Linking Cancer Metabolism to DNA Repair and Accelerated Senescence

Abstract

Conventional wisdom ascribes metabolic reprogramming in cancer to meeting increased demands for intermediates to support rapid proliferation. Prior models have proposed benefits toward cell survival, immortality and stress resistance while the recent discovery of oncometabolites has shifted attention to chromatin targets affecting gene expression. To explore further effects of cancer metabolism and epigenetic deregulation, DNA repair kinetics were examined in cells treated with metabolic intermediates, oncometabolites and/or metabolic inhibitors by tracking resolution of double strand breaks (DSBs) in irradiated MCF7 breast cancer cells. Disrupting cancer metabolism revealed roles for both glycolysis and glutaminolysis in promoting DSB repair and preventing accelerated senescence after irradiation. Targeting pathways common to glycolysis and glutaminolysis uncovered opposing effects of the hexosamine biosynthetic pathway (HBP) and tricarboxylic acid (TCA) cycle. Treating cells with the HBP metabolite N-acetylglucosamine (GlcNAc) or augmenting protein O-GlcNAcylation with small molecules or RNAi targeting O-GlcNAcase enhanced DSB repair, while targeting O-GlcNAc transferase reversed GlcNAc’s effects. Opposing the HBP, TCA metabolites including α-ketoglutarate blocked DSB resolution. Strikingly, DNA repair could be restored by the oncometabolite 2-hydroxyglutarate (2-HG). Targeting downstream effectors of histone methylation and demethylation implicated the PRC1/2 polycomb complexes as the ultimate targets for metabolic regulation, reflecting known roles for Polycomb group proteins in non-homologous end-joining (NHEJ) DSB repair. Our findings that epigenetic effects of cancer metabolic reprogramming may promote DNA repair provide a molecular mechanism by which deregulation of metabolism may not only support cell growth but also maintain cell immortality, drive therapeutic resistance and promote genomic instability.

O-GlcNAcylation Enhances Double-Strand Break Repair, Promotes Cancer Cell Proliferation, and Prevents Therapy-Induced Senescence in Irradiated Tumors

“…Among pathways highlighted by the proteomic analysis was PRC2 methylation of histones and DNA (R-HSA-212300). We had previously identified the PRC2 HMT catalytic subunit Ezh2 as a candidate mediator of O-GlcNAc-dependent regulation of DSB repair (9). Consistent with prior in vitro results that Ezh2 abundance is regulated by OGT (28), IHC analysis of Ezh2 expression in the unirradiated and irradiated shOGT or alloxan-treated shScr tumors at 7 days after irradiation displayed decreased immunostaining compared with shOGA or GlcNAcor PUGNAc-treated shScr tumors or shScr controls (Supplementary Fig.…”

“…Strikingly, OGT also modifies H2AX on the ATM phosphorylation site S139, leading to decreased gH2AX formation at sites of DNA damage (34). In turn, our prior studies revealed functional links between cellular GlcNAc levels, O-GlcNAcylation, DDR, and DSB repair after irradiation (9). Feeding cells GlcNAc or blocking OGA accelerated DSB repair kinetics, while targeting OGT resulted in unrepaired breaks, persistent IRIF, and increased cellular senescence.…”

“…The recent discovery that 2-hydroxyglutarate (2-HG) acts as an oncometabolite to promote carcinogenesis without being further metabolized has led to the discovery of epigenetic enzymes as primary targets, suggesting mechanisms based on deregulated gene expression (44). Our prior work provided an alternative model, implicating 2-HG along with the HBP metabolite GlcNAc in promoting DNA DSB repair and blocking onset of cellular senescence via their effects on chromatin modification (9).…”

“…For two-photon confocal imaging of IRIF in tumors, tissue was excised, counterstained with Hoechst 33342, and imaged as described previously (9).…”

“…Using delayed IRIF resolution in irradiated cells as a screen for defects in DSB repair, we uncovered distinct roles for glycolysis and glutaminolysis, as well as their joint influence via the hexosamine biosynthesis pathway (HBP) in regulating DSB repair and therapy-induced senescence (9).…”