title: Modulation of anti-cancer drug sensitivity through the regulation of mitochondrial activity by adenylate kinase 4.
abstract: BACKGROUND
  Adenylate kinase is a key enzyme in the high-energy phosphoryl transfer reaction in living cells. An isoform of this enzyme, adenylate kinase 4 (AK4), is localized in the mitochondrial matrix and is believed to be involved in stress, drug resistance, malignant transformation in cancer, and ATP regulation. However, the molecular basis for the AK4 functions remained to be determined.
  
  
  METHODS
  HeLa cells were transiently transfected with an AK4 small interfering RNA (siRNA), an AK4 short hairpin RNA (shRNA) plasmid, a control shRNA plasmid, an AK4 expression vector, and a control expression vector to examine the effect of the AK4 expression on cell proliferation, sensitivity to anti-cancer drug, metabolome, gene expression, and mitochondrial activity.
  
  
  RESULTS
  AK4 knockdown cells treated with short hairpin RNA increased ATP production and showed greater sensitivity to hypoxia and anti-cancer drug, cis-diamminedichloro-platinum (II) (CDDP). Subcutaneous grafting AK4 knockdown cells into nude mice revealed that the grafted cells exhibited both slower proliferation and reduced the tumor sizes in response to CDDP. AK4 knockdown cell showed a increased oxygen consumption rate with FCCP treatment, while AK4 overexpression lowered it. Metabolome analysis showed the increased levels of the tricarboxylic acid cycle intermediates, fumarate and malate in AK4 knockdown cells, while AK4 overexpression lowered them. Electron microscopy detected the increased mitochondrial numbers in AK4 knockdown cells. Microarray analysis detected the increased gene expression of two key enzymes in TCA cycle, succinate dehydrogenase A (SDHA) and oxoglutarate dehydrogenease L (OGDHL), which are components of SDH complex and OGDH complex, supporting the metabolomic results.
  
  
  CONCLUSIONS
  We found that AK4 was involved in hypoxia tolerance, resistance to anti-tumor drug, and the regulation of mitochondrial activity. These findings provide a new potential target for efficient anticancer therapies by controlling AK4 expression.
authors.affiliation: Center for Regenerative Medicine, School of Medicine, Yamaguchi University, Ube, Japan.|Department of Gastroenterology and Hepatology, School of Medicine, Yamaguchi University, Ube, Japan.
authors.forename: Koichi
authors.initials: K
authors.lastname: Fujisawa
authors.affiliation: Department of Gastroenterology and Hepatology, School of Medicine, Yamaguchi University, Ube, Japan. terais@med.niigata-u.ac.jp.|Division of Gastroenterology and Hepatology, School of Medical and Dental Sciences, Niigata University, 1-757 Asahimachidori, Chuo-Ku, Niigata, 951-8510, Japan. terais@med.niigata-u.ac.jp.
authors.forename: Shuji
authors.initials: S
authors.lastname: Terai
authors.affiliation: Department of Gastroenterology and Hepatology, School of Medicine, Yamaguchi University, Ube, Japan.
authors.forename: Taro
authors.initials: T
authors.lastname: Takami
authors.affiliation: Department of Gastroenterology and Hepatology, School of Medicine, Yamaguchi University, Ube, Japan.
authors.forename: Naoki
authors.initials: N
authors.lastname: Yamamoto
authors.affiliation: Department of Gastroenterology and Hepatology, School of Medicine, Yamaguchi University, Ube, Japan.|Department of Oncology and Laboratory Medicine, School of Medicine, Yamaguchi University, Ube, Japan.
authors.forename: Takahiro
authors.initials: T
authors.lastname: Yamasaki
authors.affiliation: Department of Gastroenterology and Hepatology, School of Medicine, Yamaguchi University, Ube, Japan.|Department of Oncology and Laboratory Medicine, School of Medicine, Yamaguchi University, Ube, Japan.
authors.forename: Toshihiko
authors.initials: T
authors.lastname: Matsumoto
authors.affiliation: Department of Organ Anatomy, School of Medicine, Yamaguchi University, Ube, Japan.
authors.forename: Kazuhito
authors.initials: K
authors.lastname: Yamaguchi
authors.affiliation: Department of Organ Anatomy, School of Medicine, Yamaguchi University, Ube, Japan.
authors.forename: Yuji
authors.initials: Y
authors.lastname: Owada
authors.affiliation: Department of Developmental and Regenerative Biology, Medical Research Institute, Tokyo Medical and Dental University, 1-5-45 Yushima, Bunkyo-ku, Tokyo, 113-8510, Japan.
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Nishina
authors.affiliation: Department of Molecular Biology, Institute of Biomedical Sciences, Tokushima University School, Tokushima, Japan.
authors.forename: Takafumi
authors.initials: T
authors.lastname: Noma
authors.affiliation: Center for Regenerative Medicine, School of Medicine, Yamaguchi University, Ube, Japan.|Department of Gastroenterology and Hepatology, School of Medicine, Yamaguchi University, Ube, Japan.
authors.forename: Isao
authors.initials: I
authors.lastname: Sakaida
mesh_terms: D000263:Adenylate Kinase / Q000378:metabolism*; D000818:Animals; D015687:Cell Hypoxia; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D019008:Drug Resistance, Neoplasm; D005260:Female; D015972:Gene Expression Regulation, Neoplastic; D006367:HeLa Cells; D006801:Humans; D055442:Metabolome; D051379:Mice; D008928:Mitochondria / Q000378:metabolism*; D009369:Neoplasms / Q000378:metabolism*; D023041:Xenograft Model Antitumor Assays
chemical_list: D000263:Adenylate Kinase
keywords: Adenylate kinase; Drug resistance; Energy metabolism; Flux analysis; Hypoxia; Metabolome; Mitochondria