CTP limitation increases expression of CTP synthase in Lactococcus lactis.
CTP synthase is encoded by the pyrG gene and catalyzes the conversion of UTP to CTP. A Lactococcus lactis pyrG mutant with a cytidine requirement was constructed, in which beta-galactosidase activity in a pyrG-lacLM transcriptional fusion was used to monitor gene expression of pyrG. A 10-fold decrease in the CTP pool induced by cytidine limitation was found to immediately increase expression of the L. lactis pyrG gene. The final level of expression of pyrG is 37-fold higher than the uninduced level. CTP limitation has pronounced effects on central cellular metabolism, and both RNA and protein syntheses are inhibited. Expression of pyrG responds only to the cellular level of CTP, since expression of pyrG has no correlation to alterations in UTP, GTP, and ATP pool sizes. In the untranslated pyrG leader sequence a potential terminator structure can be identified, and this structure is required for regulation of the pyrG gene. It is possible to fold the pyrG leader in an alternative structure that would prevent the formation of the terminator. We suggest a model for pyrG regulation in L. lactis, and probably in other gram-positive bacteria as well, in which pyrG expression is directly dependent on the CTP concentration through an attenuator mechanism. At normal CTP concentrations a terminator is preferentially formed in the pyrG leader, thereby reducing expression of CTP synthase. At low CTP concentrations the RNA polymerase pauses at a stretch of C residues in the pyrG leader, thereby allowing an antiterminator to form and transcription to proceed. This model therefore does not include any trans-acting protein for sensing the CTP concentration as previously proposed for Bacillus subtilis.
Bacterial Physiology and Genetics, BioCentrum-DTU, Technical University of Denmark, DK-2800 Kgs. Lyngby, Denmark.
Casper Møller
CM
Jørgensen
Karin
K
Hammer
Jan
J
Martinussen
D001483:Base Sequence; D019731:Carbon-Nitrogen Ligases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D003570:Cytidine Triphosphate / Q000378:metabolism*; D015964:Gene Expression Regulation, Bacterial; D013294:Lactococcus lactis / Q000201:enzymology* / Q000235:genetics / Q000254:growth & development*; D008969:Molecular Sequence Data; D009154:Mutation; D011401:Promoter Regions, Genetic; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013728:Terminator Regions, Genetic; D014158:Transcription, Genetic
D003570:Cytidine Triphosphate; D019731:Carbon-Nitrogen Ligases; C019951:CTP synthetase