scholarly journals Inhibition of the Mitochondrial Protein Import Machinery Is Selectively Cytotoxic to Acute Myeloid Leukemia (AML) Cells and Stem Cells

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. 1572-1572 ◽  
Author(s):  
Danny V Jeyaraju ◽  
Veronique Voisin ◽  
Changjiang Xu ◽  
Rose Hurren ◽  
Marcela Gronda ◽  
...  
Keyword(s):  
Stem Cells ◽  
Protein Import ◽  
Mitochondrial Protein ◽  
Mitochondrial Proteins ◽  
Leukemia Cells ◽  
Mitochondrial Protein Import ◽  
Normal Cells ◽  
Mitochondrial Stress ◽  
Inner Membrane ◽  
The Impact

Abstract A subset of AML and stem cells have increased mitochondrial stress and increased expression of mitochondrial proteases that degrade misfolded mitochondrial proteins. Given the recent findings of the interplay between mitochondrial homeostasis and mitochondrial protein import, we hypothesized that AML cells have an increased reliance on mitochondrial protein import as a compensatory mechanism for increased mitochondrial stress. To test this hypothesis, we measured expression levels of key mitochondrial protein import genes in publicly available datasets (GSE30377, GSE42414 and GSE 24759) and demonstrated their up regulation in a subset of AML cells over normal hematopoietic cells. Increased expression occurred across the spectrum of molecular mutations and cytogenetic abnormalities. Moreover, expression levels of mitochondrial protein import genes were enriched in functionally defined AML stem cells over bulk cells. To assess the impact of inhibiting mitochondrial protein import in AML, we knocked down the outer mitochondrial membrane import channel TOM40, the inner membrane import channel Tim23 and the oxidase ALR, that folds proteins through a disulfide relay system in the mitochondrial inner membrane space and is a rate limiting step for the import of a subset of mitochondrial proteins. Knockdown of these targets in OCI-AML2, TEX and U937 leukemia cells with shRNA reduced growth and viability of AML cells. Knockdown of ALR targeted the leukemia initiating cells as it abrogated engraftment of TEX leukemia cells into immune deficient mice (shRNA ALR = 5.481 +/- 0.9 % engraftment vs shRNA control= 29.44 +/- 5.4 % engraftment; p =0.0004) . Mechanistically, knockdown of mitochondrial import genes reduced levels of nuclear (ATP5A, SDHB and NDUFB8), but not mitochondrial (CoxII) encoded proteins of the OXPHOS chain. This in turn led to decreased basal oxygen consumption in leukemic cells. As a chemical approach to investigate the impact of inhibiting mitochondrial protein import in AML and normal cells, we tested the effects of MitoBloCK-6 and related analogues that selectively inhibit ALR in zebrafish, hESCs and yeast models. MitoBloCK-6 and related analogues killed leukemia cell lines (OCI-AML2, TEX, Jurkat and NB4) with an IC50of 5-10 µM. At these concentrations, MitoBloCK-6 decreased levels of nuclear (ATP5A, SDHB and NDUFB8), but not mitochondrial encoded (CoxII) proteins of the OXPHOS chain. Demonstrating the functional importance of changes in mitochondrial metabolism by these compounds, rho zero 143B rhabdomyosarcoma cells that lack mitochondrial DNA and rely solely on glycolysis were resistant to MitoBloCK-6. Finally, we tested the effects of MitoBloCK-6 on primary AML and normal cells. Treatment with MitoBloCK-6 (2 µM) inhibited clonogenic growth of (4 of 5) primary AML > 64% but produced <3% loss of clonogenic growth in normal hematopoietic cells. Finally, in an OCI-AML2 xenograft model, systemic administration of MitoBloCK-6 reduced tumor growth > 50% of control without toxicity. Thus, we have demonstrated that AML cells have a unique reliance on mitochondrial protein import and inhibition of this pathway may be a new therapeutic strategy to selectively target a subset of AML and AML stem cells. Disclosures Schimmer: Novartis: Honoraria.

scholarly journals Cooperation of translocase complexes in mitochondrial protein import

2007 ◽  
Vol 179 (4) ◽  
pp. 585-591 ◽  
Author(s):  
Stephan Kutik ◽  
Bernard Guiard ◽  
Helmut E. Meyer ◽  
Nils Wiedemann ◽  
Nikolaus Pfanner
Keyword(s):  
Outer Membrane ◽  
Protein Import ◽  
Protein Complexes ◽  
Mitochondrial Protein ◽  
Mitochondrial Proteins ◽  
Intermembrane Space ◽  
Mitochondrial Protein Import ◽  
Inner Membrane ◽  
Precursor Proteins ◽  
Preprotein Translocation

Most mitochondrial proteins are synthesized in the cytosol and imported into one of the four mitochondrial compartments: outer membrane, intermembrane space, inner membrane, and matrix. Each compartment contains protein complexes that interact with precursor proteins and promote their transport. These translocase complexes do not act as independent units but cooperate with each other and further membrane complexes in a dynamic manner. We propose that a regulated coupling of translocases is important for the coordination of preprotein translocation and efficient sorting to intramitochondrial compartments.

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.

Plant mitochondrial protein import: mechanisms and control

1999 ◽  
Vol 26 (8) ◽  
pp. 725 ◽  
Author(s):  
James Whelan
Keyword(s):  
Protein Import ◽  
Current Knowledge ◽  
Mitochondrial Protein ◽  
Mitochondrial Proteins ◽  
Mitochondrial Protein Import ◽  
Future Directions ◽  
Receptor Structure ◽  
Processing Peptidase ◽  
Targeting Signals ◽  
And Control

The characterisation of components of the plant mitochondrial import apparatus along with the availability of over one hundred nuclear-encoded mitochondrial proteins allows the study of plant mitochondrial protein import in homologous systems. From these studies it has emerged that although similarities in the import process exist with other organisms, significance differences exist, such as receptor structure, location of processing peptidase and targeting signals. These differences mean that previous studies carried out in heterologous systems must be re-evaluated. Further studies into protein import in plants need to be directed at understanding the mechanism of import and how this process may be controlled. In this review the latter points will be dealt with in terms of summarising our current knowledge and possible future directions.

scholarly journals MAS6 encodes an essential inner membrane component of the yeast mitochondrial protein import pathway

1993 ◽  
Vol 122 (5) ◽  
pp. 1003-1012 ◽  
Author(s):  
JL Emtage ◽  
RE Jensen
Keyword(s):  
Protein Import ◽  
Early Stage ◽  
Osmotic Shock ◽  
Alkali Treatment ◽  
Mitochondrial Protein ◽  
Temperature Sensitive ◽  
Restrictive Temperature ◽  
Mitochondrial Protein Import ◽  
Inner Membrane ◽  
Mitochondrial Inner Membrane

To identify new components that mediate mitochondrial protein import, we analyzed mas6, an import mutant in the yeast Saccharomyces cerevisiae. mas6 mutants are temperature sensitive for viability, and accumulate mitochondrial precursor proteins at the restrictive temperature. We show that mas6 does not correspond to any of the presently identified import mutants, and we find that mitochondria isolated from mas6 mutants are defective at an early stage of the mitochondrial protein import pathway. MAS6 encodes a 23-kD protein that contains several potential membrane spanning domains, and yeast strains disrupted for MAS6 are inviable at all temperatures and on all carbon sources. The Mas6 protein is located in the mitochondrial inner membrane and cannot be extracted from the membrane by alkali treatment. Antibodies to the Mas6 protein inhibit import into isolated mitochondria, but only when the outer membrane has been disrupted by osmotic shock. Mas6p therefore represents an essential import component located in the mitochondrial inner membrane.

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.

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 Mitochondrial protein import: An unexpected disulfide bond

2016 ◽  
Vol 214 (4) ◽  
pp. 363-365 ◽  
Author(s):  
Dejana Mokranjac
Keyword(s):  
Disulfide Bond ◽  
Protein Import ◽  
Protein Translocation ◽  
Mitochondrial Protein ◽  
Channel Gating ◽  
Mitochondrial Proteins ◽  
Mitochondrial Protein Import ◽  
Cell Biol ◽  
Molecular Nature

Most mitochondrial proteins are imported through the TIM23 translocation channel, the structure and molecular nature of which are still unclear. In this issue, Ramesh et al. (2016. J. Cell Biol. http://dx.doi.org/10.1083/jcb.201602074) show that the TIM23 subunit Tim17 contains a disulfide bond that is crucial for protein translocation and channel gating.

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 From cytosol to mitochondria: the beginning of a protein journey

2020 ◽  
Vol 401 (6-7) ◽  
pp. 645-661 ◽  
Author(s):  
Maria Clara Avendaño-Monsalve ◽  
José Carlos Ponce-Rojas ◽  
Soledad Funes
Keyword(s):  
Mitochondrial Biogenesis ◽  
Stress Responses ◽  
Protein Import ◽  
Protein Targeting ◽  
Mitochondrial Protein ◽  
Past Research ◽  
Membrane Receptors ◽  
Mitochondrial Proteins ◽  
Mitochondrial Protein Import ◽  
The Past

AbstractMitochondrial protein import is one of the key processes during mitochondrial biogenesis that involves a series of events necessary for recognition and delivery of nucleus-encoded/cytosol-synthesized mitochondrial proteins into the organelle. The past research efforts have mainly unraveled how membrane translocases ensure the correct protein sorting within the different mitochondrial subcompartments. However, early steps of recognition and delivery remain relatively uncharacterized. In this review, we discuss our current understanding about the signals on mitochondrial proteins, as well as in the mRNAs encoding them, which with the help of cytosolic chaperones and membrane receptors support protein targeting to the organelle in order to avoid improper localization. In addition, we discuss recent findings that illustrate how mistargeting of mitochondrial proteins triggers stress responses, aiming to restore cellular homeostasis.

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