scholarly journals Tim29 is a novel subunit of the human TIM22 translocase and is involved in complex assembly and stability

eLife ◽  
2016 ◽  
Vol 5 ◽  
Author(s):  
Yilin Kang ◽  
Michael James Baker ◽  
Michael Liem ◽  
Jade Louber ◽  
Matthew McKenzie ◽  
...  
Keyword(s):  
Mitochondrial Membrane ◽  
Intermembrane Space ◽  
Outer Mitochondrial Membrane ◽  
Carrier Proteins ◽  
Mitochondrial Import ◽  
Inner Membrane ◽  
Mitochondrial Inner Membrane ◽  
C Terminus ◽  
The Stability ◽  
Human Complex

The TIM22 complex mediates the import of hydrophobic carrier proteins into the mitochondrial inner membrane. While the TIM22 machinery has been well characterised in yeast, the human complex remains poorly characterised. Here, we identify Tim29 (C19orf52) as a novel, metazoan-specific subunit of the human TIM22 complex. The protein is integrated into the mitochondrial inner membrane with it’s C-terminus exposed to the intermembrane space. Tim29 is required for the stability of the TIM22 complex and functions in the assembly of hTim22. Furthermore, Tim29 contacts the Translocase of the Outer Mitochondrial Membrane, TOM complex, enabling a mechanism for transport of hydrophobic carrier substrates across the aqueous intermembrane space. Identification of Tim29 highlights the significance of analysing mitochondrial import systems across phylogenetic boundaries, which can reveal novel components and mechanisms in higher organisms.

scholarly journals Divergent Small Tim Homologues Are Associated with TbTim17 and Critical for the Biogenesis of TbTim17 Protein Complexes in Trypanosoma brucei

mSphere ◽  
2018 ◽  
Vol 3 (3) ◽  
Author(s):  
Joseph T. Smith ◽  
Ujjal K. Singha ◽  
Smita Misra ◽  
Minu Chaudhuri
Keyword(s):  
Membrane Proteins ◽  
Trypanosoma Brucei ◽  
Mitochondrial Protein ◽  
Intermembrane Space ◽  
Inner Membrane ◽  
Content Type ◽  
Mitochondrial Inner Membrane ◽  
Mitochondrial Intermembrane Space ◽  
The Stability ◽  
Tim Proteins

ABSTRACT The small Tim proteins belong to a group of mitochondrial intermembrane space chaperones that aid in the import of mitochondrial inner membrane proteins with internal targeting signals. Trypanosoma brucei , the protozoan parasite that causes African trypanosomiasis, possesses multiple small Tim proteins that include homologues of T. brucei Tim9 (TbTim9) and Tim10 (TbTim10) and a unique small Tim that shares homology with both Tim8 and Tim13 (TbTim8/13). Here, we found that these three small TbTims are expressed as soluble mitochondrial intermembrane space proteins. Coimmunoprecipitation and mass spectrometry analysis showed that the small TbTims stably associated with each other and with TbTim17, the major component of the mitochondrial inner membrane translocase in T. brucei . Yeast two-hybrid analysis indicated direct interactions among the small TbTims; however, their interaction patterns appeared to be different from those of their counterparts in yeast and humans. Knockdown of the small TbTims reduced cell growth and decreased the steady-state level of TbTim17 and T. brucei ADP/ATP carrier (TbAAC), two polytopic mitochondrial inner membrane proteins. Knockdown of small TbTims also reduced the matured complexes of TbTim17 in mitochondria. Depletion of any of the small TbTims reduced TbTim17 import moderately but greatly hampered the stability of the TbTim17 complexes in T. brucei . Altogether, our results revealed that TbTim9, TbTim10, and TbTim8/13 interact with each other, associate with TbTim17, and play a crucial role in the integrity and maintenance of the levels of TbTim17 complexes. IMPORTANCE Trypanosoma brucei is the causative agent of African sleeping sickness. The parasite’s mitochondrion represents a useful source for potential chemotherapeutic targets. Similarly to yeast and humans, mitochondrial functions depend on the import of proteins that are encoded in the nucleus and made in the cytosol. Even though the machinery involved in this mitochondrial protein import process is becoming clearer in T. brucei , a comprehensive picture of protein complex composition and function is still lacking. In this study, we characterized three T. brucei small Tim proteins, TbTim9, TbTim10, and TbTim8/13. Although the parasite does not have the classical TIM22 complex that imports mitochondrial inner membrane proteins containing internal targeting signals in yeast or humans, we found that these small TbTims associate with TbTim17, the major subunit of the TbTIM complex in T. brucei , and play an essential role in the stability of the TbTim17 complexes. Therefore, these divergent proteins are critical for mitochondrial protein biogenesis in T. brucei .

scholarly journals Mitochondria Use Different Mechanisms for Transport of Multispanning Membrane Proteins through the Intermembrane Space

2003 ◽  
Vol 23 (21) ◽  
pp. 7818-7828 ◽  
Author(s):  
Ann E. Frazier ◽  
Agnieszka Chacinska ◽  
Kaye N. Truscott ◽  
Bernard Guiard ◽  
Nikolaus Pfanner ◽  
...  
Keyword(s):  
Outer Membrane ◽  
Heat Shock Protein 70 ◽  
Mitochondrial Outer Membrane ◽  
Intermembrane Space ◽  
Carrier Proteins ◽  
Inner Membrane ◽  
Tom Complex ◽  
Mitochondrial Inner Membrane ◽  
Integral Proteins ◽  
Matrix Heat

ABSTRACT The mitochondrial inner membrane contains numerous multispanning integral proteins. The precursors of these hydrophobic proteins are synthesized in the cytosol and therefore have to cross the mitochondrial outer membrane and intermembrane space to reach the inner membrane. While the import pathways of noncleavable multispanning proteins, such as the metabolite carriers, have been characterized in detail by the generation of translocation intermediates, little is known about the mechanism by which cleavable preproteins of multispanning proteins, such as Oxa1, are transferred from the outer membrane to the inner membrane. We have identified a translocation intermediate of the Oxa1 preprotein in the translocase of the outer membrane (TOM) and found that there are differences from the import mechanisms of carrier proteins. The intermembrane space domain of the receptor Tom22 supports the stabilization of the Oxa1 intermediate. Transfer of the Oxa1 preprotein to the inner membrane is not affected by inactivation of the soluble TIM complexes. Both the inner membrane potential and matrix heat shock protein 70 are essential to release the preprotein from the TOM complex, suggesting a close functional cooperation of the TOM complex and the presequence translocase of the inner membrane. We conclude that mitochondria employ different mechanisms for translocation of multispanning proteins across the aqueous intermembrane space.

scholarly journals Monolysocardiolipin (MLCL) interactions with mitochondrial membrane proteins

2020 ◽  
Vol 48 (3) ◽  
pp. 993-1004
Author(s):  
Anna L. Duncan
Keyword(s):  
Protein Interactions ◽  
Mitochondrial Membrane ◽  
Barth Syndrome ◽  
Mitochondrial Proteins ◽  
Mitochondrial Membranes ◽  
Inner Membrane ◽  
Computational Approaches ◽  
Mitochondrial Inner Membrane ◽  
Recent Developments ◽  
The Stability

Monolysocardiolipin (MLCL) is a three-tailed variant of cardiolipin (CL), the signature lipid of mitochondria. MLCL is not normally found in healthy tissue but accumulates in mitochondria of people with Barth syndrome (BTHS), with an overall increase in the MLCL:CL ratio. The reason for MLCL accumulation remains to be fully understood. The effect of MLCL build-up and decreased CL content in causing the characteristics of BTHS are also unclear. In both cases, an understanding of the nature of MLCL interaction with mitochondrial proteins will be key. Recent work has shown that MLCL associates less tightly than CL with proteins in the mitochondrial inner membrane, suggesting that MLCL accumulation is a result of CL degradation, and that the lack of MLCL–protein interactions compromises the stability of the protein-dense mitochondrial inner membrane, leading to a decrease in optimal respiration. There is some data on MLCL–protein interactions for proteins involved in the respiratory chain and in apoptosis, but there remains much to be understood regarding the nature of MLCL–protein interactions. Recent developments in structural, analytical and computational approaches mean that these investigations are now possible. Such an understanding will be key to further insights into how MLCL accumulation impacts mitochondrial membranes. In turn, these insights will help to support the development of therapies for people with BTHS and give a broader understanding of other diseases involving defective CL content.

scholarly journals A discrete pathway for the transfer of intermembrane space proteins across the outer membrane of mitochondria

2014 ◽  
Vol 25 (25) ◽  
pp. 3999-4009 ◽  
Author(s):  
Agnieszka Gornicka ◽  
Piotr Bragoszewski ◽  
Piotr Chroscicki ◽  
Lena-Sophie Wenz ◽  
Christian Schulz ◽  
...  
Keyword(s):  
Outer Membrane ◽  
Mitochondrial Membrane ◽  
Alternative Pathway ◽  
Intermembrane Space ◽  
Outer Mitochondrial Membrane ◽  
Membrane Translocation ◽  
Tom Complex ◽  
Precursor Proteins ◽  
Core Components

Mitochondrial proteins are synthesized on cytosolic ribosomes and imported into mitochondria with the help of protein translocases. For the majority of precursor proteins, the role of the translocase of the outer membrane (TOM) and mechanisms of their transport across the outer mitochondrial membrane are well recognized. However, little is known about the mode of membrane translocation for proteins that are targeted to the intermembrane space via the redox-driven mitochondrial intermembrane space import and assembly (MIA) pathway. On the basis of the results obtained from an in organello competition import assay, we hypothesized that MIA-dependent precursor proteins use an alternative pathway to cross the outer mitochondrial membrane. Here we demonstrate that this alternative pathway involves the protein channel formed by Tom40. We sought a translocation intermediate by expressing tagged versions of MIA-dependent proteins in vivo. We identified a transient interaction between our model substrates and Tom40. Of interest, outer membrane translocation did not directly involve other core components of the TOM complex, including Tom22. Thus MIA-dependent proteins take another route across the outer mitochondrial membrane that involves Tom40 in a form that is different from the canonical TOM complex.

scholarly journals YME1L controls the accumulation of respiratory chain subunits and is required for apoptotic resistance, cristae morphogenesis, and cell proliferation

2012 ◽  
Vol 23 (6) ◽  
pp. 1010-1023 ◽  
Author(s):  
Lukas Stiburek ◽  
Jana Cesnekova ◽  
Olga Kostkova ◽  
Daniela Fornuskova ◽  
Kamila Vinsova ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Membrane Protein ◽  
Respiratory Chain ◽  
Integral Membrane Protein ◽  
Intermembrane Space ◽  
Wild Type ◽  
Inner Membrane ◽  
Mitochondrial Inner Membrane ◽  
Human Orthologue ◽  
Excessive Accumulation

Mitochondrial ATPases associated with diverse cellular activities (AAA) proteases are involved in the quality control and processing of inner-membrane proteins. Here we investigate the cellular activities of YME1L, the human orthologue of the Yme1 subunit of the yeast i‑AAA complex, using stable short hairpin RNA knockdown and expression experiments. Human YME1L is shown to be an integral membrane protein that exposes its carboxy-terminus to the intermembrane space and exists in several complexes of 600–1100 kDa. The stable knockdown of YME1L in human embryonic kidney 293 cells led to impaired cell proliferation and apoptotic resistance, altered cristae morphology, diminished rotenone-sensitive respiration, and increased susceptibility to mitochondrial membrane protein carbonylation. Depletion of YME1L led to excessive accumulation of nonassembled respiratory chain subunits (Ndufb6, ND1, and Cox4) in the inner membrane. This was due to a lack of YME1L proteolytic activity, since the excessive accumulation of subunits was reversed by overexpression of wild-type YME1L but not a proteolytically inactive YME1L variant. Similarly, the expression of wild-type YME1L restored the lamellar cristae morphology of YME1L-deficient mitochondria. Our results demonstrate the importance of mitochondrial inner-membrane proteostasis to both mitochondrial and cellular function and integrity and reveal a novel role for YME1L in the proteolytic regulation of respiratory chain biogenesis.

scholarly journals Mitochondrial Membrane Dynamics—Functional Positioning of OPA1

Antioxidants ◽  
2018 ◽  
Vol 7 (12) ◽  
pp. 186 ◽  
Author(s):  
Hakjoo Lee ◽  
Yisang Yoon
Keyword(s):  
Mitochondrial Membrane ◽  
Proteolytic Cleavage ◽  
Membrane Dynamics ◽  
Mitochondrial Morphology ◽  
Inner Membrane ◽  
Mitochondrial Inner Membrane ◽  
Functional Differences ◽  
New Information ◽  
New Development

The maintenance of mitochondrial energetics requires the proper regulation of mitochondrial morphology, and vice versa. Mitochondrial dynamins control mitochondrial morphology by mediating fission and fusion. One of them, optic atrophy 1 (OPA1), is the mitochondrial inner membrane remodeling protein. OPA1 has a dual role in maintaining mitochondrial morphology and energetics through mediating inner membrane fusion and maintaining the cristae structure. OPA1 is expressed in multiple variant forms through alternative splicing and post-translational proteolytic cleavage, but the functional differences between these variants have not been completely understood. Recent studies generated new information regarding the role of OPA1 cleavage. In this review, we will first provide a brief overview of mitochondrial membrane dynamics by describing fission and fusion that are mediated by mitochondrial dynamins. The second part describes OPA1-mediated fusion and energetic maintenance, the role of OPA1 cleavage, and a new development in OPA1 function, in which we will provide new insight for what OPA1 does and what proteolytic cleavage of OPA1 is for.

scholarly journals Protection from α-Synuclein induced dopaminergic neurodegeneration by overexpression of the mitochondrial import receptor TOM20

2020 ◽  
Vol 6 (1) ◽  
Author(s):  
Briana R. De Miranda ◽  
Emily M. Rocha ◽  
Sandra L. Castro ◽  
J. Timothy Greenamyre
Keyword(s):  
Substantia Nigra ◽  
Mitochondrial Dysfunction ◽  
Dopaminergic Neurons ◽  
Mitochondrial Membrane ◽  
Expression System ◽  
Outer Mitochondrial Membrane ◽  
Mitochondrial Import ◽  
In Vivo Models ◽  
Dopaminergic Neurodegeneration ◽  
Import Receptor

AbstractDopaminergic neurons of the substantia nigra are selectively vulnerable to mitochondrial dysfunction, which is hypothesized to be an early and fundamental pathogenic mechanism in Parkinson’s disease (PD). Mitochondrial function depends on the successful import of nuclear-encoded proteins, many of which are transported through the TOM20–TOM22 outer mitochondrial membrane import receptor machinery. Recent data suggests that post-translational modifications of α-synuclein promote its interaction with TOM20 at the outer mitochondrial membrane and thereby inhibit normal protein import, leading to dysfunction, and death of dopaminergic neurons. As such, preservation of mitochondrial import in the face of α-synuclein accumulation might be a strategy to prevent dopaminergic neurodegeneration, however, this is difficult to assess using current in vivo models of PD. To this end, we established an exogenous co-expression system, utilizing AAV2 vectors to overexpress human α-synuclein and TOM20, individually or together, in the adult Lewis rat substantia nigra to assess whether TOM20 overexpression attenuates α-synuclein-induced dopaminergic neurodegeneration. Twelve weeks after viral injection, we observed that AAV2-TOM20 expression was sufficient to prevent loss of nigral dopaminergic neurons caused by AAV2-αSyn overexpression. The observed TOM20-mediated dopaminergic neuron preservation appeared to be due, in part, to the rescued expression (and presumed import) of nuclear-encoded mitochondrial electron transport chain proteins that were inhibited by α-synuclein overexpression. In addition, TOM20 overexpression rescued the expression of the chaperone protein GRP75/mtHSP70/mortalin, a stress-response protein involved in α-synuclein-induced injury. Collectively, these data indicate that TOM20 expression prevents α-synuclein-induced mitochondrial dysfunction, which is sufficient to rescue dopaminergic neurons in the adult rat brain.

scholarly journals Mitochondrial Protein Import

2004 ◽  
Vol 279 (44) ◽  
pp. 45701-45707 ◽  
Author(s):  
Masatoshi Esaki ◽  
Hidaka Shimizu ◽  
Tomoko Ono ◽  
Hayashi Yamamoto ◽  
Takashi Kanamori ◽  
...  
Keyword(s):  
Mitochondrial Membrane ◽  
Protein Import ◽  
Protein Translocation ◽  
Mitochondrial Protein ◽  
Intermembrane Space ◽  
Outer Mitochondrial Membrane ◽  
Mitochondrial Protein Import ◽  
Tom Complex ◽  
Membrane Complex ◽  
Cytosolic Side

Protein translocation across the outer mitochondrial membrane is mediated by the translocator called the TOM (translocase of the outer mitochondrial membrane) complex. The TOM complex possesses two presequence binding sites on the cytosolic side (thecissite) and on the intermembrane space side (thetranssite). Here we analyzed the requirement of presequence elements and subunits of the TOM complex for presequence binding to thecisandtranssites of the TOM complex. The N-terminal 14 residues of the presequence of subunit 9 of F0-ATPase are required for binding to thetranssite. The interaction between the presequence and thecissite is not sufficient to anchor the precursor protein to the TOM complex. Tom7 constitutes or is close to thetranssite and has overlapping functions with the C-terminal intermembrane space domain of Tom22 in the mitochondrial protein import.

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