title: 1,25-Dihydroxyvitamin D inhibits glutamine metabolism in Harvey-ras transformed MCF10A human breast epithelial cell.
abstract: Breast cancer is the second most common cancer among women in the US. The active form of vitamin D, 1,25-dihydroxyvitamin D (1,25(OH)2D), is proposed to inhibit cellular processes and to prevent breast cancer. The current studies investigated the effect of 1,25(OH)2D on glutamine metabolism during cancer progression employing Harvey-ras oncogene transformed MCF10A human breast epithelial cells (MCF10A-ras). Treatment with 1,25(OH)2D significantly reduced intracellular glutamine and glutamate levels measured by nuclear magnetic resonance (NMR) by 23±2% each. Further, 1,25(OH)2D treatment reduced glutamine and glutamate flux, determined by [U-(13)C5] glutamine tracer kinetics, into the TCA cycle by 31±0.2% and 17±0.4%, respectively. The relative levels of mRNA and protein abundance of the major glutamine transporter, solute linked carrier family 1 member A5 (SLC1A5), was significantly decreased by 1,25(OH)2D treatment in both MCF10A-ras cells and MCF10A which overexpress ErbB2 (HER-2/neu). Consistent with these results, glutamine uptake was reduced by 1,25(OH)2D treatment and the impact was eliminated with the SLC1A5 inhibitor L-γ-Glutamyl-p-nitroanilide (GPNA). A consensus sequence to the vitamin D responsive element (VDRE) was identified in silico in the SLC1A5 gene promoter, and site-directed mutagenesis analyses with reporter gene studies demonstrate a functional negative VDRE in the promoter of the SLC1A5 gene. siRNA-SLC1A5 transfection in MCF10A-ras cells significantly reduced SLC1A5 mRNA expression as well as decreased viable cell number similar to 1,25(OH)2D treatment. SLC1A5 knockdown also induced an increase in apoptotic cells in MCF10A-ras cells. These results suggest 1,25(OH)2D alters glutamine metabolism in MCF10A-ras cells by inhibiting glutamine uptake and utilization, in part through down-regulation of SLC1A5 transcript abundance. Thus, 1,25(OH)2D down-regulation of the glutamine transporter, SLC1A5, may facilitate vitamin D prevention of breast cancer.
authors.affiliation: Interdepartmental Nutrition Program, Purdue University, West Lafayette, IN 47906, United States. Electronic address: zhou249@purdue.edu.
authors.forename: Xuanzhu
authors.initials: X
authors.lastname: Zhou
authors.affiliation: Interdepartmental Nutrition Program, Purdue University, West Lafayette, IN 47906, United States. Electronic address: wzheng87@stanford.edu.
authors.forename: Wei
authors.initials: W
authors.lastname: Zheng
authors.affiliation: Department of Chemistry, Purdue University, West Lafayette, IN 47906, United States. Electronic address: ngowda@uw.edu.
authors.forename: G A
authors.initials: GA
authors.lastname: Nagana Gowda
authors.affiliation: Department of Chemistry, Purdue University, West Lafayette, IN 47906, United States. Electronic address: draftery@uw.edu.
authors.forename: Daniel
authors.initials: D
authors.lastname: Raftery
authors.affiliation: Interdepartmental Nutrition Program, Purdue University, West Lafayette, IN 47906, United States. Electronic address: sdonkin@purdue.edu.
authors.forename: Shawn S
authors.initials: SS
authors.lastname: Donkin
authors.affiliation: Department of Animal and Avian Sciences, University of Maryland, College Park, MD 20740, United States. Electronic address: dteegard@purdue.edu.
authors.forename: Brian
authors.initials: B
authors.lastname: Bequette
authors.affiliation: Interdepartmental Nutrition Program, Purdue University, West Lafayette, IN 47906, United States. Electronic address: teegarden@purdue.edu.
authors.forename: Dorothy
authors.initials: D
authors.lastname: Teegarden
mesh_terms: D026962:Amino Acid Transport System ASC / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism; D002461:Cell Line, Transformed; D002952:Citric Acid Cycle / Q000187:drug effects / Q000235:genetics; D004847:Epithelial Cells / Q000187:drug effects* / Q000378:metabolism / Q000473:pathology; D005260:Female; D005786:Gene Expression Regulation; D018698:Glutamic Acid / Q000378:metabolism; D005973:Glutamine / Q000031:analogs & derivatives / Q000037:antagonists & inhibitors* / Q000378:metabolism / Q000494:pharmacology; D006801:Humans; D007700:Kinetics; D042361:Mammary Glands, Human / Q000187:drug effects / Q000378:metabolism / Q000473:pathology; D015778:Minor Histocompatibility Antigens / Q000235:genetics / Q000378:metabolism; D015689:Oncogene Protein p21(ras) / Q000235:genetics / Q000378:metabolism; D011401:Promoter Regions, Genetic; D012333:RNA, Messenger / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism; D034741:RNA, Small Interfering / Q000235:genetics / Q000378:metabolism; D018719:Erb-b2 Receptor Tyrosine Kinases / Q000235:genetics / Q000378:metabolism; D018167:Receptors, Calcitriol / Q000235:genetics / Q000378:metabolism; D014807:Vitamin D / Q000031:analogs & derivatives* / Q000494:pharmacology; D030181:Vitamin D Response Element
chemical_list: D026962:Amino Acid Transport System ASC; D018698:Glutamic Acid; D005973:Glutamine; D015778:Minor Histocompatibility Antigens; D015689:Oncogene Protein p21(ras); D012333:RNA, Messenger; D034741:RNA, Small Interfering; D018719:Erb-b2 Receptor Tyrosine Kinases; D018167:Receptors, Calcitriol; D014807:Vitamin D; C420506:SLC1A5 protein, human; C579457:VDR protein, human; C097949:1,25-dihydroxyvitamin D; C001300:gamma-glutamine-4-nitroanilide; C508053:ERBB2 protein, human
keywords: 1,25-Dihydroxyvitamin D; Breast cancer; Energy metabolism; Glutamine metabolism; Harvey-ras oncogene; Prevention