Abstract
Glutamine analogs are potent suppressors of general glutamine metabolism with anti-cancer activity. 6-diazo-5-oxo-L-norleucine (DON) is an orally available glutamine analog which has been recently improved by structural modification for cancer treatment. Here, we explored the chemogenomic landscape of DON sensitivity using budding yeast as model organism. We identify evolutionarily conserved proteins that mediate cell resistance to glutamine analogs, namely Ura8CTPS1/2, Hpt1HPRT1, Mec1ATR, Rad53CHK1/CHK2 and Rtg1. We describe a function of Ura8 as inducible CTP synthase responding to inhibition of glutamine metabolism and propose a model for its regulation by CTP levels and Nrd1-dependent transcription termination at a cryptic unstable transcript. Disruption of the inducible CTP synthase under DON exposure hyper-activates the Mec1-Rad53 DNA damage response (DDR) pathway, which prevents chromosome breakage. Simultaneous inhibition of CTP synthase and Mec1 kinase synergistically sensitizes cells to DON, whereas CTP synthase over-expression hampers DDR mutant sensitivity. Using genome-wide suppressor screening, we identify factors promoting DON-induced CTP depletion (TORC1, glutamine transporter) and DNA breakage in DDR mutants. Together, our results identify CTP regulation and the Mec1-Rad53 DDR axis as key glutamine analog response pathways, and provide a rationale for the combined targeting of glutamine and CTP metabolism in DDR-deficient cancers.
Highlights
Glutamine is utilized in many metabolic processes such as the biosynthesis of pyrimidine and purine nucleotides, glutathione and non-essential amino acids, and the replenishment of tricarboxylic acid (TCA) cycle metabolites
Knowledge about the pathways that influence sensitivity towards glutamine metabolism inhibitors would help to tailor the use of such glutamine-targeting therapies
We show that five evolutionarily conserved proteins operating in pyrimidine synthesis (Ura8CTPS1/2), purine salvage (Hpt1HPRT1), regulation of glutamine metabolism (Rtg1) and replication stress response (Mec1ATR and Rad53CHK1/CHK2) are crucial for resistance to the glutamine analog DON
Summary
Glutamine is utilized in many metabolic processes such as the biosynthesis of pyrimidine and purine nucleotides, glutathione and non-essential amino acids, and the replenishment of tricarboxylic acid (TCA) cycle metabolites. Cancers commonly depend on glutamine supply as nitrogen and carbon source for rapid proliferation [1]. Various steps in glutamine metabolism have been explored as therapeutic targets to reduce intracellular glutamine concentration or inhibit glutamine-dependent reactions [2]. Glutamine mimetics inhibit glutamine-dependent reactions and efficiently kills tumor cells in vivo [3], but their use is hampered by considerable toxicity [4]. While prodrug strategies [4– 6] and combination treatments [7,8] are currently explored to reduce their side effects, knowledge on the major sensitivity and resistance mechanisms may improve drug combination designs and treatment rationale. The genetic determinants of glutamine mimetic sensitivity and resistance have not been systematically mapped
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