|Topic:||Glucose Metabolism and Cancer: The Hypoxia Induced Pathway .|
|Details:||As described in the previous section, altered metabolism of glucose is a hallmark of all types of cancer. The diversion of glucose carbons into biomass in cancer cells necessitates an increased delivery of glucose into these cells and an increase in the rate of anaerobic glycolysis to lactate. This is accomplished by an increase in the expression of genes encoding glucose transporters and glycolytic enzymes. These transcriptional changes can be observed in over 70% of human cancers and is driven in part through activation of the hypoxia induced factor 1 (HIF-1) pathway and by increased expression of various proto-oncogenes and decreased expression of various tumor suppressors.
The HIF-1 pathway, which is activated by conditions of hypoxia (low oxygen tension), is a major homeostatic mechanism for cellular responses to changes in the level of oxygen within cells. HIF-1 is a heterodimeric complex composed of an α-subunit and a β-subunit. The β-subunit is constitutively expressed while the α-subunit expression and activity are increased in response to changes in cellular oxygen content. There are three related HIF complexes identified as HIF-1, HIF-2, and HIF-3 that are defined by the particular α-subunit of the complex. The activity of the HIF-1 and HIF-2 complexes are highly similar in their responses to hypoxia. Less detail is known regarding the HIF-3 complex. Humans express three α-subunit genes, HIF1α (HIF1A gene), HIF2α (EPAS1 gene, for endothelial PAS domain protein 1), and HIF3α (HIF3A gene). The PAS domain is so-called because of the three proteins in which the domain was originally identified: Per (period circadian protein), ARNT (aryl hydrocarbon receptor nuclear translocator), and Sim (simple-minded protein).The original β-subunit (HIF1β) was initially identified and characterized as ARNT. Humans express two ARNT-related genes (ARNT2 and ARNTL), however, the encoded proteins are not components of the HIF complexes. Normally the HIF1α subunits are degraded in the presence of oxygen due to polyubiquitylation. Polyubiquitylation is a key modification directing proteins for rapid degradation by the proteosome machinery. Expression of the HIF1A gene is ubiquitous, whereas expression of the HIF2A gene is more restricted being primarily found active in interstitial cells, endothelial cells, and parenchymal cells. Expression patterns of the HIF3A gene are less well defined and the gene generates multiple splice variant mRNAs, some of which lack the transcriptional transactivation domain.
The HIF α-subunits possess an oxygen-dependent degradation (ODD) domain. The ODD domain is hydroxylated by a member of the prolyl hydroxylase domain (PHD) family of proline hydroxylating enzymes. The prolyl hydroxylase family includes the enzymes that incorporate hydroxyl groups into proline residues in collagens of the extracellular matrix. The prolyl hydroxylases that hydroxylate HIF α-subunits are all ferrous (Fe2+) iron and 2-oxoglutarate (α-ketoglutarate)-dependent enzymes. The requirement of these enzymes for 2-oxoglutarate results in direct coupling of the activity this class of prolyl hydroxylases to metabolic processes that generate and utilize 2-oxoglutarate such as the TCA cycle. The 2-oxoglutarate-dependent prolyl hydroxylase enzymes are identified as PHD1 (encoded by the EGLN2 gene), PHD2 (encoded by the EGLN1 gene) and PHD3 (encoded by the EGLN3 gene). The designation of EGLN refers to the fact that these three genes are homologs of the Caenorhabditis elegans egg laying-9 (Egl-9) gene. Hydroxylation of HIF α-subunits renders the proteins susceptible to proteosomal degradation under normoxic cellular conditions. In response to proline hydroxylation the ubiquitin ligase encoded by the von Hippel-Lindau (VHL) gene binds to the HIF α-subunit proteins and catalyzes their polyubiquitination.
Expression of PHD1 is highest in the testes with lower level expression seen in brain, liver, kidney, and heart. Expression of PHD2 is observed in most tissues. Expression of PHD3 is highest within cardiac myocytes. The activities of PHD2 and PHD3 are strongly induced by changes in oxygen concentrations. The hydroxylation reactions catalyzed by the PHD enzymes require molecular oxygen (O2) in addition to the Fe2+ and 2-oxoglutarate, therefore, reductions in oxygen content will result in loss of their activity. The products of the PHD enzymes are a trans-4-hydroxyproline residues, CO2, and succinate. As indicated, expression of the HIF1A gene is ubiquitous and this pattern is maintained under normal oxygen availability (normoxic conditions). The activity of the HIF1α protein is regulated by being hydroxylated on two prolines (P405 and P531) in the ODD. The presence of the trans-4-hydroxyproline residues increases the binding of the VHL encoded protein by over 1000 fold. Given that the PHD enzymes require O2 as a substrate for the hydroxylation reaction, when conditions of hypoxia exist the HIFα subunits escape hydroxylation and are, therefore, not ubiquitinated. Under hypoxic conditions the stabilized HIFα subunits migrate to the nucleus and dimerize with the HIFβ subunit and activate the expression of target genes. The activity of HIF1α is also regulated via the hydroxylation of a specific asparagine residue (N803) found in the C-terminal transactivation domain. The N803 hydroxylation is catalyzed by another 2-oxoglutarate-dependent dioxygenase originally identified as factor-inhibiting HIF-1 (FIH1; also identified as FIH). FIH1 is encoded by the HIF1AN (hypoxia inducible factor 1 alpha subunit inhibitor) gene. The consequences of the β-hydroxylation of N803 are that HIF1α can no longer interact with the transcriptional co-activators CBP [cAMP-response element-binding protein (CREB)- binding protein] and p300 (CBP/p300) resulting in inhibition of HIF-1 activity.
Both HIF1α and HIF1β proteins are transcription factors that contain basic helix-loop-helix domains that allow them to dimerize and to bind to DNA sequences in target genes. Both HIF1α and HIF1β also have a PAS domain. The HIF1α subunits also contain two transactivation domains (TAD), which regulate the expression of HIF-1 target genes. As indicated, the transcriptional coactivator proteins CBP and p300 (CBP/p300) interact with HIF1α and this interaction occurs through the C-terminal TAD. CBP and p300 modify chromatin structure, and thereby transcriptional activity, via their lysine acetyltransferase (KAT) activity which acetylates the nucleosomal histones.
The normal role of the HIF-1 pathway is to promote the delivery of oxygen and nutrients to the oxygen-deprived tissue via the stimulation of neovascularization (angiogenesis). The microenvironment that surrounds most solid tumors is highly hypoxic and, therefore, the ability of the tumor cells to proliferate requires the ability to acquire oxygen and nutrients. This is accomplished, in large part, through the activation of the HIF-1 pathway which is considered to be a modulator in the transactivation of genes implicated in the altered metabolism observed in cancer cells. Several of the metabolic regulatory genes that are activated by HIF-1 include the pyruvate kinase M (PKM) gene, described in detail in the previous section, the fructose-1,6-bisphosphate aldolase (ALDOA) gene, the pyruvate dehydrogenase kinase 1 (PDK1) gene, the GLUT1 gene, and the lactate dehydrogenase A (LDHA) gene. The altered patterns of metabolism lead to the accumulation of diverse metabolites in the microenvironment that promote tumor growth and contribute to the ability of the tumor cells to metastasize. Activation of HIF-1 in cancer cells results in restriction of glucose entry into the mitochondrial oxidative phosphorylation pathway via inhibition of the pyruvate dehydrogenase complex, PDHc. The inhibition of the PDHc is effected via HIF-1 stimulated expression of the PDK1 gene. The regulation of the PDHc by phosphorylation is discussed in detail in the TCA Cycle page. The altered pyruvate kinase isoforms, as occurs in cancer (see previous section), coupled with the activation of the HIF-1 pathway results in diversion of glucose metabolism into the pentose phosphate pathway as well as into increased lactate production. The conversion of pyruvate into lactate is enhanced in the context of activated HIF-1 since this transcription factor activates the expression of the LDHA gene. The increased production of lactate, by cancer cells, contributes to the acidification of the tumor microenvironment which, in turn, promote further activation of the HIF-1 pathway. Lactate accumulation also results in pyruvate accumulation in cancer cells. Pyruvate is a known inhibitor of the prolyl hydroxylases that hydroxylate the HIF1α subunit proteins. Loss of HIF1α proline hydroxylation results in increased HIF1α stability and, therefore, increased HIF-1 transcriptional activity. Thus, accumulation of lactate and pyruvate, which occurs as a result of both altered pyruvate kinase gene expression and activation of the HIF-1 pathway, further promotes activation of the HIF-1 pathway leading to a controlled and enhanced metabolic profile within cancer cells.
COPYRIGHT ©2019 . All Rights Reserved.