Glutamine synthetase stabilizes the binding of GlnR to nitrogen fixation gene operators

Fernandes, Gabriela de Carvalho; Hauf, Ksenia; Sant’Anna, Fernando Hayashi; Forchhammer, Karl; Passaglia, Luciane Maria Pereira

doi:10.1111/febs.14021

Cited by 14 publications

(14 citation statements)

References 72 publications

(121 reference statements)

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“…Our results are different from some reports that P . riograndensis SBR5 GlnR is mainly the dimeric form [ 14 ] and B . subtilis GlnR is mainly monomeric form [ 20 ].…”

Section: Resultsmentioning

confidence: 99%

“…Our results are consistent with the reports that GS, in its feedback inhibited form, interacts with GlnR of P . riograndensis SBR5 [ 14 ] and GlnR and TnrA of B . subtilis [ 18 , 20 , 21 ].…”

Section: Resultsmentioning

confidence: 99%

“…Our current results are different from those obtained in P . riograndensis SBR5 where GlnR bound to the two GlnR-binding sites [PnifM(1) and PnifM(2)] of the main nif gene cluster at similar levels and whose GlnR affinity for site Ⅰ was slightly higher than for site Ⅱ [ 14 ].…”

Section: Resultsmentioning

confidence: 99%

“…GlnR is a central regulator of nitrogen metabolism in the class Bacilli, and the glnR gene in the diazotrophic Paenibacilli is not associated with the transferred nif gene cluster, indicating that Paenibacillus GlnR is indigenous. The recent studies with Surface Plasmon Resonance (SPR) experiments have demonstrated that GS stabilizes the binding of GlnR to nitrogen fixation gene operators in Paenibacillus riograndensis SBR5 [ 14 ]. However, these studies did not fully investigate the regulatory mechanism of GlnR in nitrogen fixation.…”

Section: Introductionmentioning

confidence: 99%

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Positive and negative regulation of transferred nif genes mediated by indigenous GlnR in Gram-positive Paenibacillus polymyxa

et al. 2018

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Ammonia is a major signal that regulates nitrogen fixation in most diazotrophs. Regulation of nitrogen fixation by ammonia in the Gram-negative diazotrophs is well-characterized. In these bacteria, this regulation occurs mainly at the level of nif (nitrogen fixation) gene transcription, which requires a nif-specific activator, NifA. Although Gram-positive and diazotrophic Paenibacilli have been extensively used as a bacterial fertilizer in agriculture, how nitrogen fixation is regulated in response to nitrogen availability in these bacteria remains unclear. An indigenous GlnR and GlnR/TnrA-binding sites in the promoter region of the nif cluster are conserved in these strains, indicating the role of GlnR as a regulator of nitrogen fixation. In this study, we for the first time reveal that GlnR of Paenibacillus polymyxa WLY78 is essentially required for nif gene transcription under nitrogen limitation, whereas both GlnR and glutamine synthetase (GS) encoded by glnA within glnRA operon are required for repressing nif expression under excess nitrogen. Dimerization of GlnR is necessary for binding of GlnR to DNA. GlnR in P. polymyxa WLY78 exists in a mixture of dimers and monomers. The C-terminal region of GlnR monomer is an autoinhibitory domain that prevents GlnR from binding DNA. Two GlnR-biding sites flank the -35/-10 regions of the nif promoter of the nif operon (nifBHDKENXhesAnifV). The GlnR-binding site Ⅰ (located upstream of -35/-10 regions of the nif promoter) is specially required for activating nif transcription, while GlnR-binding siteⅡ (located downstream of -35/-10 regions of the nif promoter) is for repressing nif expression. Under nitrogen limitation, GlnR dimer binds to GlnR-binding siteⅠ in a weak and transient association way and then activates nif transcription. During excess nitrogen, glutamine binds to and feedback inhibits GS by forming the complex FBI-GS. The FBI-GS interacts with the C-terminal domain of GlnR and stabilizes the binding affinity of GlnR to GlnR-binding site Ⅱ and thus represses nif transcription.

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“…Our results are different from some reports that P . riograndensis SBR5 GlnR is mainly the dimeric form [ 14 ] and B . subtilis GlnR is mainly monomeric form [ 20 ].…”

Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Positive and negative regulation of transferred nif genes mediated by indigenous GlnR in Gram-positive Paenibacillus polymyxa

et al. 2018

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“…A somewhat similar mechanism is present in Bacillus subtilis (Fisher and Wray, 2008 , 2009 ; Wray and Fisher, 2008 ). Also, work in Paenibacillus riograndensis showed that the binding of glnR to its targets is facilitated when glnR is complexed with a glutamine synthetase—again, upregulation of glnR may bypass the lack of glnA2 [Hauf et al, 2016 ; Fernandes et al, 2017 ].…”

Section: Discussionmentioning

confidence: 99%

The Unexpected Essentiality of glnA2 in Mycobacterium smegmatis Is Salvaged by Overexpression of the Global Nitrogen Regulator glnR, but Not by L-, D- or Iso-Glutamine

Rakovitsky

Goldberg

et al. 2018

Front. Microbiol.

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Nitrogen metabolism plays a central role in the physiology of microorganisms, and Glutamine Synthetase (GS) genes are present in virtually all bacteria. In M. tuberculosis, four GS genes are present, but only glnA1 is essential, whereas glnA2 was shown to be non-essential for in-vitro as well as in-vivo growth and pathogenesis, and is postulated to be involved in D-glutamine and iso-glutamine synthesis. Whilst investigating the activity of an antimicrobial compound in M. smegmatis, we found a spontaneous temperature-sensitive mutant in glnA2 (I133F), and used it to investigate the role of glnA2 in M. smegmatis. We deleted the native glnA2 and replaced it with a mutated allele. This re-created the temperature sensitivity—as after 3–4 seemingly normal division cycles, glnA2 became essential for growth. This essentiality could not be salvaged by neither L, D- nor iso-glutamine, suggesting an additional role of glnA2 in M. smegmatis over its role in M. tuberculosis. We also found that overexpression of the global nitrogen regulator glnR enabled bypassing the essentiality of glnA2, allowing the creation of a complete deletion mutant. The discrepancy between the importance of glnA2 in Mtb and M. smegmatis stresses the caution in which results in one are extrapolated to the other.

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Analysis of Protein–DNA Interactions Using Surface Plasmon Resonance and a ReDCaT Chip

Stevenson

Lawson

2021

Protein-Ligand Interactions

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Glutamine synthetase stabilizes the binding of GlnR to nitrogen fixation gene operators

Cited by 14 publications

References 72 publications

Positive and negative regulation of transferred nif genes mediated by indigenous GlnR in Gram-positive Paenibacillus polymyxa

Positive and negative regulation of transferred nif genes mediated by indigenous GlnR in Gram-positive Paenibacillus polymyxa

The Unexpected Essentiality of glnA2 in Mycobacterium smegmatis Is Salvaged by Overexpression of the Global Nitrogen Regulator glnR, but Not by L-, D- or Iso-Glutamine

Analysis of Protein–DNA Interactions Using Surface Plasmon Resonance and a ReDCaT Chip

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