scholarly journals Intramitochondrial sorting of the precursor to yeast cytochrome c oxidase subunit Va.

1993 ◽  
Vol 121 (5) ◽  
pp. 1021-1029 ◽  
Author(s):  
B R Miller ◽  
M G Cumsky
Keyword(s):  
Cytochrome C ◽  
Cytochrome C Oxidase ◽  
Mitochondrial Protein ◽  
Inner Membrane ◽  
Oxidase Subunit ◽  
Mitochondrial Inner Membrane ◽  
The Matrix ◽  
Inner Membrane Protein ◽  
Membrane Spanning ◽  
Mitochondrial Heat Shock Protein

We have continued our studies on the import pathway of the precursor to yeast cytochrome c oxidase subunit Va (pVa), a mitochondrial inner membrane protein. Previous work on this precursor demonstrated that import of pVa is unusually efficient, and that inner membrane localization is directed by a membrane-spanning domain in the COOH-terminal third of the protein. Here we report the results of studies aimed at analyzing the intramitochondrial sorting of pVa, as well as the role played by ancillary factors in import and localization of the precursor. We found that pVa was efficiently imported and correctly sorted in mitochondria prepared from yeast strains defective in the function of either mitochondrial heat shock protein (hsp)60 or hsp70. Under identical conditions the import and sorting of another mitochondrial protein, the precursor to the beta subunit of the F1 ATPase, was completely defective. Consistent with previous results demonstrating that the subunit Va precursor is loosely folded, we found that pVa could be efficiently imported into mitochondria after translation in wheat germ extracts. This results suggests that normal levels of extramitochondrial hsp70 are also not required for import of the protein. The results of this study enhance our understanding of the mechanism by which pVa is routed to the mitochondrial inner membrane. They suggest that while the NH2 terminus of pVa is exposed to the matrix and processed by the matrix metalloprotease, the protein remains anchored to the inner membrane before being assembled into a functional holoenzyme complex.

scholarly journals SMS1, a high-copy suppressor of the yeast mas6 mutant, encodes an essential inner membrane protein required for mitochondrial protein import.

1994 ◽  
Vol 5 (5) ◽  
pp. 529-538 ◽  
Author(s):  
K R Ryan ◽  
M M Menold ◽  
S Garrett ◽  
R E Jensen
Keyword(s):  
Membrane Protein ◽  
Protein Import ◽  
Mitochondrial Protein ◽  
Mitochondrial Protein Import ◽  
Yeast Saccharomyces Cerevisiae ◽  
Inner Membrane ◽  
Mitochondrial Inner Membrane ◽  
Inner Membrane Protein ◽  
Membrane Spanning ◽  
Membrane Spanning Domains

MAS6 encodes an essential inner membrane protein required for mitochondrial protein import in the yeast Saccharomyces cerevisiae (Emtage and Jensen, 1993). To identify new inner membrane import components, we isolated a high-copy suppressor (SMS1) of the mas6-1 mutant. SMS1 encodes a 16.5-kDa protein that contains several potential membrane-spanning domains. The Sms1 protein is homologous to the carboxyl-terminal domain of the Mas6 protein. Like Mas6p, Sms1p is located in the mitochondrial inner membrane and is an essential protein. Depletion of Sms1p from cells causes defects in the import of several mitochondrial precursor proteins, suggesting that Sms1p is a new inner membrane import component. Our observations raise the possibility that Sms1p and Mas6p act together to translocate proteins across the inner membrane.

scholarly journals Interactions amongCOX1,COX2, andCOX3mRNA-specific Translational Activator Proteins on the Inner Surface of the Mitochondrial Inner Membrane ofSaccharomyces cerevisiae

2003 ◽  
Vol 14 (1) ◽  
pp. 324-333 ◽  
Author(s):  
Sushma Naithani ◽  
Scott A. Saracco ◽  
Christine A. Butler ◽  
Thomas D. Fox
Keyword(s):  
Cytochrome C ◽  
Cytochrome C Oxidase ◽  
Translation System ◽  
Mitochondrial Translation ◽  
Inner Membrane ◽  
Activator Proteins ◽  
The Core ◽  
Mitochondrial Inner Membrane ◽  
The Matrix ◽  
Translational Activator

The core of the cytochrome c oxidase complex is composed of its three largest subunits, Cox1p, Cox2p, and Cox3p, which are encoded in mitochondrial DNA of Saccharomyces cerevisiae and inserted into the inner membrane from the inside. Mitochondrial translation of the COX1,COX2, and COX3 mRNAs is activated mRNA specifically by the nuclearly coded proteins Pet309p, Pet111p, and the concerted action of Pet54p, Pet122p, and Pet494p, respectively. Because the translational activators recognize sites in the 5′-untranslated leaders of these mRNAs and because untranslated mRNA sequences contain information for targeting their protein products, the activators are likely to play a role in localizing translation. Herein, we report physical associations among the mRNA-specific translational activator proteins, located on the matrix side of the inner membrane. These interactions, detected by coimmune precipitation and by two-hybrid experiments, suggest that the translational activator proteins could be organized on the surface of the inner membrane such that synthesis of Cox1p, Cox2p, and Cox3p would be colocalized in a way that facilitates assembly of the core of the cytochrome c oxidase complex. In addition, we found interactions between Nam1p/Mtf2p and the translational activators, suggesting an organized delivery of mitochondrial mRNAs to the translation system.

scholarly journals Mba1, a Novel Component of the Mitochondrial Protein Export Machinery of the Yeast Saccharomyces cerevisiae

2001 ◽  
Vol 153 (5) ◽  
pp. 1085-1096 ◽  
Author(s):  
Marc Preuss ◽  
Klaus Leonhard ◽  
Kai Hell ◽  
Rosemary A. Stuart ◽  
Walter Neupert ◽  
...  
Keyword(s):  
Mitochondrial Protein ◽  
Protein Export ◽  
Mitochondrial Translation ◽  
Yeast Saccharomyces Cerevisiae ◽  
Inner Membrane ◽  
Protein Insertion ◽  
Mitochondrial Inner Membrane ◽  
The Matrix ◽  
Biogenesis Of Mitochondria ◽  
Inner Membrane Protein

The biogenesis of mitochondria requires the integration of many proteins into the inner membrane from the matrix side. The inner membrane protein Oxa1 plays an important role in this process. We identified Mba1 as a second mitochondrial component that is required for efficient protein insertion. Like Oxa1, Mba1 specifically interacts both with mitochondrial translation products and with conservatively sorted, nuclear-encoded proteins during their integration into the inner membrane. Oxa1 and Mba1 overlap in function and substrate specificity, but both can act independently of each other. We conclude that Mba1 is part of the mitochondrial protein export machinery and represents the first component of a novel Oxa1-independent insertion pathway into the mitochondrial inner membrane.

scholarly journals Characterization of Mmp37p, aSaccharomyces cerevisiaeMitochondrial Matrix Protein with a Role in Mitochondrial Protein Import

2006 ◽  
Vol 17 (9) ◽  
pp. 4051-4062 ◽  
Author(s):  
Michelle R. Gallas ◽  
Mary K. Dienhart ◽  
Rosemary A. Stuart ◽  
Roy M. Long
Keyword(s):  
Matrix Protein ◽  
Protein Import ◽  
Mitochondrial Protein ◽  
Temperature Sensitive ◽  
Inner Membrane ◽  
Mitochondrial Inner Membrane ◽  
Close Proximity ◽  
The Matrix ◽  
Targeting Signal

Many mitochondrial proteins are encoded by nuclear genes and after translation in the cytoplasm are imported via translocases in the outer and inner membranes, the TOM and TIM complexes, respectively. Here, we report the characterization of the mitochondrial protein, Mmp37p (YGR046w) and demonstrate its involvement in the process of protein import into mitochondria. Haploid cells deleted of MMP37 are viable but display a temperature-sensitive growth phenotype and are inviable in the absence of mitochondrial DNA. Mmp37p is located in the mitochondrial matrix where it is peripherally associated with the inner membrane. We show that Mmp37p has a role in the translocation of proteins across the mitochondrial inner membrane via the TIM23-PAM complex and further demonstrate that substrates containing a tightly folded domain in close proximity to their mitochondrial targeting sequences display a particular dependency on Mmp37p for mitochondrial import. Prior unfolding of the preprotein, or extension of the region between the targeting signal and the tightly folded domain, relieves their dependency for Mmp37p. Furthermore, evidence is presented to show that Mmp37 may affect the assembly state of the TIM23 complex. On the basis of these findings, we hypothesize that the presence of Mmp37p enhances the early stages of the TIM23 matrix import pathway to ensure engagement of incoming preproteins with the mtHsp70p/PAM complex, a step that is necessary to drive the unfolding and complete translocation of the preprotein into the matrix.

Removal of a hydrophobic domain within the mature portion of a mitochondrial inner membrane protein causes its mislocalization to the matrix

1990 ◽  
Vol 10 (5) ◽  
pp. 1873-1881
Author(s):  
S M Glaser ◽  
B R Miller ◽  
M G Cumsky
Keyword(s):  
Amino Acids ◽  
Mitochondrial Matrix ◽  
Mature Protein ◽  
Wild Type ◽  
Inner Membrane ◽  
Hydrophobic Domain ◽  
Mitochondrial Inner Membrane ◽  
The Matrix ◽  
Inner Membrane Protein ◽  
Cognate Protein

We have examined the import and intramitochondrial localization of the precursor to yeast cytochrome c oxidase subunit Va, a protein of the mitochondrial inner membrane. The results of studies on the import of subunit Va derivatives carrying altered presequences suggest that the uptake of this protein is highly efficient. We found that a presequence of only 5 amino acids (Met-Leu-Ser-Leu-Arg) could direct the import and localization of subunit Va with wild-type efficiency, as judged by several different assays. We also found that subunit Va could be effectively targeted to the mitochondrial inner membrane with a heterologous presequence that failed to direct import of its cognate protein. The results presented here confirmed those of an earlier study and showed clearly that the information required to "sort" subunit Va to the inner membrane resides in the mature protein sequence, not within the presequence per se. We present additional evidence that the aforementioned sorting information is contained, at least in part, in a hydrophobic stretch of 22 amino acids residing within the C-terminal third of the protein. Removal of this domain caused subunit Va to be mislocalized to the mitochondrial matrix.

scholarly journals Removal of a hydrophobic domain within the mature portion of a mitochondrial inner membrane protein causes its mislocalization to the matrix.

1990 ◽  
Vol 10 (5) ◽  
pp. 1873-1881 ◽  
Author(s):  
S M Glaser ◽  
B R Miller ◽  
M G Cumsky
Keyword(s):  
Amino Acids ◽  
Mitochondrial Matrix ◽  
Mature Protein ◽  
Wild Type ◽  
Inner Membrane ◽  
Hydrophobic Domain ◽  
Mitochondrial Inner Membrane ◽  
The Matrix ◽  
Inner Membrane Protein ◽  
Cognate Protein

We have examined the import and intramitochondrial localization of the precursor to yeast cytochrome c oxidase subunit Va, a protein of the mitochondrial inner membrane. The results of studies on the import of subunit Va derivatives carrying altered presequences suggest that the uptake of this protein is highly efficient. We found that a presequence of only 5 amino acids (Met-Leu-Ser-Leu-Arg) could direct the import and localization of subunit Va with wild-type efficiency, as judged by several different assays. We also found that subunit Va could be effectively targeted to the mitochondrial inner membrane with a heterologous presequence that failed to direct import of its cognate protein. The results presented here confirmed those of an earlier study and showed clearly that the information required to "sort" subunit Va to the inner membrane resides in the mature protein sequence, not within the presequence per se. We present additional evidence that the aforementioned sorting information is contained, at least in part, in a hydrophobic stretch of 22 amino acids residing within the C-terminal third of the protein. Removal of this domain caused subunit Va to be mislocalized to the mitochondrial matrix.

scholarly journals An unusual mitochondrial import pathway for the precursor to yeast cytochrome c oxidase subunit Va.

1991 ◽  
Vol 112 (5) ◽  
pp. 833-841 ◽  
Author(s):  
B R Miller ◽  
M G Cumsky
Keyword(s):  
Cytochrome C ◽  
Cytochrome C Oxidase ◽  
Proteinase K ◽  
Oxidase Subunit ◽  
Mitochondrial Inner Membrane ◽  
Apocytochrome C ◽  
Precursor Proteins ◽  
Surface Receptor ◽  
High Concentrations ◽  
Hydrolysis Of

We have studied the import of the precursor to yeast cytochrome c oxidase subunit Va, a protein of the mitochondrial inner membrane. Like the majority of mitochondrial precursor proteins studied thus far, import of presubunit Va was dependent upon both a membrane potential (delta psi) and the hydrolysis of ATP. However, the levels of ATP necessary for the import of presubunit Va were significantly lower than those required for the import of a different mitochondrial precursor protein, the beta subunit of the F1-ATPase. The rate of import of presubunit Va was found to be unaffected by temperature over the range 0 to 30 degrees C, and was not facilitated by prior denaturation of the protein. These results, in conjunction with those of an earlier study demonstrating that presubunit Va could be efficiently targeted to mitochondria with minimal presequences, suggest that the subunit Va precursor normally exists in a loosely folded conformation. Presubunit Va could also be imported into mitochondria that had been pretreated with high concentrations of trypsin or proteinase K (1 mg/ml and 200 micrograms/ml, respectively). Furthermore, the rate of import into trypsin-treated mitochondria, at both 0 and 30 degrees C, was identical to that observed with the untreated organelles. Thus, import of presubunit Va is not dependent upon the function of a protease-sensitive surface receptor. When taken together, the results of this study suggest that presubunit Va follows an unusual import pathway. While this pathway uses several well-established translocation steps, in its entirety it is distinct from either the receptor-independent pathway used by apocytochrome c, or the more general pathway used by a majority of mitochondrial precursor proteins.

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