Nac-mediated repression of the serA promoter of Escherichia coli.
Escherichia coli and related bacteria contain two paralogous PII-like proteins involved in nitrogen regulation, the glnB product, PII, and the glnK product, GlnK. Previous studies have shown that cells lacking both PII and GlnK have a severe growth defect on minimal media, resulting from elevated expression of the Ntr regulon. Here, we show that this growth defect is caused by activity of the nac product, Nac, a LysR-type transcription factor that is part of the Ntr regulon. Cells with elevated Ntr expression that also contain a null mutation in nac displayed growth rates on minimal medium similar to the wild type. When expressed from high-copy plasmids, Nac imparts a growth defect to wild-type cells in an expression level-dependent manner. Neither expression of Nac nor lack thereof significantly affected Ntr gene expression, suggesting that the activity of Nac at one or more promoters outside the Ntr regulon was responsible for its effects. The growth defect of cells lacking both PII and GlnK was also eliminated upon supplementation of minimal medium with serine or glycine for solid medium or with serine or glycine and glutamine for liquid medium. These observations suggest that high Nac expression results in a reduction in serine biosynthesis. beta-Galactosidase activity expressed from a Mu d1 insertion in serA was reduced approximately 10-fold in cells with high Nac expression. We hypothesize that one role of Nac is to limit serine biosynthesis as part of a cellular mechanism to reduce metabolism in a co-ordinated manner when cells become starved for nitrogen.
Department of Biological Chemistry, University of Michigan Medical School, 1301 E. Catherine, Ann Arbor, MI 48109-0606, USA.
Timothy A
TA
Blauwkamp
Alexander J
AJ
Ninfa
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000096:biosynthesis / Q000235:genetics / Q000502:physiology*; D002352:Carrier Proteins / Q000096:biosynthesis / Q000235:genetics / Q000502:physiology*; D002364:Caseins / Q000494:pharmacology; D003470:Culture Media / Q000494:pharmacology; D004268:DNA-Binding Proteins / Q000502:physiology*; D004926:Escherichia coli / Q000187:drug effects / Q000235:genetics*; D029968:Escherichia coli Proteins / Q000096:biosynthesis / Q000235:genetics* / Q000502:physiology*; D015964:Gene Expression Regulation, Bacterial / Q000187:drug effects; D005798:Genes, Bacterial; D016153:Genes, Suppressor; D005973:Glutamine / Q000494:pharmacology; D005998:Glycine / Q000494:pharmacology; D006801:Humans; D008969:Molecular Sequence Data; D009584:Nitrogen / Q000378:metabolism; D009586:Nitrogen Fixation / Q000235:genetics; D051922:PII Nitrogen Regulatory Proteins; D011401:Promoter Regions, Genetic / Q000235:genetics*; D011492:Protein Hydrolysates / Q000494:pharmacology; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D016415:Sequence Alignment; D017385:Sequence Homology; D012694:Serine / Q000096:biosynthesis / Q000494:pharmacology; D014157:Transcription Factors / Q000502:physiology*
D001426:Bacterial Proteins; D002352:Carrier Proteins; D002364:Caseins; D003470:Culture Media; D004268:DNA-Binding Proteins; D029968:Escherichia coli Proteins; C113488:GlnK protein, Azorhizobium caulinodans; C072712:NAC protein, bacteria; C489526:Nac protein, E coli; D051922:PII Nitrogen Regulatory Proteins; D011492:Protein Hydrolysates; D011993:Recombinant Fusion Proteins; C465559:SerA protein, E coli; D014157:Transcription Factors; C026375:trypticase-soy broth; D005973:Glutamine; D012694:Serine; C404143:PIID regulatory protein, Bacteria; D009584:Nitrogen; D005998:Glycine