Adaptation to HIF-1 deficiency by upregulation of the AMP/ATP ratio and phosphofructokinase activation in hepatomas

• Published in: BMC cancer Vol. 11; no. 1; p. 198
• Main Authors: Golinska, Monika, Troy, Helen, Chung, Yuen-Li, McSheehy, Paul M, Mayr, Manuel, Yin, Xiaoke, Ly, Lucy, Williams, Kaye J, Airley, Rachel E, Harris, Adrian L, Latigo, John, Perumal, Meg, Aboagye, Eric O, Perrett, David, Stubbs, Marion, Griffiths, John R
• Format: Journal Article
• Language: English
• Published: England BioMed Central Ltd 25-05-2011; BioMed Central; BMC
• Subjects: Adaptation, Biological; Adenosine Monophosphate - metabolism; Adenosine Triphosphate - metabolism; Adenylate Kinase - metabolism; Allosteric Regulation; AMP/ATP ratio; Animals; Carcinoma, Hepatocellular - enzymology; Carcinoma, Hepatocellular - genetics; Carcinoma, Hepatocellular - metabolism; Carcinoma, Hepatocellular - pathology; Cell Proliferation; deficiency; Diagnosis; Enzyme Activation; Gene expression; Genetic aspects; Glucose - metabolism; Glucose Transport Proteins, Facilitative - metabolism; glucose uptake; Glycolysis - genetics; glycolytic enzymes; Hepa-1 tumours; Hepatoma; HIF-1β; Hypoxia-Inducible Factor 1 - deficiency; Immunohistochemistry; Liver Neoplasms - genetics; Liver Neoplasms - metabolism; Mice; Mice, Nude; PFK activation; Phosphofructokinase; Phosphofructokinases - metabolism; Physiological aspects; Proteomics; Risk factors; Up-Regulation; United Kingdom
• ISSN: 1471-2407; 1471-2407
• DOI: 10.1186/1471-2407-11-198

HIF-1 deficiency has marked effects on tumour glycolysis and growth. We therefore investigated the consequences of HIF-1 deficiency in mice, using the well established Hepa-1 wild-type (WT) and HIF-1β-deficient (c4) model. These mechanisms could be clinically relevant, since HIF-1 is now a therapeutic target.
Hepa-1 WT and c4 tumours grown in vivo were analysed by 18FDG-PET and 19FDG Magnetic Resonance Spectroscopy for glucose uptake; by HPLC for adenine nucleotides; by immunohistochemistry for GLUTs; by immunoblotting and by DIGE followed by tandem mass spectrometry for protein expression; and by classical enzymatic methods for enzyme activity.
HIF-1β deficient Hepa-1 c4 tumours grew significantly more slowly than WT tumours, and (as expected) showed significantly lower expression of many glycolytic enzymes. However, HIF-1β deficiency caused no significant change in the rate of glucose uptake in c4 tumours compared to WT when assessed in vivo by measuring fluoro-deoxyglucose (FDG) uptake. Immunohistochemistry demonstrated less GLUT-1 in c4 tumours, whereas GLUT-2 (liver type) was similar to WT. Factors that might upregulate glucose uptake independently of HIF-1 (phospho-Akt, c-Myc) were shown to have either lower or similar expression in c4 compared to WT tumours. However the AMP/ATP ratio was 4.5 fold higher (p < 0.01) in c4 tumours, and phosphofructokinase-1 (PFK-1) activity, measured at prevailing cellular ATP and AMP concentrations, was up to two-fold higher in homogenates of the deficient c4 cells and tumours compared to WT (p < 0.001), suggesting that allosteric PFK activation could explain their normal level of glycolysis. Phospho AMP-Kinase was also higher in the c4 tumours.
Despite their defective HIF-1 and consequent down-regulation of glycolytic enzyme expression, Hepa-1 c4 tumours maintain glucose uptake and glycolysis because the resulting low [ATP] high [AMP] allosterically activate PFK-1. This mechanism of resistance would keep glycolysis functioning and also result in activation of AMP-Kinase and growth inhibition; it may have major implications for the therapeutic activity of HIF inhibitors in vivo. Interestingly, this control mechanism does not involve transcriptional control or proteomics, but rather the classical activation and inhibition mechanisms of glycolytic enzymes.