title: Mitochondrial calcium uniporter activity is dispensable for MDA-MB-231 breast carcinoma cell survival.
abstract: Calcium uptake through the mitochondrial Ca2+ uniporter (MCU) is thought to be essential in regulating cellular signaling events, energy status, and survival. Functional dissection of the uniporter is now possible through the recent identification of the genes encoding for MCU protein complex subunits. Cancer cells exhibit many aspects of mitochondrial dysfunction associated with altered mitochondrial Ca2+ levels including resistance to apoptosis, increased reactive oxygen species production and decreased oxidative metabolism. We used a publically available database to determine that breast cancer patient outcomes negatively correlated with increased MCU Ca2+ conducting pore subunit expression and decreased MICU1 regulatory subunit expression. We hypothesized breast cancer cells may therefore be sensitive to MCU channel manipulation. We used the widely studied MDA-MB-231 breast cancer cell line to investigate whether disruption or increased activation of mitochondrial Ca2+ uptake with specific siRNAs and adenoviral overexpression constructs would sensitize these cells to therapy-related stress. MDA-MB-231 cells were found to contain functional MCU channels that readily respond to cellular stimulation and elicit robust AMPK phosphorylation responses to nutrient withdrawal. Surprisingly, knockdown of MCU or MICU1 did not affect reactive oxygen species production or cause significant effects on clonogenic cell survival of MDA-MB-231 cells exposed to irradiation, chemotherapeutic agents, or nutrient deprivation. Overexpression of wild type or a dominant negative mutant MCU did not affect basal cloning efficiency or ceramide-induced cell killing. In contrast, non-cancerous breast epithelial HMEC cells showed reduced survival after MCU or MICU1 knockdown. These results support the conclusion that MDA-MB-231 breast cancer cells do not rely on MCU or MICU1 activity for survival in contrast to previous findings in cells derived from cervical, colon, and prostate cancers and suggest that not all carcinomas will be sensitive to therapies targeting mitochondrial Ca2+ uptake mechanisms.
authors.affiliation: Department of Internal Medicine, Division of Cardiovascular Medicine, Roy J. and Lucille A. Carver College of Medicine, University of Iowa, Iowa City, Iowa, United States of America.
authors.forename: Duane D
authors.initials: DD
authors.lastname: Hall
authors.affiliation: Department of Internal Medicine, Division of Cardiovascular Medicine, Roy J. and Lucille A. Carver College of Medicine, University of Iowa, Iowa City, Iowa, United States of America.
authors.forename: Yuejin
authors.initials: Y
authors.lastname: Wu
authors.affiliation: Free Radical and Radiation Biology Program, Department of Radiation Oncology, Holden Comprehensive Cancer Center, Roy J. and Lucille A. Carver College of Medicine, University of Iowa, Iowa City, Iowa, United States of America.
authors.forename: Frederick E
authors.initials: FE
authors.lastname: Domann
authors.affiliation: Free Radical and Radiation Biology Program, Department of Radiation Oncology, Holden Comprehensive Cancer Center, Roy J. and Lucille A. Carver College of Medicine, University of Iowa, Iowa City, Iowa, United States of America.
authors.forename: Douglas R
authors.initials: DR
authors.lastname: Spitz
authors.affiliation: Department of Internal Medicine, Division of Cardiovascular Medicine, Roy J. and Lucille A. Carver College of Medicine, University of Iowa, Iowa City, Iowa, United States of America; Free Radical and Radiation Biology Program, Department of Radiation Oncology, Holden Comprehensive Cancer Center, Roy J. and Lucille A. Carver College of Medicine, University of Iowa, Iowa City, Iowa, United States of America; Department of Molecular Physiology and Biophysics, Roy J. and Lucille A. Carver College of Medicine, University of Iowa, Iowa City, Iowa, United States of America.
authors.forename: Mark E
authors.initials: ME
authors.lastname: Anderson
mesh_terms: D055372:AMP-Activated Protein Kinases / Q000378:metabolism; D000465:Algorithms; D000970:Antineoplastic Agents / Q000494:pharmacology; D001943:Breast Neoplasms / Q000378:metabolism / Q000473:pathology; D002118:Calcium / Q000378:metabolism*; D015220:Calcium Channels / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D002135:Calcium-Binding Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D027682:Cation Transport Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects / Q000528:radiation effects; D005260:Female; D006367:HeLa Cells; D006801:Humans; D008928:Mitochondria / Q000378:metabolism*; D033681:Mitochondrial Membrane Transport Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000378:metabolism; D011839:Radiation, Ionizing; D017382:Reactive Oxygen Species / Q000378:metabolism
chemical_list: D000970:Antineoplastic Agents; D015220:Calcium Channels; D002135:Calcium-Binding Proteins; D027682:Cation Transport Proteins; C116241:MICU1 protein, human; D033681:Mitochondrial Membrane Transport Proteins; D034741:RNA, Small Interfering; D017382:Reactive Oxygen Species; C034703:mitochondrial calcium uniporter; D055372:AMP-Activated Protein Kinases; D002118:Calcium