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The role of hypoxia inducible factor-1 alpha in bypassing oncogene-induced senescence.

ABSTRACT: Oncogene induced senescence (OIS) is a sustained anti-proliferative response acutely induced in primary cells via activation of mitogenic oncogenes such as Ras/BRAF. This mechanism acts as an initial barrier preventing normal cells transformation into malignant cell. Besides oncogenic activation and DNA damage response (DDR), senescence is modulated by a plethora of other factors, and one of the most important one is oxygen tension of the tissue. The aim of this study was to determine the impact of hypoxia on RasV12-induced senescence in human diploid fibroblasts (HDFs). We showed here that hypoxia prevents execution of oncogene induced senescence (OIS), through a strong down-regulation of senescence hallmarks, such as SA- ?-galactosidase, H3K9me3, HP1?, p53, p21CIP1 and p16INK4a in association with induction of hypoxia inducible factor-1? (HIF-1?). In addition, hypoxia also decreased marks of H-RasV12-induced DDR in both cell lines through down-regulation of ATM/ATR, Chk1 and Chk2 phosphorylation as well as decreased ?-H2AX positivity. Utilizing shRNA system targeting HIF-1? we show that HIF-1? is directly involved in down regulation of p53 and its target p21CIP1 but not p16INK4a. In line with this finding we found that knock down of HIF-1? leads to a strong induction of apoptotic response, but not restoration of senescence in Ras expressing HDFs in hypoxia. This indicates that HIF-1? is an important player in early steps of tumorigenesis, leading to suppression of senescence through its negative regulation of p53 and p21CIP1. In our work we describe a mechanism through which hypoxia and specifically HIF-1? preclude cells from maintaining senescence-driven anti proliferative response. These findings indicate the possible mechanism through which hypoxic environment helps premalignant cells to evade impingement of cellular failsafe pathways.

SUBMITTER: Kilic Eren M

PROVIDER: S-EPMC4077769 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Excessive alcohol use increases the risk of developing respiratory infections partially due to impaired alveolar macrophage (AM) phagocytic capacity. Previously, we showed that chronic ethanol (EtOH) exposure led to mitochondrial derangements and diminished oxidative phosphorylation in AM. Since oxidative phosphorylation is needed to meet the energy demands of phagocytosis, EtOH mediated decreases in oxidative phosphorylation likely contribute to impaired AM phagocytosis. Treatment with the peroxisome proliferator-activated receptor gamma (PPARγ) ligand, pioglitazone (PIO), improved EtOH-mediated decreases in oxidative phosphorylation. In other models, hypoxia-inducible factor-1 alpha (HIF-1α) has been shown to mediate the switch from oxidative phosphorylation to glycolysis; however, the role of HIF-1α in chronic EtOH mediated derangements in AM has not been explored. We hypothesize that AM undergo a metabolic shift from oxidative phosphorylation to a glycolytic phenotype in response to chronic EtOH exposure. Further, we speculate that HIF-1α is a critical mediator of this metabolic switch. To test these hypotheses, primary mouse AM (mAM) were isolated from a mouse model of chronic EtOH consumption and a mouse AM cell line (MH-S) were exposed to EtOH in vitro. Expression of HIF-1α, glucose transporters (Glut1 and 4), and components of the glycolytic pathway (Pfkfb3 and PKM2), were measured by qRT-PCR and western blot. Lactate levels (lactate assay), cell energy phenotype (extracellular flux analyzer), glycolysis stress tests (extracellular flux analyzer), and phagocytic function (fluorescent microscopy) were conducted. EtOH exposure increased expression of HIF-1α, Glut1, Glut4, Pfkfb3, and PKM2 and shifted AM to a glycolytic phenotype. Pharmacological stabilization of HIF-1α via cobalt chloride treatment in vitro mimicked EtOH-induced AM derangements (increased glycolysis and diminished phagocytic capacity). Further, PIO treatment diminished HIF-1α levels and reversed glycolytic shift following EtOH exposure. These studies support a critical role for HIF-1α in mediating the glycolytic shift in energy metabolism of AM during excessive alcohol use.

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The role of hypoxia inducible factor-1 alpha in bypassing oncogene-induced senescence.

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