Binding mitochondria to cryogel monoliths allows detection of proteins specifically released following permeability transition

Teilum, Maria; Hansson, Magnus; Dainiak, Maria B.; Månsson, Roland; Surve, Sheryl; Elmér, Eskil; Önnerfjord, Patrik; Mattiasson, Gustav

doi:10.1016/j.ab.2005.08.032

Cited by 20 publications

(9 citation statements)

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“…To identify the proteins specifically released from PT-induced mitochondria, differential analysis between samples prepared from PT-induced mitochondria and from nontreated mitochondria, as performed in this study, is essential. Recently to eliminate the background proteins, Teilum et al (23) prepared the released protein samples by using mitochondria immobilized on a sponge-like material, cryogel monoliths. As a result of the proteome analysis of thus prepared samples, they showed 68 proteins released from Ca 2ϩ -treated mitochondria.…”

Section: Discussionmentioning

confidence: 99%

Differential Permeabilization Effects of Ca2+ and Valinomycin on the Inner and Outer Mitochondrial Membranes as Revealed by Proteomics Analysis of Proteins Released from Mitochondria

Yamada

Yamamoto

Yamazaki

et al. 2009

Molecular & Cellular Proteomics

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It is well established that cytochrome c is released from mitochondria when the permeability transition (PT) of this organelle is induced by Ca 2؉ . Our previous study showed that valinomycin also caused the release of cytochrome c from mitochondria but without inducing this PT (Shinohara, Y., Almofti, M. R., Yamamoto, T., Ishida, T., Kita, F., Kanzaki, H., Ohnishi, M., Yamashita, K., Shimizu, S., and Terada, H. (2002) Permeability transition-independent release of mitochondrial cytochrome c induced by valinomycin. Eur. J. Biochem. 269, 5224 -5230). These results indicate that cytochrome c may be released from mitochondria with or without the induction of PT. In the present study, we examined the protein species released from valinomycin-and Ca 2؉ -treated mitochondria by LC-MS/MS analysis. As a result, the proteins located in the intermembrane space were found to be specifically released from valinomycin-treated mitochondria, whereas those in the intermembrane space and in the matrix were released from Ca 2؉ -treated mitochondria. These results were confirmed by Western analysis. Furthermore to examine how the protein release occurred, we examined the correlation between the species of released proteins and those of the abundant proteins in mitochondria. Consequently most of the proteins released from mitochondria treated with either agent were highly expressed proteins in mitochondria, indicating that the release occurred not selectively but in a manner dependent on the concentration of the proteins. Based on these results, the permeabilization effects of Ca Mitochondria are well known as the organelle for energy conversion in all eukaryotes. This energy conversion, i.e. ATP synthesis, is performed by using the electrochemical gradient of H ϩ across the inner mitochondrial membrane. To enable effective energy conversion, the mitochondrial inner membrane is highly resistant to the permeation of solutes and ions. However, under certain conditions, such as in the presence of Ca 2ϩ and inorganic phosphate, the permeability of this inner membrane is known to be markedly increased. This phenomenon is referred to as the permeability transition (PT) 1 and is believed to result from the formation of a proteinaceous pore, referred to as the PT pore, which makes the inner membrane permeable to various solutes and ions smaller than 1.5 kDa (1-3). The physiological importance of the PT has long been uncertain; however, recent studies have revealed that the changes in the permeability of the inner mitochondrial membrane due to the induction of PT cause the release of cytochrome c into the cytosol and that the released cytochrome c then triggers subsequent steps of programmed cell death, which is known as apoptosis (4 -6). Thus, the PT is considered to be one of the major regulatory steps of apoptosis. However, the questions as to how the PT is induced and how cytochrome c is released accompanied by the induction of PT have remained unanswered.To characterize the features of the mitochondrial PT and to understand the mechani...

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Section: Discussionmentioning

confidence: 99%

Differential Permeabilization Effects of Ca2+ and Valinomycin on the Inner and Outer Mitochondrial Membranes as Revealed by Proteomics Analysis of Proteins Released from Mitochondria

Yamada

Yamamoto

Yamazaki

et al. 2009

Molecular & Cellular Proteomics

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“…The macroporous MG monoliths (porosity 1 -100 lm) were successfully used as chromatography media for separation of viruses [68], cell organelles [35], microbial [28,59,61] and mammalian [19,40] cells or as cell culture 3-D-scaffolds [16,41,69] with controlled degradability [72].…”

Section: Applications Of Mgsmentioning

confidence: 99%

“…Their main applications are found in biocatalysis with immobilized cells [12,27,54,64,65] and enzymes [38,57,58], in bioseparations for purification of targets from particulatecontaining fluids [18, 30 -33, 37, 66, 67], chromatography of cell organelles [35], viruses [68], microbial [28,61,62], and mammalian [16,19,40] cells and in biomedical applications as 3-D matrixes for culture of mammalian cells [41,69].…”

Section: Applications Of Mgsmentioning

confidence: 99%

Macroporous gels prepared at subzero temperatures as novel materials for chromatography of particulate‐containing fluids and cell culture applications

Plieva

Galaev

Mattìasson³

2007

J of Separation Science

180

134

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Macroporous gels (MGs) with a broad variety of morphologies are prepared using the cryotropic gelation technique, i. e. gelation at subzero temperatures. These highly elastic hydrophilic materials can be produced from practically any gel-forming system with a broad range of porosity extending from elastic and porous gels with pore sizes up to 1.0 microm to elastic and sponge-like gels with pore sizes up to 100 microm. The versatility of the cryogelation technique is demonstrated by use of different chemical reactions (hydrogen bond formation, chemical cross-linking of polymers, free radical polymerization) mainly in an aqueous medium. Appropriate control over solvent crystallization (formation of solvent crystals) and rate of chemical reaction during the cryogelation allows the reproducible preparation of cryogels with tailored properties. Different approaches, such as chemical modification of reactive groups, grafting of the pore surface with an appropriate polymer, or direct copolymerization with functional monomers are used for control of the surface chemistry of MGs. Typically, MGs with pore sizes up to 1.0 microm are produced in the shape of beads and MGs with pore size up to 100 microm are prepared as monoliths, discs, and sheets. The difference in porous structure of MGs defines the main applications of these porous materials. Elastic beaded MGs are mostly used as carriers for cell and enzyme immobilization or for capture of low-molecular weight targets from particulate-containing fluids in expanded-bed mode. However, the elastic and sponge-like MG monoliths with interconnected pores measuring hundreds of mum have been successfully used as monolithic columns for chromatography of particulate-containing fluids (crude cell homogenates, viruses, whole cells, wastewater effluents) and as three-dimensional scaffolds for mammalian cell culture applications.

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“…Several investigations have made key contributions to our understanding of mitochondrial subproteomes (11)(12)(13)(14)(15)(16)(17), linking subproteomic changes to ischemic injury. However, how ischemic injury modifies the mitochondrial proteome (including the identities of affected proteins, their sub-organelle location and biological functions), has not been defined.…”

Section: Introductionmentioning

confidence: 99%

Altered Proteome Biology of Cardiac Mitochondria Under Stress Conditions

et al. 2008

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Myocardial ischemia-reperfusion induces mitochondrial dysfunction and, depending upon the degree of injury, may lead to cardiac cell death. However, our ability to understand mitochondrial dysfunction has been hindered by an absence of molecular markers defining the various degrees of injury. To address this paucity of knowledge, we sought to characterize the impact of ischemic damage on mitochondrial proteome biology. We hypothesized that ischemic injury induces differential alterations in various mitochondrial sub-compartments, that these proteomic changes are specific to the severity of injury, and that they are important to subsequent cellular adaptations to myocardial ischemic injury. Accordingly, an in vitro model of cardiac mitochondria injury in mice was established to examine two stress conditions: reversible injury (induced by mild calcium overload) and irreversible injury (induced by hypotonic stimuli). Both forms of injury had a drastic impact on the proteome biology of cardiac mitochondria. Altered mitochondrial function was concomitant with significant protein loss/shedding from the injured organelles. In the setting of mild calcium overload, mitochondria retained functionality despite the release of numerous proteins, and the majority of mitochondria remained intact. In contrast, hypotonic stimuli caused severe damage to mitochondrial structure and function, induced increased oxidative modification of mitochondrial proteins, and brought about detrimental changes to the sub proteomes of the inner mitochondrial membrane and matrix. Using an established in vivo murine model of regional myocardial ischemic injury, we validated key observations made by the in vitro model. This preclinical investigation provides function and sub-organelle location information on a repertoire of cardiac mitochondrial proteins sensitive to ischemia reperfusion stress and highlights protein clusters potentially involved in mitochondrial dysfunction in the setting of ischemic injury.

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Binding mitochondria to cryogel monoliths allows detection of proteins specifically released following permeability transition

Cited by 20 publications

References 50 publications

Differential Permeabilization Effects of Ca2+ and Valinomycin on the Inner and Outer Mitochondrial Membranes as Revealed by Proteomics Analysis of Proteins Released from Mitochondria

Differential Permeabilization Effects of Ca2+ and Valinomycin on the Inner and Outer Mitochondrial Membranes as Revealed by Proteomics Analysis of Proteins Released from Mitochondria

Macroporous gels prepared at subzero temperatures as novel materials for chromatography of particulate‐containing fluids and cell culture applications

Altered Proteome Biology of Cardiac Mitochondria Under Stress Conditions

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