Quantitative Proteomic Comparison of Rat Mitochondria from Muscle, Heart, and Liver

Abstract

Mitochondria, through oxidative phosphorylation, are the primary source of energy production in all tissues under aerobic conditions. Although critical to life, energy production is not the only function of mitochondria, and the composition of this organelle is tailored to meet the specific needs of each cell type. As an organelle, the mitochondrion has been a popular subject for proteomic analysis, but quantitative proteomic methods have yet to be applied to tease apart subtle differences among mitochondria from different tissues or muscle types. Here we used mass spectrometry-based proteomics to analyze mitochondrial proteins extracted from rat skeletal muscle, heart, and liver tissues. Based on 689 proteins identified with high confidence, mitochondria from the different tissues are qualitatively quite similar. However, striking differences emerged from the quantitative comparison of protein abundance between the tissues. Furthermore we applied similar methods to analyze mitochondrial matrix and intermembrane space proteins extracted from the same mitochondrial source, providing evidence for the submitochondrial localization of a number of proteins in skeletal muscle and liver. Several proteins not previously thought to reside in mitochondria were identified, and their presence in this organelle was confirmed by protein correlation profiling. Hierarchical clustering of microarray expression data provided further evidence that some of the novel mitochondrial candidates identified in the proteomic survey might be associated with mitochondria. These data reveal several important distinctions between mitochondrial and submitochondrial proteomes from skeletal muscle, heart, and liver Different mammalian tissues have distinct energy needs, and mitochondria morphology can vary widely, although the structure is not exclusively linked to respiration (1). Morphology and structure of mitochondria in mammalian skeletal muscle, heart, and liver are very different (2). In skeletal muscle mitochondria are distributed between sarcomeres and tightly embedded in the microfilaments (F-actin) and microtubules. Indeed the intimate association of mitochondria with the myofilaments minimizes diffusion distance and facilitates conversion of chemical energy to mechanical work. Mammalian skeletal muscle is a highly specialized tissue composed of fibers with a diverse range of properties. The aerobic type I fibers contain many mitochondria and rely on oxidative phosphorylation for ATP. Type IIa fibers are very rich in small mitochondria and are characterized by a high capacity for oxidative ATP generation. These mitochondria can only oxidize glycolytic products unlike those in type I fibers that can also utilize fatty acids and ketones. Type IIb fibers rely upon anaerobic glycolysis, contain few mitochondria, and have high levels of glycogen (3). Given these functional differences it is reasonable to assume that mitochondrial and mitochondria-related proteins are present in different amounts depending on the specific energy requirement...

Application of capillary isotachophoresis‐based multidimensional separations coupled with electrospray ionization‐tandem mass spectrometry for characterization of mouse brain mitochondrial proteome

“…Distinct proteins (553 out of 2191; or 2082 nonredundant proteins) presented in this study overlapped with the MitoP2-database, corresponding to 76% mouse mitochondrial proteome coverage. On the other hand, approximately 45% of the proteins identified by Forner et al [30] (a total of 887 proteins collected from rat tissues of muscle, heart, and liver) had an apparent orthologue against the 2004 mouse MitoP database [59]. The proteomic results of mouse brain mitochondria achieved by Kislinger et al [29] were also compared to the mitochondrial protein reference set of the MitoP2-database.…”

“…Consequently, considerable efforts have been devoted to the application of various LC techniques in single or multidimensional separation format prior to MS detection for the analysis of mitochondrial proteins [17][18][19][20][21][22][23][24][25][26][27][28][29][30][31]. In particular, the peptide-based shotgun proteomic studies fully exploit the resolution and sensitivity achievable with multidimensional LC-MS or SDS-PAGE/LC-MS approaches, allowing many additional mitochondrial proteins to be identified.…”

Can Peripheral Blood Mononuclear Cells Be Used as a Proxy for Mitochondrial Dysfunction in Vital Organs During Hemorrhagic Shock and Resuscitation?

“…In contrast to both liver and kidney, PBMC and cardiac mitochondrial function declined dramatically with reperfusion. While direct reperfusion injury with the generation of toxic reactive oxygen species and peroxynitrite may account for the acute changes seen in cardiac mitochondrial function (32,33), PBMC’s in our model were not subjected to hypoxia and the serum pO2 did not decline during hemorrhagic shock or resuscitation. Although it is unclear how resuscitation directly impairs PBMC mitochondrial function, the release of nitric oxide and IL-6 into the circulation upon reperfusion could potentially inhibit mitochondrial function directly (34,35).…”

“…Although the majority of mammalian cells contain mitochondria, there is striking diversity across tissue types in terms of mitochondrial protein expression, function, and capacity for oxidative phosphorylation (32). As such, it is not surprising that the different organs in our study demonstrated a variable response to hemorrhagic shock and resuscitation.…”

Proteomic survey towards the tissue-specific proteins of mouse mitochondria

“…In total, 399 mitochondrial proteins were identified, in which approximately 50% were not generally detected in all the tissues. This group implemented a quantitative proteomics approach to compare the mitochondrial proteins extracted from the rat tissues, heart, liver and skeletal muscle, and observed that approximate one third of the mitochondrial proteins identified were predominant to one of the three tissues [14]. The largest scale screening for the tissue-specific proteins of cytosol, microsomes, mitochondria and nuclei in the mouse brain, heart, kidney, liver, lung and placenta was performed using a gel-free multidimensional protein identification technology developed by Kislinger et al [15].…”