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.