate dehydrogenase shuttle, comprising the cytoplasmic glycerol-3-phosphate dehydrogenase, Gpdlp, and the mitochondrial glycerol-3-phosphate dehydrogenase, Gut2p, are the most important mechanisms for mitochondrial oxidation of cytosolic NADH. In this review we summarize the recent results showing (i) the contribution of each of the mechanisms involved in mitochondrial oxidation of the cytosolic NADH, under different physiological situations; (ii) the kinetic and structural properties of these metabolic pathways in order to channel NADH from cytosolic dehydrogenases to the inner mitochondrial membrane and (iii) the organization in supramolecular complexes and, the peculiar ensuing kinetic regulation of some of the enzymes (i.e. Gut2p inhibition by external NADH dehydrogenase activity) leading to a highly integrated functioning of enzymes having a similar physiological function. The cell physiological consequences of such an organized and regulated network are discussed.