Signal Presequences Increase Mitochondrial Permeability and Open the Multiple Conductance Channel

Kushnareva, Yulia; Campo, María Luisa; Kinnally, Kathleen W.; Sokolove, Patricia M.

doi:10.1006/abbi.1999.1190

Cited by 37 publications

(17 citation statements)

References 41 publications

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“…Something similar occurs in the case of TIM23. Whereas both isolated Tim23p and purified inner membranes form an open pore when reconstituted into bilayers or proteoliposomes (18,33), our studies with mitoplasts account for silent membranes that only reveal TIM23 channel activity in the presence of amino-terminal signal peptides (39 with the observation that transitions between noncontiguous current levels are recurrent and become increasingly frequent as the channel grows to be open 4 these data stand for a large, single dynamic channel with a cooperative behavior.…”

Section: Discussionmentioning

confidence: 68%

Awaking TIM22, a Dynamic Ligand-gated Channel for Protein Insertion in the Mitochondrial Inner Membrane

Peixoto

Graña

Roy

et al. 2007

Journal of Biological Chemistry

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Aqueous channels are at the core of the translocase of the outer membrane (TOM) and the translocase of the inner membrane for the transport of preproteins (TIM23), the translocases mediating the transport of proteins across the outer and inner mitochondrial membranes. Yet, the existence of a channel associated to the translocase of the inner membrane for the insertion of multitopic protein (TIM22) complex has been arguable, as its function relates to the insertion of multispanning proteins into the inner membrane. For the first time, we report conditions for detecting a channel activity associated to the TIM22 translocase in organelle, i.e. intact mitoplasts. An internal signal peptide in the intermembrane space of mitochondria is a requisite to inducing this channel, which is otherwise silent. The channel showed slightly cationic and high conductance activity of 1000 pS with a predominant half-open substate. Despite their different composition, the channels of the three mitochondrial translocases were thus remarkably similar, in agreement with their common task as pores transiently trapping proteins en route to their final destination. The opening of the TIM22 channel was a step-up process depending on the signal peptide concentration. Interestingly, low membrane potentials kept the channel fully open, providing a threshold level of the peptide is present. Our results portray TIM22 as a dynamic channel solely active in the presence of its cargo proteins. In its fully open conformation, favored by the combined action of internal signal peptide and low membrane potential, the channel could embrace the in-transit protein. As insertion progressed and initial interaction with the signal peptide faded, the channel would close, sustaining its role as a shunt that places trapped proteins into the membrane.Most cells depend upon mitochondria to accomplish their programmed role. Along with mitochondria's own biogenesis, functioning and death entirely rely on protein translocation. The elevated protein content of mitochondria (ϳ700 different proteins in yeast (1)) and the four different compartments enclosed by their two membranes add to the complexity of protein trafficking pathways in this organelle. In addition, the need to maintain a low permeability of the inner membrane for coupling oxidative phosphorylation must be balanced with the passage of large molecules as proteins through water-filled channels. Focused on the route to their final destination inside mitochondria, three multisubunit complexes or translocases are depicted as the main machineries for specific recognition, import, and sorting of precursor proteins synthesized in the cytoplasm (for review, see Refs. 2-7).The translocase of the outer membrane (TOM) 3 is the common gate for the transport of every mitochondrial protein across the outer membrane (for review, see Ref. 8), and the TIM23 translocase imports matrix-targeted preproteins with cleavable amino-terminal presequences. Few proteins of the inner membrane (IM) containing a single transmembrane seg...

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Section: Discussionmentioning

confidence: 68%

Awaking TIM22, a Dynamic Ligand-gated Channel for Protein Insertion in the Mitochondrial Inner Membrane

Peixoto

Graña

Roy

et al. 2007

Journal of Biological Chemistry

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“…Mitochondria, 0.5 mg of protein/ml (Mc), and 2.5 M rotenone (R) were added to the incubation medium composed of 125 mM KNO 3 , 50 M EGTA, 10 mM Hepes-Tris buffer, pH 7.2, and supplemented with 5 mM succinate-Tris, pH 7.2; a and b, 10 M safranin O was added to monitor the inner membrane potential; 3WP and 5WP, 3 and 5 g/ml BTM-WP1, respectively; FCCP, 1 M FCCP; 2P and 3P, 2 and 3 g/ml BTM-P1, respectively. dependent swelling of mitochondria at concentrations of KCl lower than 50 mM; higher concentrations of KCl inhibited all these effects (40).…”

Section: Discussionmentioning

confidence: 90%

“…Most of the signal sequences of mitochondrial proteins with a net positive charge have been shown to induce potentialdependent swelling of mitochondria and to activate the mitochondrial multiple conductance channel (38,40) that has been suggested to be distinct from both the classical permeability transition pore and the mastoparan-induced permeability increase (40). Many membrane-active peptides have been reported to function as a defense system against bacteria in plants and animals (41)(42)(43).…”

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confidence: 99%

Mitochondria Permeabilization by a Novel Polycation Peptide BTM-P1

Lemeshko

Arias

Ordúz

2005

Journal of Biological Chemistry

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Bacillus thuringiensis subsp. medellin is known to produce the Cry11Bb protein of 94 kDa, which is toxic for mosquito larvae due to permeabilization of the plasma membrane of midgut epithelial cells. Earlier we found that a 2.8-kDa novel peptide BTM-P1, which was artificially synthesized taking into account the primary structure of Cry11Bb endotoxin, is active against several species of bacteria. In this work we show that BTM-P1 induces cyclosporin A-insensitive swelling of rat liver mitochondria in various salt solutions but not in the sucrose medium. Inorganic phosphate and Ca 2؉ significantly increased this effect of the peptide. The uncoupling action of BTM-P1 on oxidative phosphorylation was stronger in the potassiumcontaining media and correlated with a decrease of the inner membrane potential of mitochondria. In isotonic KNO 3 , KCl, or NH 4 NO 3 media, a complete drop of the inner membrane potential was observed at 1-2 g/ml of the peptide. The peptide-induced swelling was increased by energization of mitochondria in the potassium-containing media, but it was inhibited in the NaNO 3 , NH 4 NO 3 , and Tris-NO 3 media. All mitochondrial effects of the peptide were completely prevented by adding a single N-terminal tryptophan residue to the peptide sequence. We suggest a mechanism of membrane permeabilization that includes a transmembrane-and surface potential-dependent insertion of the polycation peptide into the lipid bilayer and its oligomerization leading to formation of ion channels and also to the mitochondrial permeability transition pore opening in a cyclosporin A-insensitive manner.

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“…For example, although pore-like conductance can be reconstituted with purified ANT alone or in combination with other proteins [13,19,21], MPT pore conductance occurs in triple ANT knockout yeast [22]. Mitochondrial targeting peptides and exogenous pore-forming amphipathic peptides, such as alamethicin and mastoparan, also induce onset of a Ca 2+ -dependent and CsA-sensitive MPT [9,23,24].…”

Section: Models Of Permeability Transition Pore Structure and Functionmentioning

confidence: 99%

Cyclophilin D as a Drug Target

Waldmeier¹,

Zimmermann²,

Qian³

et al. 2003

CMC

214

146

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The mitochondrial permeability transition (MPT) plays an important role in damage-induced cell death, and agents inhibiting the MPT may have a therapeutic potential for treating human conditions such as ischemia/reperfusion injury, trauma, and neurodegenerative diseases. The mitochondrial matrix protein, cyclophilin D (CYP D), a member of a family of highly homologous peptidylprolyl cis-trans isomerases (PPIases), plays a decisive role in MPT, being an integral constituent of the MPT pore. Other putative MPT pore proteins include the adenine nucleotide translocator (ANT) and the voltage-dependent anion channel (VDAC). In an alternative model, the MPT pore is formed by clusters of misfolded membrane proteins outlining aqueous channels that are regulated by CYP D and other chaperone-like proteins. Like cyclophilin A (CYP A) and other cyclophilin family members, CYP D is targeted by the immunosuppressant cyclosporin A (CsA). CsA is cytoprotective in many cellular and animal models, but protection may result from either inhibition of the MPT through an interaction with CYP D or inhibition of calcineurin-mediated dephosphorylation of BAD through an interaction with CYP A. The relevance of MPT inhibition by CsA for its cytoprotective effects is well documented in many cellular models. Mechanisms of action in vivo are more difficult to define, and accordingly the evidence is as yet less compelling in in vivo animal models of ischemia/reperfusion injury, trauma and neurodegenerative diseases. Notwithstanding, CYP D is a drug target of high interest. Structural considerations suggest feasibility of designing CYP D ligands without immunosuppressant properties. This is highly desirable, since they have the potential of being useful therapeutic agents in a variety of disease states. It might be a tougher challenge to obtain compounds specific for CYP D vs. other cyclophilins, and/or of small molecular weight, allowing brain penetration to make them suitable for treating neurodegenerative diseases.

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Signal Presequences Increase Mitochondrial Permeability and Open the Multiple Conductance Channel

Cited by 37 publications

References 41 publications

Awaking TIM22, a Dynamic Ligand-gated Channel for Protein Insertion in the Mitochondrial Inner Membrane

Awaking TIM22, a Dynamic Ligand-gated Channel for Protein Insertion in the Mitochondrial Inner Membrane

Mitochondria Permeabilization by a Novel Polycation Peptide BTM-P1

Cyclophilin D as a Drug Target

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