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Aerobic Inactivation ofRhizobium meliloti NifAinEscherichia coli IsMediated bylonandTwoNewlyIdentified Genes, snoBandsnoC

Aerobic Inactivation ofRhizobium meliloti NifAinEscherichia coli IsMediated bylonandTwoNewlyIdentified Genes, snoBandsnoC,EVA HUALA,ANNE L. MOON,FREDE

Aerobic Inactivation ofRhizobium meliloti NifAinEscherichia coli IsMediated bylonandTwoNewlyIdentified Genes, snoBandsnoC  
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level through thesensitivity ofNifAtooxygen.We havepreviously reported thattheNifA protein issensitive tooxygen inEscherichia coli as wellas inR.meliloti. To investigate whether the posttranslational regulation ofNifAisdependent on hostfactors conserved between R.meliloti andE.coli, we carried outa TnSmutagenesis ofE.coli andisolated mutantswithincreased NifAactivity underaerobic conditions. Fifteen insertion mutations occurred atthree unlinked loci. Onelocus isthepreviously character- ized Iongene;theother twoloci, which we havenamedsnoBandsnoC,define previously uncharacterized E. coli genes.Theproducts ofsnoCandIonaffect therateofNifAdegradation, whereas theproduct ofsnoBmay affect bothNifAdegradation andinactivation. A snoBlondoublemutantshoweda higher level ofNifA accumulation thandida Ionmutant, suggesting thatthesnoBproduct affects theability ofNifAtobedegraded byalon-independent pathway. Theeffects ofasnoCmutation anda Ionmutation were notadditive, suggesting thatthesnoCandIonproducts function inthesame degradative pathway. Nitrogen fixation inRhizobium meliloti andotherdiaz- otrophs isaninherently oxygensensitive process. Produc- tion ofnitrogenase inthepresence ofoxygeniseffectively prevented, since NifA,thetranscriptional activator ofthe nitrogenase structural genes, isregulated attwolevels by oxygen. First, theenvironmental sensor-regulator pairfixLJ stimulates nifAtranscription onlyundermicroaerobic con- ditions (9,11). Second, thenifA geneproduct (NifA) isitself inactive underaerobic conditions (2,14,15). Inactivation of R.meliloti NifAunderaerobic conditions also occurs when NifAisexpressed inEscherichia coli, suggesting either that NifAinactivation byoxygendoesnotrequire thepresence of anyhostfactors orthatsuchfactors arepresent inbothR. meliloti andE.coli. Incontrast totheNifAofR.meliloti, NifAfromthefree-living diazotroph Klebsiella pneumoniae isoxygensensitive onlyinthepresence ofnifL, whichis cotranscribed withnifAinK.pneumoniae butforwhichno homolog hasbeenfoundinanysymbiotic nitrogen-fixing species. HowthenifL product modulates NifAactivity inK. pneumoniae inresponse totheoxygenlevel remains un- clear. Previous workonR.meliloti NifAdemonstrated that smallchanges intheaminoacidsequence oftheamino terminus canhaveaprofound effect onthelevel ofNifA accumulation whenNifAisexpressed inE.coli (15). NifA translated fromthefirst in-frame AUG codon(NifA-AUG1) accumulates toverylowlevels whencells aregrownaero- bically andreaches slightly higher levels whencells are grownunder1%oxygen. Incontrast, several variant NifA proteins withaltered amino-terminal sequences accumulate
Published in 1991.
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