The NtrY/NtrX System of Sinorhizobium meliloti GR4 Regulates Motility, EPS I Production, and Nitrogen Metabolism but Is Dispensable for Symbiotic Nitrogen Fixation

Abstract

Sinorhizobium meliloti can translocate over surfaces. However, little is known about the regulatory mechanisms that control this trait and its relevance for establishing symbiosis with alfalfa plants. To gain insights into this field, we isolated Tn5 mutants of S. meliloti GR4 with impaired surface motility. In mutant strain GRS577, the transposon interrupted the ntrY gene encoding the sensor kinase of the NtrY/NtrX two-component regulatory system. GRS577 is impaired in flagella synthesis and overproduces succinoglycan, which is responsible for increased biofilm formation. The mutant also shows altered cell morphology and higher susceptibility to salt stress. GRS577 induces nitrogen-fixing nodules in alfalfa but exhibits decreased competitive nodulation. Complementation experiments indicate that both ntrY and ntrX account for all the phenotypes displayed by the ntrY::Tn5 mutant. Ectopic overexpression of VisNR, the motility master regulator, was sufficient to rescue motility and competitive nodulation of the transposant. A transcriptome profiling of GRS577 confirmed differential expression of exo and flagellar genes, and led to the demonstration that NtrY/NtrX allows for optimal expression of denitrification and nifA genes under microoxic conditions in response to nitrogen compounds. This study extends our knowledge of the complex role played by NtrY/NtrX in S. meliloti.

Genome‐wide identification of genes directly regulated by ChvI and a consensus sequence for ChvI binding in Sinorhizobium meliloti

“…Genes with altered expression due to syrA (exopolysaccharide regulator) overexpression showed especially strong overlap with ChvI regulon genes, which had led us to propose that SyrA may interact with ExoS/ChvI (Barnett and Long, ). In comparing the list of ChvI direct target genes (Groups 1 and 2) to all other S. meliloti transcriptome data sets, we also identified significant overlap (using hypergeometric probability tests, data not shown) with the transcriptomes for NCR247‐treatment (Penterman et al, ) and acid shock (Hellweg et al, ), and for podJ (Fields et al, ), cbrA (Gibson et al, ), ntrY (Calatrava‐Morales et al, ) and emrR mutants (Santos et al, ). Phenotypes common to these four mutants are increased exopolysaccharide production, decreased swimming motility and increased sensitivity to SDS and/or DOC.…”

Transcriptional regulation of ectoine catabolism in response to multiple metabolic and environmental cues

“…The NtrYX system (Fig. 9A) has been implicated in a number of biologically rather varied cellular processes in Alphaproteobacteria [for a recent description of the NtrYX system and additional references see (Fernandez et al, 2017)], notably also in the metabolism and assimilation of nitrogen-containing compounds (Pawlowski et al, 1991;Carrica Mdel et al, 2012;Cheng et al, 2014;Bonato et al, 2016;Calatrava-Morales et al, 2017). Our data now subscribe a function to the NtrYX system of R. pomeroyi with respect to the catabolism of ectoine when it is used as sole carbon source while still allowing its use as sole nitrogen source (Fig.…”

2‐Tridecanone impacts surface‐associated bacterial behaviours and hinders plant–bacteria interactions

“…qPCR was performed on an iCycler iQ5 (Bio‐Rad, Hercules, CA) according to Calatrava‐Morales et al . (). The primer sequences for qPCR are listed in Supporting Information Table S6.…”

“…RNA was also extracted from cells grown for 7 h on the surface of semisolid MM as described previously (Nogales et al, 2010) in the presence of volatile 2-TDC (1 mmol) or ethanol (control). RNA extraction, cDNA synthesis from 10 lg of total RNA, Cy3-and Cy5-labelling, hybridization to Sm14kOLI microarrays, image acquisition and data analysis were performed as previously reported (Calatrava-Morales et al, 2017). Normalization and t-statistics were carried out using the EMMA 2.8.2 microarray data analysis software (Dondrup et al, 2009).…”

“…About 1 lg of RNA was reversely transcribed using Superscript II reverse transcriptase (Invitrogen) and random hexamers (Roche) as primers. qPCR was performed on an iCycler iQ5 (Bio-Rad, Hercules, CA) according to Calatrava-Morales et al (2017). The primer sequences for qPCR are listed in Supporting Information Table S6.…”