Small mitochondrial protein NERCLIN regulates cardiolipin homeostasis and mitochondrial ultrastructure

ABSTRACTCardiolipin (CL) is an essential phospholipid for mitochondrial structure and function. Here we present a small mitochondrial protein, NERCLIN, as a negative regulator of CL homeostasis and mitochondrial ultrastructure. Primate-specific NERCLIN is expressed ubiquitously from GRPEL2 locus on a tightly regulated low level, but induced by heat stress. NERCLIN overexpression severely disrupts mitochondrial cristae structure and induces mitochondrial fragmentation. Proximity labeling suggested interactions of NERCLIN with CL synthesis and prohibitin complexes on the matrix side of the inner mitochondrial membrane. Lipid analysis indicated that NERCLIN regulates mitochondrial CL content. The regulation may occur directly through interaction with PTPMT1, a proximal partner on the CL synthesis pathway, as its product phosphatidylglycerol was also reduced by NERCLIN. We propose that NERCLIN contributes to stress-induced adaptation of mitochondrial dynamics and turnover by regulating the mitochondrial CL content. Our findings add NERCLIN to the group of recently identified small mitochondrial proteins with important regulatory functions.

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Skeletal Phenotypes Due to Abnormalities in Mitochondrial Protein Homeostasis and Import

International Journal of Molecular Sciences ◽

10.3390/ijms21218327 ◽

2020 ◽

Vol 21 (21) ◽

pp. 8327

Author(s):

Tian Zhao ◽

Caitlin Goedhart ◽

Gerald Pfeffer ◽

Steven C Greenway ◽

Matthew Lines ◽

...

Keyword(s):

Mitochondrial Dysfunction ◽

Mitochondrial Disease ◽

Protein Import ◽

Mitochondrial Protein ◽

Membrane Lipid ◽

Disease Genes ◽

Protein Homeostasis ◽

Inner Mitochondrial Membrane ◽

Mitochondrial Functions ◽

Insight Into

Mitochondrial disease represents a collection of rare genetic disorders caused by mitochondrial dysfunction. These disorders can be quite complex and heterogeneous, and it is recognized that mitochondrial disease can affect any tissue at any age. The reasons for this variability are not well understood. In this review, we develop and expand a subset of mitochondrial diseases including predominantly skeletal phenotypes. Understanding how impairment ofdiverse mitochondrial functions leads to a skeletal phenotype will help diagnose and treat patients with mitochondrial disease and provide additional insight into the growing list of human pathologies associated with mitochondrial dysfunction. The underlying disease genes encode factors involved in various aspects of mitochondrial protein homeostasis, including proteases and chaperones, mitochondrial protein import machinery, mediators of inner mitochondrial membrane lipid homeostasis, and aminoacylation of mitochondrial tRNAs required for translation. We further discuss a complex of frequently associated phenotypes (short stature, cataracts, and cardiomyopathy) potentially explained by alterations to steroidogenesis, a process regulated by mitochondria. Together, these observations provide novel insight into the consequences of impaired mitochondrial protein homeostasis.

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Transmembrane BAX Inhibitor-1 Motif Containing Protein 5 (TMBIM5) Sustains Mitochondrial Structure, Shape, and Function by Impacting the Mitochondrial Protein Synthesis Machinery

Cells ◽

10.3390/cells9102147 ◽

2020 ◽

Vol 9 (10) ◽

pp. 2147

Author(s):

Bruno Seitaj ◽

Felicia Maull ◽

Li Zhang ◽

Verena Wüllner ◽

Christina Wolf ◽

...

Keyword(s):

Cell Death ◽

Protein Synthesis ◽

Mitochondrial Membrane ◽

Mitochondrial Protein ◽

Mitochondrial Protein Synthesis ◽

Inner Mitochondrial Membrane ◽

Mitochondrial Structure ◽

Protein Synthesis Machinery ◽

Atp Generation ◽

And Function

The Transmembrane Bax Inhibitor-1 motif (TMBIM)-containing protein family is evolutionarily conserved and has been implicated in cell death susceptibility. The only member with a mitochondrial localization is TMBIM5 (also known as GHITM or MICS1), which affects cristae organization and associates with the Parkinson’s disease-associated protein CHCHD2 in the inner mitochondrial membrane. We here used CRISPR-Cas9-mediated knockout HAP1 cells to shed further light on the function of TMBIM5 in physiology and cell death susceptibility. We found that compared to wild type, TMBIM5-knockout cells were smaller and had a slower proliferation rate. In these cells, mitochondria were more fragmented with a vacuolar cristae structure. In addition, the mitochondrial membrane potential was reduced and respiration was attenuated, leading to a reduced mitochondrial ATP generation. TMBIM5 did not associate with Mic10 and Mic60, which are proteins of the mitochondrial contact site and cristae organizing system (MICOS), nor did TMBIM5 knockout affect their expression levels. TMBIM5-knockout cells were more sensitive to apoptosis elicited by staurosporine and BH3 mimetic inhibitors of Bcl-2 and Bcl-XL. An unbiased proteomic comparison identified a dramatic downregulation of proteins involved in the mitochondrial protein synthesis machinery in TMBIM5-knockout cells. We conclude that TMBIM5 is important to maintain the mitochondrial structure and function possibly through the control of mitochondrial biogenesis.

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Mitochondrial dynamics: overview of molecular mechanisms

Essays in Biochemistry ◽

10.1042/ebc20170104 ◽

2018 ◽

Vol 62 (3) ◽

pp. 341-360 ◽

Cited By ~ 197

Author(s):

Lisa Tilokani ◽

Shun Nagashima ◽

Vincent Paupe ◽

Julien Prudent

Keyword(s):

Membrane Fusion ◽

Mitochondrial Membrane ◽

Molecular Mechanisms ◽

Calcium Influx ◽

Mitochondrial Dynamics ◽

Mitochondrial Fission ◽

Membrane Lipid ◽

Inner Mitochondrial Membrane ◽

Step Process ◽

Shape Distribution

Mitochondria are highly dynamic organelles undergoing coordinated cycles of fission and fusion, referred as ‘mitochondrial dynamics’, in order to maintain their shape, distribution and size. Their transient and rapid morphological adaptations are crucial for many cellular processes such as cell cycle, immunity, apoptosis and mitochondrial quality control. Mutations in the core machinery components and defects in mitochondrial dynamics have been associated with numerous human diseases. These dynamic transitions are mainly ensured by large GTPases belonging to the Dynamin family. Mitochondrial fission is a multi-step process allowing the division of one mitochondrion in two daughter mitochondria. It is regulated by the recruitment of the GTPase Dynamin-related protein 1 (Drp1) by adaptors at actin- and endoplasmic reticulum-mediated mitochondrial constriction sites. Drp1 oligomerization followed by mitochondrial constriction leads to the recruitment of Dynamin 2 to terminate membrane scission. Inner mitochondrial membrane constriction has been proposed to be an independent process regulated by calcium influx. Mitochondrial fusion is driven by a two-step process with the outer mitochondrial membrane fusion mediated by mitofusins 1 and 2 followed by inner membrane fusion, mediated by optic atrophy 1. In addition to the role of membrane lipid composition, several members of the machinery can undergo post-translational modifications modulating these processes. Understanding the molecular mechanisms controlling mitochondrial dynamics is crucial to decipher how mitochondrial shape meets the function and to increase the knowledge on the molecular basis of diseases associated with morphology defects. This article will describe an overview of the molecular mechanisms that govern mitochondrial fission and fusion in mammals.

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Residues of Tim44 Involved in both Association with the Translocon of the Inner Mitochondrial Membrane and Regulation of Mitochondrial Hsp70 Tethering

Molecular and Cellular Biology ◽

10.1128/mcb.00007-08 ◽

2008 ◽

Vol 28 (13) ◽

pp. 4424-4433 ◽

Cited By ~ 32

Author(s):

Dirk Schiller ◽

Yu Chin Cheng ◽

Qinglian Liu ◽

William Walter ◽

Elizabeth A. Craig

Keyword(s):

Amino Acid ◽

Protein Import ◽

Protein Translocation ◽

Mitochondrial Protein ◽

Inner Mitochondrial Membrane ◽

The Matrix ◽

Mitochondrial Hsp70 ◽

Amino Acid Segment

ABSTRACT Translocation of proteins from the cytosol across the mitochondrial inner membrane is driven by the action of the import motor, which is associated with the translocon on the matrix side of the membrane. It is well established that an essential peripheral membrane protein, Tim44, tethers mitochondrial Hsp70 (mtHsp70), the core of the import motor, to the translocon. This Tim44-mtHsp70 interaction, which can be recapitulated in vitro, is destabilized by binding of mtHsp70 to a substrate polypeptide. Here we report that the N-terminal 167-amino-acid segment of mature Tim44 is sufficient for both interaction with mtHsp70 and destabilization of a Tim44-mtHsp70 complex caused by client protein binding. Amino acid alterations within a 30-amino-acid segment affected both the release of mtHsp70 upon peptide binding and the interaction of Tim44 with the translocon. Our results support the idea that Tim44 plays multiple roles in mitochondrial protein import by recruiting Ssc1 and its J protein cochaperone to the translocon and coordinating their interactions to promote efficient protein translocation in vivo.

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Effect of denervation-induced muscle disuse on mitochondrial protein import

AJP Cell Physiology ◽

10.1152/ajpcell.00181.2010 ◽

2011 ◽

Vol 300 (1) ◽

pp. C138-C145 ◽

Cited By ~ 38

Author(s):

Kaustabh Singh ◽

David A. Hood

Keyword(s):

Protein Import ◽

Mitochondrial Protein ◽

Ros Generation ◽

Ros Production ◽

Mitochondrial Protein Import ◽

Mitochondrial Content ◽

Muscle Disuse ◽

The Matrix ◽

And Function ◽

Mitochondrial Heat Shock Protein

This study determined whether muscle disuse affects mitochondrial protein import and whether changes in protein import are related to mitochondrial content and function. Protein import was measured using a model of unilateral peroneal nerve denervation in rats for 3 ( n = 10), 7 ( n = 12), or 14 ( n = 14) days. We compared the import of preproteins into the matrix of subsarcolemmal (SS) and intermyofibrillar (IMF) mitochondria isolated from the denervated and the contralateral control tibialis anterior muscles. Denervation led to 50% and 29% reductions in protein import after 14 days of disuse in SS and IMF mitochondria, respectively. This was accompanied by significant decreases in mitochondrial state 3 respiration, muscle mass, and whole muscle cytochrome c oxidase activity. To investigate the mechanisms involved, we assessed disuse-related changes in 1) protein import machinery components and 2) mitochondrial function, reflected by respiration and reactive oxygen species (ROS) production. Denervation significantly reduced the expression of translocases localized in the inner membrane (Tim23), outer membrane (Tom20), and mitochondrial heat shock protein 70 (mtHsp70), especially in the SS subfraction. Denervation also resulted in elevated ROS generation, and exogenous ROS was found to markedly reduce protein import. Thus our data indicate that protein import kinetics are closely related to alterations in mitochondrial respiratory capacity ( r = 0.95) and are negatively impacted by ROS. Deleterious changes in the protein import system likely facilitate the reduction in mitochondrial content and the increase in organelle dysfunction (i.e., increased ROS production and decreased respiration) during chronic disuse, which likely contribute to the activation of degradative pathways leading to muscle atrophy.

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MICOS and phospholipid transfer by Ups2–Mdm35 organize membrane lipid synthesis in mitochondria

The Journal of Cell Biology ◽

10.1083/jcb.201602007 ◽

2016 ◽

Vol 213 (5) ◽

pp. 525-534 ◽

Cited By ~ 84

Author(s):

Mari J. Aaltonen ◽

Jonathan R. Friedman ◽

Christof Osman ◽

Bénédicte Salin ◽

Jean-Paul di Rago ◽

...

Keyword(s):

Lipid Synthesis ◽

Membrane Lipid ◽

Intermembrane Space ◽

Lipid Transfer ◽

Inner Mitochondrial Membrane ◽

Phospholipid Transfer ◽

Mitochondrial Intermembrane Space ◽

In Trans ◽

And Function ◽

Specific Lipid

Mitochondria exert critical functions in cellular lipid metabolism and promote the synthesis of major constituents of cellular membranes, such as phosphatidylethanolamine (PE) and phosphatidylcholine. Here, we demonstrate that the phosphatidylserine decarboxylase Psd1, located in the inner mitochondrial membrane, promotes mitochondrial PE synthesis via two pathways. First, Ups2–Mdm35 complexes (SLMO2–TRIAP1 in humans) serve as phosphatidylserine (PS)-specific lipid transfer proteins in the mitochondrial intermembrane space, allowing formation of PE by Psd1 in the inner membrane. Second, Psd1 decarboxylates PS in the outer membrane in trans, independently of PS transfer by Ups2–Mdm35. This latter pathway requires close apposition between both mitochondrial membranes and the mitochondrial contact site and cristae organizing system (MICOS). In MICOS-deficient cells, limiting PS transfer by Ups2–Mdm35 and reducing mitochondrial PE accumulation preserves mitochondrial respiration and cristae formation. These results link mitochondrial PE metabolism to MICOS, combining functions in protein and lipid homeostasis to preserve mitochondrial structure and function.

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A Conserved Mitochondrial Chaperone-Protease Complex Involved in Protein Homeostasis

Frontiers in Molecular Biosciences ◽

10.3389/fmolb.2021.767088 ◽

2021 ◽

Vol 8 ◽

Author(s):

Mauro Serricchio ◽

Peter Bütikofer

Keyword(s):

Quality Control ◽

Protein Complex ◽

Elevated Temperatures ◽

Protein Complexes ◽

Mitochondrial Dynamics ◽

Mitochondrial Protein ◽

Culture Conditions ◽

Inner Mitochondrial Membrane ◽

Knock Out ◽

Mitochondrial Functions

Mitochondria are essential organelles involved in cellular energy production. The inner mitochondrial membrane protein stomatin-like protein 2 (SLP-2) is a member of the SPFH (stomatin, prohibitin, flotilin, and HflK/C) superfamily and binds to the mitochondrial glycerophospholipid cardiolipin, forming cardiolipin-enriched membrane domains to promote the assembly and/or stabilization of protein complexes involved in oxidative phosphorylation. In addition, human SLP-2 anchors a mitochondrial processing complex required for proteolytic regulation of proteins involved in mitochondrial dynamics and quality control. We now show that deletion of the gene encoding the Trypanosoma brucei homolog TbSlp2 has no effect on respiratory protein complex stability and mitochondrial functions under normal culture conditions and is dispensable for growth of T. brucei parasites. In addition, we demonstrate that TbSlp2 binds to the metalloprotease TbYme1 and together they form a large mitochondrial protein complex. The two proteins negatively regulate each other’s expression levels by accelerating protein turnover. Furthermore, we show that TbYme1 plays a role in heat-stress resistance, as TbYme1 knock-out parasites displayed mitochondrial fragmentation and loss of viability when cultured at elevated temperatures. Unbiased interaction studies uncovered putative TbYme1 substrates, some of which were differentially affected by the absence of TbYme1. Our results support emerging evidence for the presence of mitochondrial quality control pathways in this ancient eukaryote.

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SnoN as a novel negative regulator of TGF-β/Smad signaling: a target for tailoring organ fibrosis

AJP Heart and Circulatory Physiology ◽

10.1152/ajpheart.00453.2014 ◽

2015 ◽

Vol 308 (2) ◽

pp. H75-H82 ◽

Cited By ~ 18

Author(s):

Matthew R. Zeglinski ◽

Mark Hnatowich ◽

Davinder S. Jassal ◽

Ian M. C. Dixon

Keyword(s):

Transforming Growth Factor ◽

Tissue Injury ◽

Transforming Growth Factor Β ◽

Negative Regulator ◽

Matricellular Protein ◽

Matrix Remodeling ◽

Structural Support ◽

The Matrix ◽

Protein Components ◽

And Function

Remodeling of the extracellular matrix is beneficial during the acute wound healing stage following tissue injury. In the short term, resident fibroblasts and myofibroblasts regulate the matrix remodeling process through production of matricellular protein components that provide structural support to the damaged tissue. This process is largely governed by the transforming growth factor-β1 (TGF-β1) pathway, a critical mediator of the remodeling process. In the long term, chronic activation of the TGF-β1 pathway promotes excessive synthesis and deposition of matrix proteins, including fibrillar collagens, which ultimately leads to organ failure. SnoN (and its alternatively-spliced isoforms SnoN2, SnoA, and SnoI) is one of four members of a family of negative regulators of TGF-β1 signaling that includes Ski and functional Smad-suppressing elements on chromosomes 15 and 18. SnoN has been shown to be structurally and functionally similar to Ski and has been demonstrated to directly interact with Ski to abrogate gene expression. Despite this, little progress has been made in delineating a specific role for SnoN in the regulation of myofibroblast phenotype and function. This review outlines the current body of knowledge of what we refer to as the “Ski-Sno superfamily,” with a focus on the structural and functional importance of SnoN in mediating the fibrotic response by myofibroblasts following tissue injury.

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Identification of Tam41 maintaining integrity of the TIM23 protein translocator complex in mitochondria

The Journal of Cell Biology ◽

10.1083/jcb.200603087 ◽

2006 ◽

Vol 174 (5) ◽

pp. 631-637 ◽

Cited By ~ 77

Author(s):

Yasushi Tamura ◽

Yoshihiro Harada ◽

Koji Yamano ◽

Kazuaki Watanabe ◽

Daigo Ishikawa ◽

...

Keyword(s):

Protein Import ◽

Mitochondrial Protein ◽

Molecular Size ◽

Yeast Cells ◽

Temperature Sensitive ◽

Inner Mitochondrial Membrane ◽

Mitochondrial Protein Import ◽

The Matrix

Newly synthesized mitochondrial proteins are imported into mitochondria with the aid of protein translocator complexes in the outer and inner mitochondrial membranes. We report the identification of yeast Tam41, a new member of mitochondrial protein translocator systems. Tam41 is a peripheral inner mitochondrial membrane protein facing the matrix. Disruption of the TAM41 gene led to temperature-sensitive growth of yeast cells and resulted in defects in protein import via the TIM23 translocator complex at elevated temperature both in vivo and in vitro. Although Tam41 is not a constituent of the TIM23 complex, depletion of Tam41 led to a decreased molecular size of the TIM23 complex and partial aggregation of Pam18 and -16. Import of Pam16 into mitochondria without Tam41 was retarded, and the imported Pam16 formed aggregates in vitro. These results suggest that Tam41 facilitates mitochondrial protein import by maintaining the functional integrity of the TIM23 protein translocator complex from the matrix side of the inner membrane.

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Global Regulatory Functions of Oaf1p and Pip2p (Oaf2p), Transcription Factors That Regulate Genes Encoding Peroxisomal Proteins in Saccharomyces cerevisiae

Molecular and Cellular Biology ◽

10.1128/mcb.18.11.6560 ◽

1998 ◽

Vol 18 (11) ◽

pp. 6560-6570 ◽

Cited By ~ 101

Author(s):

Igor V. Karpichev ◽

Gillian M. Small

Keyword(s):

Saccharomyces Cerevisiae ◽

Transcription Factors ◽

Binding Site ◽

Consensus Sequence ◽

Mitochondrial Protein ◽

Northern Analysis ◽

Consensus Binding Site ◽

Genes Encoding ◽

And Function ◽

Regulatory Functions

ABSTRACT Two transcription factors, Oaf1p and Pip2p (Oaf2p), are key components in the pathway by which several Saccharomyces cerevisiae genes encoding peroxisomal proteins are activated in the presence of a fatty acid such as oleate. By searching the S. cerevisiae genomic database for the consensus sequence that acts as a target for these transcription factors, we identified 40 genes that contain a putative Oaf1p-Pip2p binding site in their promoter region. Quantitative Northern analysis confirmed that the expression of 22 of the genes identified is induced by oleate and that either one or both of these transcription factors are required for the activation. In addition to known peroxisomal proteins, the regulated genes encode novel peroxisomal proteins, a mitochondrial protein, and proteins of unknown location and function. We demonstrate that Oaf1p regulates certain genes in the absence of Pip2p and that both of these transcription factors play a role in maintaining the glucose-repressed state of one gene. Furthermore, we provide evidence that the defined consensus binding site is not required for the regulation of certain oleate-responsive genes.

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