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Hypoxia induces a lipogenic cancer cell phenotype via HIF1α-dependent and -independent pathways.

Valli A, Rodriguez M, Moutsianas L, Fischer R, Fedele V, Huang HL, Van Stiphout R, Jones D, Mccarthy M, Vinaxia M, Igarashi K, Sato M, Soga T, Buffa F, Mccullagh J, Yanes O, Harris A, Kessler B - Oncotarget (2015)

Bottom Line: To study the role of HIF1α in these processes, we used HCT116 colorectal cancer cells expressing endogenous HIF1α and cells in which the hif1α gene was deleted to characterize HIF1α-dependent and independent effects on hypoxia regulated lipid metabolites.Palmitate, stearate, PLD3 and PAFC16 were regulated in a HIF-independent manner.Our results demonstrate the impact of hypoxia on lipid metabolites, of which a distinct subset is regulated by HIF1α.

View Article: PubMed Central - PubMed

Affiliation: Weatherall Institute of Molecular Medicine, University of Oxford, Oxford, UK.

ABSTRACT
The biochemistry of cancer cells diverges significantly from normal cells as a result of a comprehensive reprogramming of metabolic pathways. A major factor influencing cancer metabolism is hypoxia, which is mediated by HIF1α and HIF2α. HIF1α represents one of the principal regulators of metabolism and energetic balance in cancer cells through its regulation of glycolysis, glycogen synthesis, Krebs cycle and the pentose phosphate shunt. However, less is known about the role of HIF1α in modulating lipid metabolism. Lipids serve cancer cells to provide molecules acting as oncogenic signals, energetic reserve, precursors for new membrane synthesis and to balance redox biological reactions. To study the role of HIF1α in these processes, we used HCT116 colorectal cancer cells expressing endogenous HIF1α and cells in which the hif1α gene was deleted to characterize HIF1α-dependent and independent effects on hypoxia regulated lipid metabolites. Untargeted metabolomics integrated with proteomics revealed that hypoxia induced many changes in lipids metabolites. Enzymatic steps in fatty acid synthesis and the Kennedy pathway were modified in a HIF1α-dependent fashion. Palmitate, stearate, PLD3 and PAFC16 were regulated in a HIF-independent manner. Our results demonstrate the impact of hypoxia on lipid metabolites, of which a distinct subset is regulated by HIF1α.

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Glycerol derivatives and phospholipids are dependent on HIF1α(a) TAG normalized levels detected by 1H-NMR in the organic phase of the cell extracts, reported as mean ±sd (n=3). (b) Glycerol normalized levels detected by 1H-NMR in the aqueous phase of the cell extracts, reported as mean ±sd (n=3). (c) Glycerophosphate normalized levels detected by CE/MS in the aqueous phase of the cell extracts, reported as mean ±sd (n=3). (d) Metabolic pathway generating the precursors utilized in the Kennedy pathway. Abbreviations:CMP, Cytidine monophosphate; Pi, phosphate inorganic; CDP-choline, Cytidine-diphosphocholine; PLD3, Phospholipase D3. (e) Choline, (f) phosphocholine, (g) phosphatidylcholine and (h) MAG normalized levels reported as mean ±sd, were detected by 1H-NMR in the organic and aqueous phase of the cell extracts (n=3). (i) Phospholipase D3 normalized levels detected by label-free quantitative proteomics analysis in HCT116 cells data are reported as mean ±sd (n=3).
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Figure 6: Glycerol derivatives and phospholipids are dependent on HIF1α(a) TAG normalized levels detected by 1H-NMR in the organic phase of the cell extracts, reported as mean ±sd (n=3). (b) Glycerol normalized levels detected by 1H-NMR in the aqueous phase of the cell extracts, reported as mean ±sd (n=3). (c) Glycerophosphate normalized levels detected by CE/MS in the aqueous phase of the cell extracts, reported as mean ±sd (n=3). (d) Metabolic pathway generating the precursors utilized in the Kennedy pathway. Abbreviations:CMP, Cytidine monophosphate; Pi, phosphate inorganic; CDP-choline, Cytidine-diphosphocholine; PLD3, Phospholipase D3. (e) Choline, (f) phosphocholine, (g) phosphatidylcholine and (h) MAG normalized levels reported as mean ±sd, were detected by 1H-NMR in the organic and aqueous phase of the cell extracts (n=3). (i) Phospholipase D3 normalized levels detected by label-free quantitative proteomics analysis in HCT116 cells data are reported as mean ±sd (n=3).

Mentions: Hypoxia caused an increase of TAG levels and the absence of HIF1α strongly reinforced this effect in hif1α−/− cells. This effect was also observed in hif1α−/− normoxic cells, indicating that HIF1α suppresses hypoxic TAG accumulation (Figure 6a; Table 1). Hydrolysis of TAG generates free glycerol that can be phosphorylated to glycerophosphate. Interestingly, the level of these two metabolites showed an opposite distribution with HIF1α causing an accumulation of glycerol and a suppression of glycerophosphate in hypoxic wild type HCT116 cells (Figure 6b and c; Table 1). The levels of MAG, choline (Cho) and phopsphocholine (PCho), all involved in phosphatidylcholine (PC) biosynthesis through the Kennedy pathway (Figure 6d), were unaltered in normoxia and hypoxia-induced wild type cells. Surprisingly, only hif1α−/− cells accumulated MAG, Cho, PCho and PC levels under hypoxia, thus underlining the suppressive HIF1α-dependent effect on this metabolic pathway (Figure 6e, f, g and h; Table 1). The levels of phospholipase D3 (PLD3), mediating PC catabolism resulting in phosphatidate and Cho (Figure 6d), were down regulated in both wild type and hif1α−/− hypoxic cells in a HIF1α-independent manner. Levels were unchanged under normoxia (Figure 6i; Table 2).


Hypoxia induces a lipogenic cancer cell phenotype via HIF1α-dependent and -independent pathways.

Valli A, Rodriguez M, Moutsianas L, Fischer R, Fedele V, Huang HL, Van Stiphout R, Jones D, Mccarthy M, Vinaxia M, Igarashi K, Sato M, Soga T, Buffa F, Mccullagh J, Yanes O, Harris A, Kessler B - Oncotarget (2015)

Glycerol derivatives and phospholipids are dependent on HIF1α(a) TAG normalized levels detected by 1H-NMR in the organic phase of the cell extracts, reported as mean ±sd (n=3). (b) Glycerol normalized levels detected by 1H-NMR in the aqueous phase of the cell extracts, reported as mean ±sd (n=3). (c) Glycerophosphate normalized levels detected by CE/MS in the aqueous phase of the cell extracts, reported as mean ±sd (n=3). (d) Metabolic pathway generating the precursors utilized in the Kennedy pathway. Abbreviations:CMP, Cytidine monophosphate; Pi, phosphate inorganic; CDP-choline, Cytidine-diphosphocholine; PLD3, Phospholipase D3. (e) Choline, (f) phosphocholine, (g) phosphatidylcholine and (h) MAG normalized levels reported as mean ±sd, were detected by 1H-NMR in the organic and aqueous phase of the cell extracts (n=3). (i) Phospholipase D3 normalized levels detected by label-free quantitative proteomics analysis in HCT116 cells data are reported as mean ±sd (n=3).
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Figure 6: Glycerol derivatives and phospholipids are dependent on HIF1α(a) TAG normalized levels detected by 1H-NMR in the organic phase of the cell extracts, reported as mean ±sd (n=3). (b) Glycerol normalized levels detected by 1H-NMR in the aqueous phase of the cell extracts, reported as mean ±sd (n=3). (c) Glycerophosphate normalized levels detected by CE/MS in the aqueous phase of the cell extracts, reported as mean ±sd (n=3). (d) Metabolic pathway generating the precursors utilized in the Kennedy pathway. Abbreviations:CMP, Cytidine monophosphate; Pi, phosphate inorganic; CDP-choline, Cytidine-diphosphocholine; PLD3, Phospholipase D3. (e) Choline, (f) phosphocholine, (g) phosphatidylcholine and (h) MAG normalized levels reported as mean ±sd, were detected by 1H-NMR in the organic and aqueous phase of the cell extracts (n=3). (i) Phospholipase D3 normalized levels detected by label-free quantitative proteomics analysis in HCT116 cells data are reported as mean ±sd (n=3).
Mentions: Hypoxia caused an increase of TAG levels and the absence of HIF1α strongly reinforced this effect in hif1α−/− cells. This effect was also observed in hif1α−/− normoxic cells, indicating that HIF1α suppresses hypoxic TAG accumulation (Figure 6a; Table 1). Hydrolysis of TAG generates free glycerol that can be phosphorylated to glycerophosphate. Interestingly, the level of these two metabolites showed an opposite distribution with HIF1α causing an accumulation of glycerol and a suppression of glycerophosphate in hypoxic wild type HCT116 cells (Figure 6b and c; Table 1). The levels of MAG, choline (Cho) and phopsphocholine (PCho), all involved in phosphatidylcholine (PC) biosynthesis through the Kennedy pathway (Figure 6d), were unaltered in normoxia and hypoxia-induced wild type cells. Surprisingly, only hif1α−/− cells accumulated MAG, Cho, PCho and PC levels under hypoxia, thus underlining the suppressive HIF1α-dependent effect on this metabolic pathway (Figure 6e, f, g and h; Table 1). The levels of phospholipase D3 (PLD3), mediating PC catabolism resulting in phosphatidate and Cho (Figure 6d), were down regulated in both wild type and hif1α−/− hypoxic cells in a HIF1α-independent manner. Levels were unchanged under normoxia (Figure 6i; Table 2).

Bottom Line: To study the role of HIF1α in these processes, we used HCT116 colorectal cancer cells expressing endogenous HIF1α and cells in which the hif1α gene was deleted to characterize HIF1α-dependent and independent effects on hypoxia regulated lipid metabolites.Palmitate, stearate, PLD3 and PAFC16 were regulated in a HIF-independent manner.Our results demonstrate the impact of hypoxia on lipid metabolites, of which a distinct subset is regulated by HIF1α.

View Article: PubMed Central - PubMed

Affiliation: Weatherall Institute of Molecular Medicine, University of Oxford, Oxford, UK.

ABSTRACT
The biochemistry of cancer cells diverges significantly from normal cells as a result of a comprehensive reprogramming of metabolic pathways. A major factor influencing cancer metabolism is hypoxia, which is mediated by HIF1α and HIF2α. HIF1α represents one of the principal regulators of metabolism and energetic balance in cancer cells through its regulation of glycolysis, glycogen synthesis, Krebs cycle and the pentose phosphate shunt. However, less is known about the role of HIF1α in modulating lipid metabolism. Lipids serve cancer cells to provide molecules acting as oncogenic signals, energetic reserve, precursors for new membrane synthesis and to balance redox biological reactions. To study the role of HIF1α in these processes, we used HCT116 colorectal cancer cells expressing endogenous HIF1α and cells in which the hif1α gene was deleted to characterize HIF1α-dependent and independent effects on hypoxia regulated lipid metabolites. Untargeted metabolomics integrated with proteomics revealed that hypoxia induced many changes in lipids metabolites. Enzymatic steps in fatty acid synthesis and the Kennedy pathway were modified in a HIF1α-dependent fashion. Palmitate, stearate, PLD3 and PAFC16 were regulated in a HIF-independent manner. Our results demonstrate the impact of hypoxia on lipid metabolites, of which a distinct subset is regulated by HIF1α.

Show MeSH
Related in: MedlinePlus