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Transcriptome Profiling of Wild-Type and pga-Knockout Mutant Strains Reveal the Role of Exopolysaccharide in Aggregatibacter actinomycetemcomitans.

Shanmugam M, El Abbar F, Ramasubbu N - PLoS ONE (2015)

Bottom Line: Our study demonstrated that the absence of the pga locus affects the genes involved in peptidoglycan recycling, glycogen storage, and virulence.Further, using confocal microscopy and plating assays, we show that the viability of pga mutant strain is significantly reduced during biofilm growth.Thus, this study highlights the importance of pga genes and the exopolysaccharide in the virulence of A. actinomycetemcomitans.

View Article: PubMed Central - PubMed

Affiliation: Department of Oral Biology, Rutgers School of Dental Medicine, Newark, New Jersey, United States of America.

ABSTRACT
Exopolysaccharides have a diverse set of functions in most bacteria including a mechanistic role in protecting bacteria against environmental stresses. Among the many functions attributed to the exopolysaccharides, biofilm formation, antibiotic resistance, immune evasion and colonization have been studied most extensively. The exopolysaccharide produced by many Gram positive as well as Gram negative bacteria including the oral pathogen Aggregatibacter actinomycetemcomitans is the homopolymer of β(1,6)-linked N-acetylglucosamine. Recently, we reported that the PGA-deficient mutant of A. actinomycetemcomitans failed to colonize or induce bone resorption in a rat model of periodontal disease, and the colonization genes, apiA and aae, were significantly down regulated in the mutant strain. To understand the role of exopolysaccharide and the pga locus in the global expression of A. actinomycetemcomitans, we have used comparative transcriptome profiling to identify differentially expressed genes in the wild-type strain in relation to the PGA-deficient strain. Transcriptome analysis revealed that about 50% of the genes are differently expressed (P < 0.05 and fold change >1.5). Our study demonstrated that the absence of the pga locus affects the genes involved in peptidoglycan recycling, glycogen storage, and virulence. Further, using confocal microscopy and plating assays, we show that the viability of pga mutant strain is significantly reduced during biofilm growth. Thus, this study highlights the importance of pga genes and the exopolysaccharide in the virulence of A. actinomycetemcomitans.

No MeSH data available.


Related in: MedlinePlus

Pathway analysis plot.Number of up- and down-regulated genes in A. actinomycetemcomitans IDH781 strain. In this comparison, up-regulated genes in the the mutant strain EA1002 deficient in PGA synthesis are shown in light blue. Data are grouped according to the major role categories. Positive and negative numbers correspond to the up- and down-regulated genes, respectively. Statistical values for the various pathways are provided in S2 and S3 Tables.
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pone.0134285.g002: Pathway analysis plot.Number of up- and down-regulated genes in A. actinomycetemcomitans IDH781 strain. In this comparison, up-regulated genes in the the mutant strain EA1002 deficient in PGA synthesis are shown in light blue. Data are grouped according to the major role categories. Positive and negative numbers correspond to the up- and down-regulated genes, respectively. Statistical values for the various pathways are provided in S2 and S3 Tables.

Mentions: To gain a better perspective of the various pathways and genes that were enriched, the software KegArray (Version 1.2.3; [18]) was used for the pathway analysis and to group the differentially expressed genes according to function (Fig 2). Furthermore, gene enrichment analysis was also computed with the CLC Genomics GSEA tool [24] to obtain statistical values associated with each pathway. When necessary, we also utilized the Comparative GO analysis for further statistical parameters associated with each pathway [25]. Using such combined analyses, we observed that genes associated with amino sugar, fatty acid, pyruvate and starch/sucrose metabolism, as well as glycolysis, pentose phosphate pathway, peptidoglycan turnover, citrate cycle, and two-component system are all up-regulated in EA1002 to a significant extent (Fig 2; light blue bars). This suggests that strain EA1002 undergoes a substantial change in metabolism during biofilm growth. Particularly noteworthy change occurred with the ribosomal protein coding genes where almost all of these were down-regulated in EA1002 compared to IDH781.


Transcriptome Profiling of Wild-Type and pga-Knockout Mutant Strains Reveal the Role of Exopolysaccharide in Aggregatibacter actinomycetemcomitans.

Shanmugam M, El Abbar F, Ramasubbu N - PLoS ONE (2015)

Pathway analysis plot.Number of up- and down-regulated genes in A. actinomycetemcomitans IDH781 strain. In this comparison, up-regulated genes in the the mutant strain EA1002 deficient in PGA synthesis are shown in light blue. Data are grouped according to the major role categories. Positive and negative numbers correspond to the up- and down-regulated genes, respectively. Statistical values for the various pathways are provided in S2 and S3 Tables.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4519337&req=5

pone.0134285.g002: Pathway analysis plot.Number of up- and down-regulated genes in A. actinomycetemcomitans IDH781 strain. In this comparison, up-regulated genes in the the mutant strain EA1002 deficient in PGA synthesis are shown in light blue. Data are grouped according to the major role categories. Positive and negative numbers correspond to the up- and down-regulated genes, respectively. Statistical values for the various pathways are provided in S2 and S3 Tables.
Mentions: To gain a better perspective of the various pathways and genes that were enriched, the software KegArray (Version 1.2.3; [18]) was used for the pathway analysis and to group the differentially expressed genes according to function (Fig 2). Furthermore, gene enrichment analysis was also computed with the CLC Genomics GSEA tool [24] to obtain statistical values associated with each pathway. When necessary, we also utilized the Comparative GO analysis for further statistical parameters associated with each pathway [25]. Using such combined analyses, we observed that genes associated with amino sugar, fatty acid, pyruvate and starch/sucrose metabolism, as well as glycolysis, pentose phosphate pathway, peptidoglycan turnover, citrate cycle, and two-component system are all up-regulated in EA1002 to a significant extent (Fig 2; light blue bars). This suggests that strain EA1002 undergoes a substantial change in metabolism during biofilm growth. Particularly noteworthy change occurred with the ribosomal protein coding genes where almost all of these were down-regulated in EA1002 compared to IDH781.

Bottom Line: Our study demonstrated that the absence of the pga locus affects the genes involved in peptidoglycan recycling, glycogen storage, and virulence.Further, using confocal microscopy and plating assays, we show that the viability of pga mutant strain is significantly reduced during biofilm growth.Thus, this study highlights the importance of pga genes and the exopolysaccharide in the virulence of A. actinomycetemcomitans.

View Article: PubMed Central - PubMed

Affiliation: Department of Oral Biology, Rutgers School of Dental Medicine, Newark, New Jersey, United States of America.

ABSTRACT
Exopolysaccharides have a diverse set of functions in most bacteria including a mechanistic role in protecting bacteria against environmental stresses. Among the many functions attributed to the exopolysaccharides, biofilm formation, antibiotic resistance, immune evasion and colonization have been studied most extensively. The exopolysaccharide produced by many Gram positive as well as Gram negative bacteria including the oral pathogen Aggregatibacter actinomycetemcomitans is the homopolymer of β(1,6)-linked N-acetylglucosamine. Recently, we reported that the PGA-deficient mutant of A. actinomycetemcomitans failed to colonize or induce bone resorption in a rat model of periodontal disease, and the colonization genes, apiA and aae, were significantly down regulated in the mutant strain. To understand the role of exopolysaccharide and the pga locus in the global expression of A. actinomycetemcomitans, we have used comparative transcriptome profiling to identify differentially expressed genes in the wild-type strain in relation to the PGA-deficient strain. Transcriptome analysis revealed that about 50% of the genes are differently expressed (P < 0.05 and fold change >1.5). Our study demonstrated that the absence of the pga locus affects the genes involved in peptidoglycan recycling, glycogen storage, and virulence. Further, using confocal microscopy and plating assays, we show that the viability of pga mutant strain is significantly reduced during biofilm growth. Thus, this study highlights the importance of pga genes and the exopolysaccharide in the virulence of A. actinomycetemcomitans.

No MeSH data available.


Related in: MedlinePlus