title: Steroidogenesis in MA-10 mouse Leydig cells is altered via fatty acid import into the mitochondria.
abstract: Mitochondria are home to many cellular processes, including oxidative phosphorylation and fatty acid metabolism, and in steroid-synthesizing cells, they are involved in cholesterol import and metabolism, which is the initiating step in steroidogenesis. The formation of macromolecular protein complexes aids in the regulation and efficiency of these mitochondrial functions, though because of their dynamic nature, they are hard to identify. To overcome this problem, we used Blue-Native PAGE with whole-gel mass spectrometry on isolated mitochondria from control and hormone-treated MA-10 mouse tumor Leydig cells. The presence of multiple mitochondrial protein complexes was shown. Although these were qualitatively similar under control and human chorionic gonadotropin (hCG)-stimulated conditions, quantitative differences in the components of the complexes emerged after hCG treatment. A prominent decrease was observed with proteins involved in fatty acid import into the mitochondria, implying that mitochondrial beta-oxidation is not essential for steroidogenesis. To confirm this observation, we inhibited fatty acid import utilizing the CPT1a inhibitor etomoxir, resulting in increased steroid production. Conversely, stimulation of mitochondrial beta-oxidation with metformin resulted in a dose-dependent reduction in steroidogenesis. These changes were accompanied by changes in mitochondrial respiration and in the lactic acid formed during glycolysis. Taken together, these results suggest that upon hormonal stimulation, mitochondria efficiently import cholesterol for steroid production at the expense of other lipids necessary for energy production, specifically fatty acids required for beta-oxidation.
authors.affiliation: The Research Institute of the McGill University Health Centre and Department of Medicine, McGill University, Montreal, Quebec, Canada.
authors.forename: Malena B
authors.initials: MB
authors.lastname: Rone
authors.affiliation: The Research Institute of the McGill University Health Centre and Department of Medicine, McGill University, Montreal, Quebec, Canada.
authors.forename: Andrew S
authors.initials: AS
authors.lastname: Midzak
authors.affiliation: The Research Institute of the McGill University Health Centre and Department of Medicine, McGill University, Montreal, Quebec, Canada.
authors.forename: Daniel B
authors.initials: DB
authors.lastname: Martinez-Arguelles
authors.affiliation: The Research Institute of the McGill University Health Centre and Department of Medicine, McGill University, Montreal, Quebec, Canada.
authors.forename: Jinjiang
authors.initials: J
authors.lastname: Fan
authors.affiliation: Protein Characterization Laboratory, Cancer Research Technology Program, Leidos Biomedical Research, Inc., Frederick National Laboratory for Cancer Research, NCI, Frederick, Maryland.
authors.forename: Xiaoying
authors.initials: X
authors.lastname: Ye
authors.affiliation: Protein Characterization Laboratory, Cancer Research Technology Program, Leidos Biomedical Research, Inc., Frederick National Laboratory for Cancer Research, NCI, Frederick, Maryland.
authors.forename: Josip
authors.initials: J
authors.lastname: Blonder
authors.affiliation: The Research Institute of the McGill University Health Centre and Department of Medicine, McGill University, Montreal, Quebec, Canada Departments of Biochemistry and Pharmacology & Therapeutics, McGill University, Montreal, Quebec, Canada vassilios.papadopoulos@mcgill.ca.
authors.forename: Vassilios
authors.initials: V
authors.lastname: Papadopoulos
mesh_terms: D000818:Animals; D001692:Biological Transport; D002460:Cell Line; D005227:Fatty Acids / Q000378:metabolism*; D005786:Gene Expression Regulation / Q000187:drug effects / Q000502:physiology; D007004:Hypoglycemic Agents / Q000494:pharmacology; D007985:Leydig Cells / Q000378:metabolism*; D008297:Male; D008687:Metformin / Q000494:pharmacology; D051379:Mice; D008928:Mitochondria / Q000378:metabolism*; D010084:Oxidation-Reduction; D013256:Steroids / Q000096:biosynthesis*; D059467:Transcriptome
chemical_list: D005227:Fatty Acids; D007004:Hypoglycemic Agents; D013256:Steroids; D008687:Metformin
keywords: Respiratory Bioflux Analysis; beta-oxidation; etomoxir; metformin; proteomics; steroid biosynthesis; translocator protein