title: Lethality of glnD null mutations in Azotobacter vinelandii is suppressible by prevention of glutamine synthetase adenylylation.
abstract: GlnD is a pivotal protein in sensing intracellular levels of fixed nitrogen and has been best studied in enteric bacteria, where it reversibly uridylylates two related proteins, PII and GlnK. The uridylylation state of these proteins determines the activities of glutamine synthetase (GS) and NtrC. Results presented here demonstrate that glnD is an essential gene in Azotobacter vinelandii. Null glnD mutations were introduced into the A. vinelandii genome, but none could be stably maintained unless a second mutation was present that resulted in unregulated activity of GS. One mutation, gln-71, occurred spontaneously to give strain MV71, which failed to uridylylate the GlnK protein. The second, created by design, was glnAY407F (MV75), altering the adenylylation site of GS. The gln-71 mutation is probably located in glnE, encoding adenylyltransferase, because introducing the Escherichia coli glnE gene into MV72, a glnD(+) derivative of MV71, restored the regulation of GS activity. GlnK-UMP is therefore apparently required for GS to be sufficiently deadenylylated in A. vinelandii for growth to occur. The DeltaglnD GS(c) isolates were Nif(-), which could be corrected by introducing a nifL mutation, confirming a role for GlnD in mediating nif gene regulation via some aspect of the NifL/NifA interaction. MV71 was unexpectedly NtrC(+), suggesting that A. vinelandii NtrC activity might be regulated differently than in enteric organisms.
authors.affiliation: Department of Plant Pathology, College of Agriculture, PO Box 210036, The University of Arizona, Tucson, AZ 85721, USA1.
authors.forename: Rita
authors.initials: R
authors.lastname: Colnaghi
authors.affiliation: Department of Plant Pathology, College of Agriculture, PO Box 210036, The University of Arizona, Tucson, AZ 85721, USA1.
authors.forename: Paul
authors.initials: P
authors.lastname: Rudnick
authors.affiliation: Department of Plant Pathology, College of Agriculture, PO Box 210036, The University of Arizona, Tucson, AZ 85721, USA1.
authors.forename: Luhong
authors.initials: L
authors.lastname: He
authors.affiliation: Department of Plant Pathology, College of Agriculture, PO Box 210036, The University of Arizona, Tucson, AZ 85721, USA1.
authors.forename: Andrew
authors.initials: A
authors.lastname: Green
authors.affiliation: Department of Plant Pathology, College of Agriculture, PO Box 210036, The University of Arizona, Tucson, AZ 85721, USA1.
authors.forename: Dalai
authors.initials: D
authors.lastname: Yan
authors.affiliation: Department of Plant Pathology, College of Agriculture, PO Box 210036, The University of Arizona, Tucson, AZ 85721, USA1.
authors.forename: Ethan
authors.initials: E
authors.lastname: Larson
authors.affiliation: Department of Plant Pathology, College of Agriculture, PO Box 210036, The University of Arizona, Tucson, AZ 85721, USA1.
authors.forename: Christina
authors.initials: C
authors.lastname: Kennedy
mesh_terms: D000225:Adenine / Q000378:metabolism*; D016948:Azotobacter vinelandii / Q000235:genetics* / Q000254:growth & development / Q000378:metabolism; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D015153:Blotting, Western; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000235:genetics; D029968:Escherichia coli Proteins; D005974:Glutamate-Ammonia Ligase / Q000235:genetics / Q000378:metabolism*; D009154:Mutation; D009586:Nitrogen Fixation / Q000235:genetics; D009713:Nucleotidyltransferases / Q000235:genetics*; D051922:PII Nitrogen Regulatory Proteins; D017384:Sequence Deletion; D015398:Signal Transduction; D015534:Trans-Activators; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism; D014529:Uridine / Q000378:metabolism
chemical_list: D001426:Bacterial Proteins; D002352:Carrier Proteins; D004268:DNA-Binding Proteins; D029968:Escherichia coli Proteins; C113488:GlnK protein, Azorhizobium caulinodans; C061338:NifA protein, Bacteria; D051922:PII Nitrogen Regulatory Proteins; D015534:Trans-Activators; D014157:Transcription Factors; C497876:glnG protein, E coli; C084296:nifL protein, Bacteria; C497878:glnK protein, E coli; D009713:Nucleotidyltransferases; C020658:glutamine-synthetase adenylyltransferase; C022515:regulatory protein uridylyltransferase; C081173:glutamine synthetase I; D005974:Glutamate-Ammonia Ligase; D000225:Adenine; D014529:Uridine