scholarly journals Ups1p, a conserved intermembrane space protein, regulates mitochondrial shape and alternative topogenesis of Mgm1p

2006 ◽  
Vol 173 (5) ◽  
pp. 651-658 ◽  
Author(s):  
Hiromi Sesaki ◽  
Cory D. Dunn ◽  
Miho Iijima ◽  
Kelly A. Shepard ◽  
Michael P. Yaffe ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Mitochondrial Protein ◽  
Yeast Cells ◽  
Mitochondrial Morphology ◽  
Intermembrane Space ◽  
Genome Maintenance ◽  
Human Homologue ◽  
Inner Membrane ◽  
Rhomboid Protease ◽  
And Control

Mgm1p is a conserved dynamin-related GTPase required for fusion, morphology, inheritance, and the genome maintenance of mitochondria in Saccharomyces cerevisiae. Mgm1p undergoes unconventional processing to produce two functional isoforms by alternative topogenesis. Alternative topogenesis involves bifurcate sorting in the inner membrane and intramembrane proteolysis by the rhomboid protease Pcp1p. Here, we identify Ups1p, a novel mitochondrial protein required for the unique processing of Mgm1p and for normal mitochondrial shape. Our results demonstrate that Ups1p regulates the sorting of Mgm1p in the inner membrane. Consistent with its function, Ups1p is peripherally associated with the inner membrane in the intermembrane space. Moreover, the human homologue of Ups1p, PRELI, can fully replace Ups1p in yeast cells. Together, our findings provide a conserved mechanism for the alternative topogenesis of Mgm1p and control of mitochondrial morphology.

scholarly journals Prohibitins Regulate Membrane Protein Degradation by the m-AAA Protease in Mitochondria

1999 ◽  
Vol 19 (5) ◽  
pp. 3435-3442 ◽  
Author(s):  
Gregor Steglich ◽  
Walter Neupert ◽  
Thomas Langer
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cell Growth ◽  
Membrane Proteins ◽  
Eukaryotic Cells ◽  
Mitochondrial Morphology ◽  
Regulatory Effect ◽  
Inner Membrane ◽  
Functional Activities ◽  
Native Molecular Mass ◽  
Affect Cell Growth

ABSTRACT Prohibitins comprise a protein family in eukaryotic cells with potential roles in senescence and tumor suppression. Phb1p and Phb2p, members of the prohibitin family in Saccharomyces cerevisiae, have been implicated in the regulation of the replicative life span of the cells and in the maintenance of mitochondrial morphology. The functional activities of these proteins, however, have not been elucidated. We demonstrate here that prohibitins regulate the turnover of membrane proteins by the m-AAA protease, a conserved ATP-dependent protease in the inner membrane of mitochondria. The m-AAA protease is composed of the homologous subunits Yta10p (Afg3p) and Yta12p (Rca1p). Deletion ofPHB1 or PHB2 impairs growth of Δyta10 or Δyta12 cells but does not affect cell growth in the presence of the m-AAA protease. A prohibitin complex with a native molecular mass of approximately 2 MDa containing Phb1p and Phb2p forms a supercomplex with them-AAA protease. Proteolysis of nonassembled inner membrane proteins by the m-AAA protease is accelerated in mitochondria lacking Phb1p or Phb2p, indicating a negative regulatory effect of prohibitins on m-AAA protease activity. These results functionally link members of two conserved protein families in eukaryotes to the degradation of membrane proteins in mitochondria.

scholarly journals Mutant huntingtin disrupts mitochondrial proteostasis by interacting with TIM23

2019 ◽  
Vol 116 (33) ◽  
pp. 16593-16602 ◽  
Author(s):  
Svitlana Yablonska ◽  
Vinitha Ganesan ◽  
Lisa M. Ferrando ◽  
JinHo Kim ◽  
Anna Pyzel ◽  
...  
Keyword(s):  
Cell Lines ◽  
Protein Import ◽  
Mitochondrial Protein ◽  
Biological Significance ◽  
Intermembrane Space ◽  
Mutant Huntingtin ◽  
Mitochondrial Protein Import ◽  
Inner Membrane ◽  
Mitochondrial Proteome ◽  
Brain Tissues

Mutant huntingtin (mHTT), the causative protein in Huntington’s disease (HD), associates with the translocase of mitochondrial inner membrane 23 (TIM23) complex, resulting in inhibition of synaptic mitochondrial protein import first detected in presymptomatic HD mice. The early timing of this event suggests that it is a relevant and direct pathophysiologic consequence of mHTT expression. We show that, of the 4 TIM23 complex proteins, mHTT specifically binds to the TIM23 subunit and that full-length wild-type huntingtin (wtHTT) and mHTT reside in the mitochondrial intermembrane space. We investigated differences in mitochondrial proteome between wtHTT and mHTT cells and found numerous proteomic disparities between mHTT and wtHTT mitochondria. We validated these data by quantitative immunoblotting in striatal cell lines and human HD brain tissue. The level of soluble matrix mitochondrial proteins imported through the TIM23 complex is lower in mHTT-expressing cell lines and brain tissues of HD patients compared with controls. In mHTT-expressing cell lines, membrane-bound TIM23-imported proteins have lower intramitochondrial levels, whereas inner membrane multispan proteins that are imported via the TIM22 pathway and proteins integrated into the outer membrane generally remain unchanged. In summary, we show that, in mitochondria, huntingtin is located in the intermembrane space, that mHTT binds with high-affinity to TIM23, and that mitochondria from mHTT-expressing cells and brain tissues of HD patients have reduced levels of nuclearly encoded proteins imported through TIM23. These data demonstrate the mechanism and biological significance of mHTT-mediated inhibition of mitochondrial protein import, a mechanism likely broadly relevant to other neurodegenerative diseases.

scholarly journals Tim18p, a New Subunit of the TIM22 Complex That Mediates Insertion of Imported Proteins into the Yeast Mitochondrial Inner Membrane

2000 ◽  
Vol 20 (4) ◽  
pp. 1187-1193 ◽  
Author(s):  
Carla M. Koehler ◽  
Michael P. Murphy ◽  
Nikolaus A. Bally ◽  
Danielle Leuenberger ◽  
Wolfgang Oppliger ◽  
...  
Keyword(s):  
Protein Complexes ◽  
Integral Membrane Protein ◽  
Yeast Cells ◽  
Temperature Sensitive ◽  
Intermembrane Space ◽  
Inner Membrane ◽  
Protein Insertion ◽  
Mitochondrial Inner Membrane ◽  
Precursor Proteins ◽  
Synthetically Lethal

ABSTRACT Import of carrier proteins from the cytoplasm into the mitochondrial inner membrane of yeast is mediated by a distinct system consisting of two soluble 70-kDa protein complexes in the intermembrane space and a 300-kDa complex in the inner membrane, the TIM22 complex. The TIM22 complex contains the peripheral subunits Tim9p, Tim10p, and Tim12p and the integral membrane subunits Tim22p and Tim54p. We identify here an additional subunit, an 18-kDa integral membrane protein termed Tim18p. This protein is made as a 21.9-kDa precursor which is imported into mitochondria and processed to its mature form. When mitochondria are gently solubilized, Tim18p comigrates with the other subunits of the TIM22 complex on nondenaturing gels and is coimmunoprecipitated with Tim54p and Tim12p. Tim18p does not cofractionate with the TIM23 complex upon immunoprecipitation or nondenaturing gel electrophoresis. Deletion of Tim18p decreases the growth rate of yeast cells by a factor of two and is synthetically lethal with temperature-sensitive mutations in Tim9p or Tim10p. It also impairs the import of several precursor proteins into isolated mitochondria, and lowers the apparent mass of the TIM22 complex. We suggest that Tim18p functions in the assembly and stabilization of the TIM22 complex but does not directly participate in protein insertion into the inner membrane.

scholarly journals Phosphatidylserine transport by Ups2–Mdm35 in respiration-active mitochondria

2016 ◽  
Vol 214 (1) ◽  
pp. 77-88 ◽  
Author(s):  
Non Miyata ◽  
Yasunori Watanabe ◽  
Yasushi Tamura ◽  
Toshiya Endo ◽  
Osamu Kuge
Keyword(s):  
Yeast Cells ◽  
Intermembrane Space ◽  
Diauxic Shift ◽  
Inner Membrane ◽  
Mitochondrial Inner Membrane ◽  
Mitochondrial Functions ◽  
Phospholipid Transfer ◽  
Metabolic Transition ◽  
Mitochondrial Intermembrane Space

Phosphatidylethanolamine (PE) is an essential phospholipid for mitochondrial functions and is synthesized mainly by phosphatidylserine (PS) decarboxylase at the mitochondrial inner membrane. In Saccharomyces cerevisiae, PS is synthesized in the endoplasmic reticulum (ER), such that mitochondrial PE synthesis requires PS transport from the ER to the mitochondrial inner membrane. Here, we provide evidence that Ups2–Mdm35, a protein complex localized at the mitochondrial intermembrane space, mediates PS transport for PE synthesis in respiration-active mitochondria. UPS2- and MDM35-null mutations greatly attenuated conversion of PS to PE in yeast cells growing logarithmically under nonfermentable conditions, but not fermentable conditions. A recombinant Ups2–Mdm35 fusion protein exhibited phospholipid-transfer activity between liposomes in vitro. Furthermore, UPS2 expression was elevated under nonfermentable conditions and at the diauxic shift, the metabolic transition from glycolysis to oxidative phosphorylation. These results demonstrate that Ups2–Mdm35 functions as a PS transfer protein and enhances mitochondrial PE synthesis in response to the cellular metabolic state.

Multiple pathways for mitochondrial protein traffic

2009 ◽  
Vol 390 (8) ◽  
Author(s):  
Toshiya Endo ◽  
Koji Yamano
Keyword(s):  
Outer Membrane ◽  
Protein Trafficking ◽  
Protein Import ◽  
Mitochondrial Protein ◽  
Intermembrane Space ◽  
Mitochondrial Protein Import ◽  
Inner Membrane ◽  
Protein Traffic

Abstract Mitochondria are two-membrane bounded organelles consisting of 1000–2000 different proteins, most of which are synthesized in the cytosol and subsequently imported into mitochondria. The imported proteins are further sorted to one of the four compartments, the outer membrane, intermembrane space, inner membrane, and matrix, mostly following one of the five major pathways. Mitochondrial protein import and sorting are mediated by the translocator complexes in the membranes and chaperones in the aqueous compartments operating along the import pathways. Here, we summarize the expanding knowledge on the roles of translocators, chaperones, and related components in the multiple pathways for mitochondrial protein trafficking.

scholarly journals Exposure of Saccharomyces cerevisiae to Acetaldehyde Induces Sulfur Amino Acid Metabolism and Polyamine Transporter Genes, Which Depend on Met4p and Haa1p Transcription Factors, Respectively

2004 ◽  
Vol 70 (4) ◽  
pp. 1913-1922 ◽  
Author(s):  
Agustín Aranda ◽  
Marcel-lí del Olmo
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Protein Biosynthesis ◽  
Sulfur Metabolism ◽  
Cell Cycle Control ◽  
Control Cell ◽  
Mitochondrial Protein ◽  
Vitamin B1 ◽  
Global Gene Expression ◽  
Growth Conditions ◽  
Yeast Cells

ABSTRACT Acetaldehyde is a toxic compound produced by Saccharomyces cerevisiae cells under several growth conditions. The adverse effects of this molecule are important, as significant amounts accumulate inside the cells. By means of global gene expression analyses, we have detected the effects of acetaldehyde addition in the expression of about 400 genes. Repressed genes include many genes involved in cell cycle control, cell polarity, and the mitochondrial protein biosynthesis machinery. Increased expression is displayed in many stress response genes, as well as other families of genes, such as those encoding vitamin B1 biosynthesis machinery and proteins for aryl alcohol metabolism. The induction of genes involved in sulfur metabolism is dependent on Met4p and other well-known factors involved in the transcription of MET genes under nonrepressing conditions of sulfur metabolism. Moreover, the deletion of MET4 leads to increased acetaldehyde sensitivity. TPO genes encoding polyamine transporters are also induced by acetaldehyde; in this case, the regulation is dependent on the Haa1p transcription factor. In this paper, we discuss the connections between acetaldehyde and the processes affected by this compound in yeast cells with reference to the microarray data.

scholarly journals Tim23–Tim50 pair coordinates functions of translocators and motor proteins in mitochondrial protein import

2009 ◽  
Vol 184 (1) ◽  
pp. 129-141 ◽  
Author(s):  
Yasushi Tamura ◽  
Yoshihiro Harada ◽  
Takuya Shiota ◽  
Koji Yamano ◽  
Kazuaki Watanabe ◽  
...  
Keyword(s):  
Motor Proteins ◽  
Protein Import ◽  
Protein Translocation ◽  
Mitochondrial Protein ◽  
Intermembrane Space ◽  
Mitochondrial Protein Import ◽  
Inner Membrane ◽  
The Matrix ◽  
General Protein ◽  
Mitochondrial Hsp70

Mitochondrial protein traffic requires coordinated operation of protein translocator complexes in the mitochondrial membrane. The TIM23 complex translocates and inserts proteins into the mitochondrial inner membrane. Here we analyze the intermembrane space (IMS) domains of Tim23 and Tim50, which are essential subunits of the TIM23 complex, in these functions. We find that interactions of Tim23 and Tim50 in the IMS facilitate transfer of precursor proteins from the TOM40 complex, a general protein translocator in the outer membrane, to the TIM23 complex. Tim23–Tim50 interactions also facilitate a late step of protein translocation across the inner membrane by promoting motor functions of mitochondrial Hsp70 in the matrix. Therefore, the Tim23–Tim50 pair coordinates the actions of the TOM40 and TIM23 complexes together with motor proteins for mitochondrial protein import.

scholarly journals Suppression of a Defect in Mitochondrial Protein Import Identifies Cytosolic Proteins Required for Viability of Yeast Cells Lacking Mitochondrial DNA

Genetics ◽  
2003 ◽  
Vol 165 (1) ◽  
pp. 35-45
Author(s):  
Cory D Dunn ◽  
Robert E Jensen
Keyword(s):  
Mitochondrial Dna ◽  
Protein Import ◽  
Mitochondrial Protein ◽  
Yeast Cells ◽  
Mitochondrial Protein Import ◽  
Inner Membrane ◽  
Cytosolic Proteins ◽  
Mitochondrial Inner Membrane ◽  
Genes Encoding ◽  
Negative Phenotype

Abstract The TIM22 complex, required for the insertion of imported polytopic proteins into the mitochondrial inner membrane, contains the nonessential Tim18p subunit. To learn more about the function of Tim18p, we screened for high-copy suppressors of the inability of tim18Δ mutants to live without mitochondrial DNA (mtDNA). We identified several genes encoding cytosolic proteins, including CCT6, SSB1, ICY1, TIP41, and PBP1, which, when overproduced, rescue the mtDNA dependence of tim18Δ cells. Furthermore, these same plasmids rescue the petite-negative phenotype of cells lacking other components of the mitochondrial protein import machinery. Strikingly, disruption of the genes identified by the different suppressors produces cells that are unable to grow without mtDNA. We speculate that loss of mtDNA leads to a lowered inner membrane potential, and subtle changes in import efficiency can no longer be tolerated. Our results suggest that increased amounts of Cct6p, Ssb1p, Icy1p, Tip41p, and Pbp1p help overcome the problems resulting from a defect in protein import.

Loss of the mitochondrial Hsp70 functions causes aggregation of mitochondria in yeast cells

2001 ◽  
Vol 114 (19) ◽  
pp. 3565-3574 ◽  
Author(s):  
Akemi Kawai ◽  
Shuh-ichi Nishikawa ◽  
Aiko Hirata ◽  
Toshiya Endo
Keyword(s):  
Protein Synthesis ◽  
Elevated Temperature ◽  
Protein Import ◽  
Mitochondrial Protein ◽  
Yeast Cells ◽  
Mitochondrial Morphology ◽  
Temperature Sensitive ◽  
Restrictive Temperature ◽  
Mitochondrial Matrix ◽  
Mitochondrial Hsp70

Ssc1p, a member of the Hsp70 family in the mitochondrial matrix of budding yeast, mediates protein import into mitochondria and prevents irreversible aggregation of proteins in the mitochondrial matrix during folding/assembly or at elevated temperature. Here, we show that functional inactivation of the mitochondrial Hsp70 system causes aggregation of mitochondria. When temperature-sensitive mitochondrial Hsp70 mutant cells were incubated at restrictive temperature, a tubular network of mitochondria was collapsed to form aggregates. Inhibition of protein synthesis in the cytosol did not suppress the mitochondrial aggregation and functional impairment of Tim23, a subunit of mitochondrial protein translocator in the inner membrane, did not cause mitochondrial aggregation. Therefore defects of the Hsp70 function in protein import into mitochondria or resulting accumulation of precursor forms of mitochondrial proteins outside the mitochondria are not the causal reason for the aberrant mitochondrial morphology. By contrast, deletion of Mdj1p, a functional partner for mitochondrial Hsp70 in prevention of irreversible protein aggregation in the matrix, but not in protein import into mitochondria, caused aggregation of mitochondria, which was enhanced at elevated temperature (37°C). The aggregation of mitochondria at 37°C was reversed when the temperature was lowered to 23°C unless protein synthesis was blocked. On the basis of these results, we propose that the mitochondrial matrix contains a protein that is responsible for the maintenance of mitochondrial morphology and requires mitochondrial Hsp70 for its function.

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 .

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