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Cellular responses during morphological transformation in Azospirillum brasilense and Its flcA knockout mutant.

Hou X, McMillan M, Coumans JV, Poljak A, Raftery MJ, Pereg L - PLoS ONE (2014)

Bottom Line: Comparison of 2-DE protein profiles of wild-type (Sp7) and a flcA deletion mutant (Sp7-flcAΔ) revealed a total of 33 differentially expressed 2-DE gel spots, with 22 of these spots confidently separated to allow protein identification.Analysis of these spots by liquid chromatography-tandem mass spectrometry (LC-MS/MS) and MASCOT database searching identified 48 proteins (≥10% emPAI in each spot).This work is a step forward in connecting the Azospirillum phenome and genome.

View Article: PubMed Central - PubMed

Affiliation: School of Science and Technology, University of New England, Armidale, New South Wales, Australia; Department of Microbiology and Immunology, Shanxi Medical University, Taiyuan, Shanxi, China.

ABSTRACT
FlcA is a response regulator controlling flocculation and the morphological transformation of Azospirillum cells from vegetative to cyst-like forms. To understand the cellular responses of Azospirillum to conditions that cause morphological transformation, proteins differentially expressed under flocculation conditions in A. brasilense Sp7 and its flcA knockout mutant were investigated. Comparison of 2-DE protein profiles of wild-type (Sp7) and a flcA deletion mutant (Sp7-flcAΔ) revealed a total of 33 differentially expressed 2-DE gel spots, with 22 of these spots confidently separated to allow protein identification. Analysis of these spots by liquid chromatography-tandem mass spectrometry (LC-MS/MS) and MASCOT database searching identified 48 proteins (≥10% emPAI in each spot). The functional characteristics of these proteins included carbon metabolism (beta-ketothiolase and citrate synthase), nitrogen metabolism (Glutamine synthetase and nitric oxide synthase), stress tolerance (superoxide dismutase, Alkyl hydroperoxidase and ATP-dependent Clp protease proteolytic subunit) and morphological transformation (transducer coupling protein). The observed differences between Sp7 wild-type and flcA- strains enhance our understanding of the morphological transformation process and help to explain previous phenotypical observations. This work is a step forward in connecting the Azospirillum phenome and genome.

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Related in: MedlinePlus

Plant-root binding abilities of A. brasilense strain.Colonization of wheat root by Sp7 (1 and 4), Sp7-flcAΔ (2 and 5) and Sp72001 (3 and 6). A. brasilense strains harbor the reporter plasmid pLA-lacZ, containing a constitutively expressed lacZ gene and were stained with X-gal. Sp7 (1, 4) has strong binding ability to wheat roots and can be found all over the root surface; Sp7-flcAΔ (2, 5) and Sp72001 (3, 6) lost the ability to bind to wheat roots and could only be found in lateral root emergence areas. Scale bars indicate 50 µm (group A) and 5 µm (group B) (Magnification ×100 in group A and ×1000 in group B).
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pone-0114435-g002: Plant-root binding abilities of A. brasilense strain.Colonization of wheat root by Sp7 (1 and 4), Sp7-flcAΔ (2 and 5) and Sp72001 (3 and 6). A. brasilense strains harbor the reporter plasmid pLA-lacZ, containing a constitutively expressed lacZ gene and were stained with X-gal. Sp7 (1, 4) has strong binding ability to wheat roots and can be found all over the root surface; Sp7-flcAΔ (2, 5) and Sp72001 (3, 6) lost the ability to bind to wheat roots and could only be found in lateral root emergence areas. Scale bars indicate 50 µm (group A) and 5 µm (group B) (Magnification ×100 in group A and ×1000 in group B).

Mentions: As shown previously for flcA− mutant Sp72001 [6], Sp7-flcAΔ was also impaired in wheat root-surface colonization (Fig. 2) and lacZ-labeled Sp7-flcAΔ cells were detected by X-gal staining only in the crevices of lateral root emergence sites (Fig. 2). This is in contrast to the wild-type strain Sp7, which intensively colonized the surface of wheat roots.


Cellular responses during morphological transformation in Azospirillum brasilense and Its flcA knockout mutant.

Hou X, McMillan M, Coumans JV, Poljak A, Raftery MJ, Pereg L - PLoS ONE (2014)

Plant-root binding abilities of A. brasilense strain.Colonization of wheat root by Sp7 (1 and 4), Sp7-flcAΔ (2 and 5) and Sp72001 (3 and 6). A. brasilense strains harbor the reporter plasmid pLA-lacZ, containing a constitutively expressed lacZ gene and were stained with X-gal. Sp7 (1, 4) has strong binding ability to wheat roots and can be found all over the root surface; Sp7-flcAΔ (2, 5) and Sp72001 (3, 6) lost the ability to bind to wheat roots and could only be found in lateral root emergence areas. Scale bars indicate 50 µm (group A) and 5 µm (group B) (Magnification ×100 in group A and ×1000 in group B).
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4264754&req=5

pone-0114435-g002: Plant-root binding abilities of A. brasilense strain.Colonization of wheat root by Sp7 (1 and 4), Sp7-flcAΔ (2 and 5) and Sp72001 (3 and 6). A. brasilense strains harbor the reporter plasmid pLA-lacZ, containing a constitutively expressed lacZ gene and were stained with X-gal. Sp7 (1, 4) has strong binding ability to wheat roots and can be found all over the root surface; Sp7-flcAΔ (2, 5) and Sp72001 (3, 6) lost the ability to bind to wheat roots and could only be found in lateral root emergence areas. Scale bars indicate 50 µm (group A) and 5 µm (group B) (Magnification ×100 in group A and ×1000 in group B).
Mentions: As shown previously for flcA− mutant Sp72001 [6], Sp7-flcAΔ was also impaired in wheat root-surface colonization (Fig. 2) and lacZ-labeled Sp7-flcAΔ cells were detected by X-gal staining only in the crevices of lateral root emergence sites (Fig. 2). This is in contrast to the wild-type strain Sp7, which intensively colonized the surface of wheat roots.

Bottom Line: Comparison of 2-DE protein profiles of wild-type (Sp7) and a flcA deletion mutant (Sp7-flcAΔ) revealed a total of 33 differentially expressed 2-DE gel spots, with 22 of these spots confidently separated to allow protein identification.Analysis of these spots by liquid chromatography-tandem mass spectrometry (LC-MS/MS) and MASCOT database searching identified 48 proteins (≥10% emPAI in each spot).This work is a step forward in connecting the Azospirillum phenome and genome.

View Article: PubMed Central - PubMed

Affiliation: School of Science and Technology, University of New England, Armidale, New South Wales, Australia; Department of Microbiology and Immunology, Shanxi Medical University, Taiyuan, Shanxi, China.

ABSTRACT
FlcA is a response regulator controlling flocculation and the morphological transformation of Azospirillum cells from vegetative to cyst-like forms. To understand the cellular responses of Azospirillum to conditions that cause morphological transformation, proteins differentially expressed under flocculation conditions in A. brasilense Sp7 and its flcA knockout mutant were investigated. Comparison of 2-DE protein profiles of wild-type (Sp7) and a flcA deletion mutant (Sp7-flcAΔ) revealed a total of 33 differentially expressed 2-DE gel spots, with 22 of these spots confidently separated to allow protein identification. Analysis of these spots by liquid chromatography-tandem mass spectrometry (LC-MS/MS) and MASCOT database searching identified 48 proteins (≥10% emPAI in each spot). The functional characteristics of these proteins included carbon metabolism (beta-ketothiolase and citrate synthase), nitrogen metabolism (Glutamine synthetase and nitric oxide synthase), stress tolerance (superoxide dismutase, Alkyl hydroperoxidase and ATP-dependent Clp protease proteolytic subunit) and morphological transformation (transducer coupling protein). The observed differences between Sp7 wild-type and flcA- strains enhance our understanding of the morphological transformation process and help to explain previous phenotypical observations. This work is a step forward in connecting the Azospirillum phenome and genome.

Show MeSH
Related in: MedlinePlus