Expression of the nifA gene of Herbaspirillum seropedicae: role of the NtrC and NifA binding sites and of the −24/−12 promoter element

Souza, Emanuel Maltempi de; Pedrosa, Fábio O.; Rigo, L. U.; Machado, Hidevaldo B.; Yates, M. G.

doi:10.1099/00221287-146-6-1407

Cited by 38 publications

(26 citation statements)

References 40 publications

(24 reference statements)

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“…seropedicae strains SmR1 (Nif + , Sm R ) and SmR54 (Nif - , nifA ::Tn5-B21, Sm R , Km R ) [47] and Azospirillum(A . ) brasilense strains FP2 (Nif + , Sm R , Nal R , Tc R , Km R ) and FP10 (Nif - , nifA – , Sm R ,Nal R ) [48].…”

Section: Methodsmentioning

confidence: 99%

Metabolic profiling of two maize (Zea mays L.) inbred lines inoculated with the nitrogen fixing plant-interacting bacteria Herbaspirillum seropedicae and Azospirillum brasilense

et al. 2017

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Maize roots can be colonized by free-living atmospheric nitrogen (N2)-fixing bacteria (diazotrophs). However, the agronomic potential of non-symbiotic N2-fixation in such an economically important species as maize, has still not been fully exploited. A preliminary approach to improve our understanding of the mechanisms controlling the establishment of such N2-fixing associations has been developed, using two maize inbred lines exhibiting different physiological characteristics. The bacterial-plant interaction has been characterized by means of a metabolomic approach. Two established model strains of Nif+ diazotrophic bacteria, Herbaspirillum seropedicae and Azospirillum brasilense and their Nif- couterparts defficient in nitrogenase activity, were used to evaluate the impact of the bacterial inoculation and of N2 fixation on the root and leaf metabolic profiles. The two N2-fixing bacteria have been used to inoculate two genetically distant maize lines (FV252 and FV2), already characterized for their contrasting physiological properties. Using a well-controlled gnotobiotic experimental system that allows inoculation of maize plants with the two diazotrophs in a N-free medium, we demonstrated that both maize lines were efficiently colonized by the two bacterial species. We also showed that in the early stages of plant development, both bacterial strains were able to reduce acetylene, suggesting that they contain functional nitrogenase activity and are able to efficiently fix atmospheric N2 (Fix+). The metabolomic approach allowed the identification of metabolites in the two maize lines that were representative of the N2 fixing plant-bacterial interaction, these included mannitol and to a lesser extend trehalose and isocitrate. Whilst other metabolites such as asparagine, although only exhibiting a small increase in maize roots following bacterial infection, were specific for the two Fix+ bacterial strains, in comparison to their Fix- counterparts. Moreover, a number of metabolites exhibited a maize-genotype specific pattern of accumulation, suggesting that the highly diverse maize genetic resources could be further exploited in terms of beneficial plant-bacterial interactions for optimizing maize growth, with reduced N fertilization inputs.

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Section: Methodsmentioning

confidence: 99%

Metabolic profiling of two maize (Zea mays L.) inbred lines inoculated with the nitrogen fixing plant-interacting bacteria Herbaspirillum seropedicae and Azospirillum brasilense

et al. 2017

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“…Indicator plates for bacterial strains carrying gusA fusions contained 50 μg ml −1 5‐bromo‐4‐chloro‐3‐indolyl‐β‐ D ‐glucuronide (X‐gluc) [25]. H. seropedicae SMR1 [26] is a spontaneously streptomycin‐resistant derivative of the wild‐type strain Z78 [8] and was used to construct the mutant P nif :: gusA LR15. H. seropedicae IM40 is a Nif − mutant which carries a chromosomal nifH :: lacZ fusion [27].…”

Section: Methodsmentioning

confidence: 99%

Endophytic Herbaspirillum seropedicae expresses nif genes in gramineous plants

Roncato-Maccari

Ramos

Pedrosa

et al. 2003

FEMS Microbiology Ecology

138

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The interactions between maize, sorghum, wheat and rice plants and Herbaspirillum seropedicae were examined microscopically following inoculation with the H. seropedicae LR15 strain, a Nif(+) (Pnif::gusA) mutant obtained by the insertion of a gusA-kanamycin cassette into the nifH gene of the H. seropedicae wild-type strain. The expression of the Pnif::gusA fusion was followed during the association of the diazotroph with the gramineous species. Histochemical analysis of seedlings of maize, sorghum, wheat and rice grown in vermiculite showed that strain LR15 colonized root surfaces and inner tissues. In early steps of the endophytic association, H. seropedicae colonized root exudation sites, such as axils of secondary roots and intercellular spaces of the root cortex; it then occupied the vascular tissue and there expressed nif genes. The expression of nif genes occurred in roots, stems and leaves as detected by the GUS reporter system. The expression of nif genes was also observed in bacterial colonies located in the external mucilaginous root material, 8 days after inoculation. Moreover, the colonization of plant tissue by H. seropedicae did not depend on the nitrogen-fixing ability, since similar numbers of cells were isolated from roots or shoots of the plants inoculated with Nif(+) or Nif(-) strains.

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“…4), where the upstream regions are specially AϩT rich (Ϸ60%) with respect to coding regions (Ϸ47.5%). They also contain conserved Ϫ24͞Ϫ12 consensus sequences for 54 -dependent promoters (25) and two or three putative NifA-binding sites (26,27). Moreover, in the nifH upstream region, there are two more interesting features that stress the relevance of its regulatory function: an inverted repeat comprising the ATG translation start codon and a DNA region with high bending index between the two NifA-binding sites (data not shown).…”

Section: Ammonium Starvation-induced Nif Genes Under Aerobic Conditionsmentioning

confidence: 99%

Gene function analysis in environmental isolates: The nif regulon of the strict iron oxidizing bacterium Leptospirillum ferrooxidans

Parro

Moreno‐Paz

2003

Proc. Natl. Acad. Sci. U.S.A.

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A random genomic library from an environmental isolate of the Gram-negative bacterium Leptospirillum ferrooxidans has been printed on a microarray. Gene expression analysis was carried out with total RNA extracted from L. ferrooxidans cultures in the presence or absence of ammonium as nitrogen source under aerobic conditions. Although practically nothing is known about the genome sequence of this bacterium, this approach allowed us the selection and sequencing of only those clones bearing genes that showed an altered expression pattern. By sequence comparison, we have identified most of the genes of nitrogen fixation regulon in L. ferrooxidans, like the nifHDKENX operon, encoding the structural components of Mo-Fe nitrogenase; nifSU-hesBhscBA-fdx operon, for Fe-S cluster assembly; the amtB gene (ammonium transporter); modA (molybdenum ABC type transporter); some regulatory genes like ntrC, nifA (the specific activator of nif genes); or two glnB-like genes (encoding the PII regulatory protein). Our results show that shotgun DNA microarrays are very powerful tools to accomplish gene expression studies with environmental bacteria whose genome sequence is still unknown, avoiding the time and effort necessary for whole genome sequencing projects.shotgun DNA microarrays ͉ gene expression ͉ nitrogen fixation ͉ iron oxidizer G ene expression analysis by DNA microarrays (biochips) has enjoyed a tremendous increase during the last few years (1). As whole genome sequences are available at increasing rates, DNA microarrays are used for the functional analysis of the newly and already characterized genes. A previous knowledge of DNA sequence usually is a prerequisite for biochip construction from already known genes. It is possible to spot all ORFs of complete genomes from bacteria to humans. This strategy requires one to undertake expensive and time-consuming genome sequencing projects to study the function of just a particular set of genes. This is a genuinely tedious task if we want to apply this approach to those bacterial strains isolated from the environment. Here we use a shotgun DNA microarray strategy (2) for gene expression analysis in environmental isolates, bypassing the need for previous knowledge of the genome sequence.The Gram-negative bacterium Leptospirillum ferrooxidans is one of the most abundant bacteria in mineral-processing bioreactors, as well as in certain natural environments like the extremely acidic Tinto River (in the southwestern region of Spain), which has pH values between 0.8 and 2.4 and high metal concentrations along the entire 90 km of its length (3, 4). L. ferrooxidans is a strict iron-oxidizing acidophile considered one of the main responsible agents for maintaining the pH balance and hence the physicochemical properties of the ecosystem. This process is achieved by accelerating the limiting step for pyrite (FeS 2 ) leaching: the production of ferric iron (Fe 3ϩ ) (5, 6). L. ferrooxidans is also very important because of its capacity to extract heavy metals from minerals or contamina...

show abstract

Expression of the nifA gene of Herbaspirillum seropedicae: role of the NtrC and NifA binding sites and of the −24/−12 promoter element

Cited by 38 publications

References 40 publications

Metabolic profiling of two maize (Zea mays L.) inbred lines inoculated with the nitrogen fixing plant-interacting bacteria Herbaspirillum seropedicae and Azospirillum brasilense

Metabolic profiling of two maize (Zea mays L.) inbred lines inoculated with the nitrogen fixing plant-interacting bacteria Herbaspirillum seropedicae and Azospirillum brasilense

Endophytic Herbaspirillum seropedicae expresses nif genes in gramineous plants

Gene function analysis in environmental isolates: The nif regulon of the strict iron oxidizing bacterium Leptospirillum ferrooxidans

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