Mitochondrial thiol modification by a targeted electrophile inhibits metabolism in breast adenocarcinoma cells by inhibiting enzyme activity and protein levels
Many cancer cells follow an aberrant metabolic program to maintain energy for rapid cell proliferation. Metabolic reprogramming often involves the upregulation of glutaminolysis to generate reducing equivalents for the electron transport chain and amino acids for protein synthesis. Critical enzymes...
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doaj-28598c4d4dc14203b4dba36a2bf4aa842020-11-25T01:44:42ZengElsevierRedox Biology2213-23172016-08-018C13614810.1016/j.redox.2016.01.002Mitochondrial thiol modification by a targeted electrophile inhibits metabolism in breast adenocarcinoma cells by inhibiting enzyme activity and protein levelsM. Ryan Smith0Praveen K. Vayalil1Fen Zhou2Gloria A. Benavides3Reena R. Beggs4Hafez Golzarian5Bhavitavya Nijampatnam6Patsy G. Oliver7Robin A.J. Smith8Michael P. Murphy9Sadanandan E. Velu10Aimee Landar11Department of Pathology, University of Alabama at Birmingham, AL, USADepartment of Pathology, University of Alabama at Birmingham, AL, USADepartment of Pathology, University of Alabama at Birmingham, AL, USADepartment of Pathology, University of Alabama at Birmingham, AL, USADepartment of Pathology, University of Alabama at Birmingham, AL, USADepartment of Chemistry, University of Alabama at Birmingham, AL, USADepartment of Chemistry, University of Alabama at Birmingham, AL, USADepartment of Radiation Oncology, University of Alabama at Birmingham, AL, USADepartment of Chemistry, University of Otago, Dunedin, New ZealandMRC Mitochondrial Biology Unit, Cambridge, UKDepartment of Chemistry, University of Alabama at Birmingham, AL, USADepartment of Pathology, University of Alabama at Birmingham, AL, USAMany cancer cells follow an aberrant metabolic program to maintain energy for rapid cell proliferation. Metabolic reprogramming often involves the upregulation of glutaminolysis to generate reducing equivalents for the electron transport chain and amino acids for protein synthesis. Critical enzymes involved in metabolism possess a reactive thiolate group, which can be modified by certain oxidants. In the current study, we show that modification of mitochondrial protein thiols by a model compound, iodobutyl triphenylphosphonium (IBTP), decreased mitochondrial metabolism and ATP in MDA-MB 231 (MB231) breast adenocarcinoma cells up to 6 days after an initial 24 h treatment. Mitochondrial thiol modification also depressed oxygen consumption rates (OCR) in a dose-dependent manner to a greater extent than a non-thiol modifying analog, suggesting that thiol reactivity is an important factor in the inhibition of cancer cell metabolism. In non-tumorigenic MCF-10A cells, IBTP also decreased OCR; however the extracellular acidification rate was significantly increased at all but the highest concentration (10 µM) of IBTP indicating that thiol modification can have significantly different effects on bioenergetics in tumorigenic versus non-tumorigenic cells. ATP and other adenonucleotide levels were also decreased by thiol modification up to 6 days post-treatment, indicating a decreased overall energetic state in MB231 cells. Cellular proliferation of MB231 cells was also inhibited up to 6 days post-treatment with little change to cell viability. Targeted metabolomic analyses revealed that thiol modification caused depletion of both Krebs cycle and glutaminolysis intermediates. Further experiments revealed that the activity of the Krebs cycle enzyme, aconitase, was attenuated in response to thiol modification. Additionally, the inhibition of glutaminolysis corresponded to decreased glutaminase C (GAC) protein levels, although other protein levels were unaffected. This study demonstrates for the first time that mitochondrial thiol modification inhibits metabolism via inhibition of both aconitase and GAC in a breast cancer cell model.http://www.sciencedirect.com/science/article/pii/S2213231716300027BioenergeticsIBTPRedox signalingSeahorse extracellular flux analysisTricarboxylic acid cycleKrebs cycle |
collection |
DOAJ |
language |
English |
format |
Article |
sources |
DOAJ |
author |
M. Ryan Smith Praveen K. Vayalil Fen Zhou Gloria A. Benavides Reena R. Beggs Hafez Golzarian Bhavitavya Nijampatnam Patsy G. Oliver Robin A.J. Smith Michael P. Murphy Sadanandan E. Velu Aimee Landar |
spellingShingle |
M. Ryan Smith Praveen K. Vayalil Fen Zhou Gloria A. Benavides Reena R. Beggs Hafez Golzarian Bhavitavya Nijampatnam Patsy G. Oliver Robin A.J. Smith Michael P. Murphy Sadanandan E. Velu Aimee Landar Mitochondrial thiol modification by a targeted electrophile inhibits metabolism in breast adenocarcinoma cells by inhibiting enzyme activity and protein levels Redox Biology Bioenergetics IBTP Redox signaling Seahorse extracellular flux analysis Tricarboxylic acid cycle Krebs cycle |
author_facet |
M. Ryan Smith Praveen K. Vayalil Fen Zhou Gloria A. Benavides Reena R. Beggs Hafez Golzarian Bhavitavya Nijampatnam Patsy G. Oliver Robin A.J. Smith Michael P. Murphy Sadanandan E. Velu Aimee Landar |
author_sort |
M. Ryan Smith |
title |
Mitochondrial thiol modification by a targeted electrophile inhibits metabolism in breast adenocarcinoma cells by inhibiting enzyme activity and protein levels |
title_short |
Mitochondrial thiol modification by a targeted electrophile inhibits metabolism in breast adenocarcinoma cells by inhibiting enzyme activity and protein levels |
title_full |
Mitochondrial thiol modification by a targeted electrophile inhibits metabolism in breast adenocarcinoma cells by inhibiting enzyme activity and protein levels |
title_fullStr |
Mitochondrial thiol modification by a targeted electrophile inhibits metabolism in breast adenocarcinoma cells by inhibiting enzyme activity and protein levels |
title_full_unstemmed |
Mitochondrial thiol modification by a targeted electrophile inhibits metabolism in breast adenocarcinoma cells by inhibiting enzyme activity and protein levels |
title_sort |
mitochondrial thiol modification by a targeted electrophile inhibits metabolism in breast adenocarcinoma cells by inhibiting enzyme activity and protein levels |
publisher |
Elsevier |
series |
Redox Biology |
issn |
2213-2317 |
publishDate |
2016-08-01 |
description |
Many cancer cells follow an aberrant metabolic program to maintain energy for rapid cell proliferation. Metabolic reprogramming often involves the upregulation of glutaminolysis to generate reducing equivalents for the electron transport chain and amino acids for protein synthesis. Critical enzymes involved in metabolism possess a reactive thiolate group, which can be modified by certain oxidants. In the current study, we show that modification of mitochondrial protein thiols by a model compound, iodobutyl triphenylphosphonium (IBTP), decreased mitochondrial metabolism and ATP in MDA-MB 231 (MB231) breast adenocarcinoma cells up to 6 days after an initial 24 h treatment. Mitochondrial thiol modification also depressed oxygen consumption rates (OCR) in a dose-dependent manner to a greater extent than a non-thiol modifying analog, suggesting that thiol reactivity is an important factor in the inhibition of cancer cell metabolism. In non-tumorigenic MCF-10A cells, IBTP also decreased OCR; however the extracellular acidification rate was significantly increased at all but the highest concentration (10 µM) of IBTP indicating that thiol modification can have significantly different effects on bioenergetics in tumorigenic versus non-tumorigenic cells. ATP and other adenonucleotide levels were also decreased by thiol modification up to 6 days post-treatment, indicating a decreased overall energetic state in MB231 cells. Cellular proliferation of MB231 cells was also inhibited up to 6 days post-treatment with little change to cell viability. Targeted metabolomic analyses revealed that thiol modification caused depletion of both Krebs cycle and glutaminolysis intermediates. Further experiments revealed that the activity of the Krebs cycle enzyme, aconitase, was attenuated in response to thiol modification. Additionally, the inhibition of glutaminolysis corresponded to decreased glutaminase C (GAC) protein levels, although other protein levels were unaffected. This study demonstrates for the first time that mitochondrial thiol modification inhibits metabolism via inhibition of both aconitase and GAC in a breast cancer cell model. |
topic |
Bioenergetics IBTP Redox signaling Seahorse extracellular flux analysis Tricarboxylic acid cycle Krebs cycle |
url |
http://www.sciencedirect.com/science/article/pii/S2213231716300027 |
work_keys_str_mv |
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