The plasma membrane lactate transporter MCT4, but not MCT1, is up-regulated by hypoxia through a HIF-1alpha-dependent mechanism.
The monocarboxylate transporter MCT4 mediates lactic acid efflux from most tissues that are dependent on glycolysis for their ATP production. Here we demonstrate that expression of MCT4 mRNA and protein was increased >3-fold by a 48-h exposure to 1% O(2), whereas MCT1 expression was not increased. The effect was mimicked by CoCl(2) (50 microm), suggesting transcriptional regulation by hypoxia-inducible factor 1alpha (HIF-1alpha). The predicted promoters for human MCT1, MCT2, and MCT4 were cloned into the pGL3 vector and shown to be active (luciferase luminescence) under basal conditions. Only the MCT4 promoter was activated (>2-fold) by hypoxia. No response was found in cells lacking HIF-1alpha. Four potential hypoxia-response elements were identified, but deletion analysis implicated only two in the hypoxia response. These were just upstream from the transcription start site and also found in the mouse MCT4 promoter. Mutation of site 2 totally abolished the hypoxic response, whereas mutation of site 1 only reduced the response. Gel-shift analysis demonstrated that nuclear extracts of hypoxic but not normoxic HeLa cells contained two transcription factors that bound to DNA probes containing these hypoxia-response elements. The major shifted band was abolished by mutation of site 2, and supershift analysis confirmed that HIF-1alpha bound to this site. Binding of the second factor was abolished by mutation of site 1. We conclude that MCT4, like other glycolytic enzymes, is up-regulated by hypoxia through a HIF-1alpha-mediated mechanism. This adaptive response allows the increased lactic acid produced during hypoxia to be rapidly lost from the cell.
Ullah
Mohammed S
MS
Department of Biochemistry, University of Bristol, School of Medical Sciences, University Walk, Bristol BS8 1TD, United Kingdom.
Davies
Andrew J
AJ
Halestrap
Andrew P
AP
D000222:Adaptation, Physiological; D000818:Animals; D016466:CHO Cells; D019556:COS Cells; D018797:Cell Cycle Proteins / Q000096:biosynthesis* / Q000502:physiology; D015687:Cell Hypoxia; D002462:Cell Membrane / Q000502:physiology; D002522:Chlorocebus aethiops; D006224:Cricetinae; D020869:Gene Expression Profiling; D006367:HeLa Cells; D006801:Humans; D051795:Hypoxia-Inducible Factor 1, alpha Subunit / Q000502:physiology*; D019344:Lactic Acid / Q000378:metabolism; D027501:Monocarboxylic Acid Transporters / Q000096:biosynthesis* / Q000502:physiology; D009124:Muscle Proteins / Q000096:biosynthesis* / Q000502:physiology; D032383:Myocytes, Cardiac; D015513:Oncogene Proteins / Q000096:biosynthesis* / Q000502:physiology; D010100:Oxygen; D011401:Promoter Regions, Genetic; D012333:RNA, Messenger / Q000096:biosynthesis
D018797:Cell Cycle Proteins; C497442:HIF1A protein, human; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; C115081:MCTS1 protein, human; D027501:Monocarboxylic Acid Transporters; D009124:Muscle Proteins; D015513:Oncogene Proteins; D012333:RNA, Messenger; C406962:SLC16A4 protein, human; D019344:Lactic Acid; D010100:Oxygen