Abstract
In eukaryotes, adventitious oxidation of erythrose-4-phosphate, an intermediate of the pentose phosphate pathway (PPP), generates 4-phosphoerythronate (4PE), which inhibits 6-phosphogluconate dehydrogenase. 4PE is detoxified by metabolite-proofreading phosphatases such as yeast Pho13. Here, we report that a similar function is carried out in Bacillus subtilis by CpgA, a checkpoint protein known to be important for ribosome assembly, cell morphology and resistance to cell wall-targeting antibiotics. We find that ΔcpgA cells are intoxicated by glucose or other carbon sources that feed into the PPP, and that CpgA has high phosphatase activity with 4PE. Inhibition of 6-phosphogluconate dehydrogenase (GndA) leads to intoxication by 6-phosphogluconate, a potent inhibitor of phosphoglucose isomerase (PGI). The coordinated shutdown of PPP and glycolysis leads to metabolic gridlock. Overexpression of GndA, PGI, or yeast Pho13 suppresses glucose intoxication of ΔcpgA cells, but not cold sensitivity, a phenotype associated with ribosome assembly defects. Our results suggest that CpgA is a multifunctional protein, with genetically separable roles in ribosome assembly and metabolite proofreading.
Highlights
In eukaryotes, adventitious oxidation of erythrose-4-phosphate, an intermediate of the pentose phosphate pathway (PPP), generates 4-phosphoerythronate (4PE), which inhibits 6-phosphogluconate dehydrogenase. 4PE is detoxified by metabolite-proofreading phosphatases such as yeast Pho[13]
Several substrates of the PrkC kinase/PrpC phosphatase system have been linked to cell wall assembly and intrinsic antibiotic resistance, including WalR, GlmR/YvcK, GpsB, and CpgA5,6,15–20
We assessed the role of these genes in intrinsic resistance to cefuroxime (CEF), a β-lactam antibiotic that serves as a sensitive indicator of cell wall assembly defects in B. subtilis
Summary
Adventitious oxidation of erythrose-4-phosphate, an intermediate of the pentose phosphate pathway (PPP), generates 4-phosphoerythronate (4PE), which inhibits 6-phosphogluconate dehydrogenase. 4PE is detoxified by metabolite-proofreading phosphatases such as yeast Pho[13]. We report that a similar function is carried out in Bacillus subtilis by CpgA, a checkpoint protein known to be important for ribosome assembly, cell morphology and resistance to cell wall-targeting antibiotics. Overexpression of GndA, PGI, or yeast Pho[13] suppresses glucose intoxication of ΔcpgA cells, but not cold sensitivity, a phenotype associated with ribosome assembly defects. CpgA (circularly-permuted GTPase) is important for normal cell morphology, proper deposition of the peptidoglycan sacculus, and intrinsic resistance to antibiotics affecting both the ribosome and cell wall synthesis[5,10]. Whether or not these various phenotypes are related to the ribosomeassembly role is not resolved
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