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Identifying feasible metabolic routes in Mycobacterium smegmatis and possible alterations under diverse nutrient conditions.

Baloni P, Padiadpu J, Singh A, Gupta KR, Chandra N - BMC Microbiol. (2014)

Bottom Line: Next, we probe cellular metabolism with an array of alternative carbon and nitrogen sources and identify those that are utilized and favour growth as well as those that do not support growth.We obtain mechanistic insights that explain observed phenotypic behaviour by studying gene-expression profiles and pathways inferred from the genome sequence.Comparison of transcriptome and phenome analysis of Msm and Mtb provides a rationale for understanding commonalities in metabolic adaptability.

View Article: PubMed Central - PubMed

Affiliation: Molecular Biophysics Unit, IISc, Bangalore, 560012, India. baloni@mbu.iisc.ernet.in.

ABSTRACT

Background: Many studies on M. tuberculosis have emerged from using M. smegmatis MC(2)155 (Msm), since they share significant similarities and yet Msm is non-pathogenic and faster growing. Although several individual molecules have been studied from Msm, many questions remain open about its metabolism as a whole and its capability to be versatile. Adaptability and versatility are emergent properties of a system, warranting a molecular systems perspective to understand them.

Results: We identify feasible metabolic pathways in Msm in reference condition with transcriptome, phenotypic microarray, along with functional annotation of the genome. Together with transcriptome data, specific genes from a set of alternatives have been mapped onto different pathways. About 257 metabolic pathways can be considered to be feasible in Msm. Next, we probe cellular metabolism with an array of alternative carbon and nitrogen sources and identify those that are utilized and favour growth as well as those that do not support growth. In all, about 135 points in the entire metabolic map are probed. Analyzing growth patterns under these conditions, lead us to hypothesize different pathways that can become active in various conditions and possible alternate routes that may be induced, thus explaining the observed physiological adaptations.

Conclusions: The study provides the first detailed analysis of feasible pathways towards adaptability. We obtain mechanistic insights that explain observed phenotypic behaviour by studying gene-expression profiles and pathways inferred from the genome sequence. Comparison of transcriptome and phenome analysis of Msm and Mtb provides a rationale for understanding commonalities in metabolic adaptability.

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Representation of the feasible pathways identified from transcriptomics and phenotypic analysis. (a) Selected metabolic pathways with enzymes vertically ordered from bottom to top. They are colored in grey scale based on the extent of normalized gene expression. Multiple boxes present in a single layer depict the alternate genes encoding similar enzymes. (b) Representative pathways with the level of gene expression in reference condition (c) Schematic illustrating alternate enzymes being highly expressed in glycolysis, highlighted in orange (d) Multiple sequence alignment of tpi (triose phosphate isomerase) for Mtb and two Msm proteins. Sequence identity and conservation along with catalytic site residues (*) are shown.
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Fig3: Representation of the feasible pathways identified from transcriptomics and phenotypic analysis. (a) Selected metabolic pathways with enzymes vertically ordered from bottom to top. They are colored in grey scale based on the extent of normalized gene expression. Multiple boxes present in a single layer depict the alternate genes encoding similar enzymes. (b) Representative pathways with the level of gene expression in reference condition (c) Schematic illustrating alternate enzymes being highly expressed in glycolysis, highlighted in orange (d) Multiple sequence alignment of tpi (triose phosphate isomerase) for Mtb and two Msm proteins. Sequence identity and conservation along with catalytic site residues (*) are shown.

Mentions: In order to identify metabolic pathways active in Msm in log phase cultures in the reference condition, we map inferred enzyme abundances from gene expression values of individual genes, for all pathways in the organism listed in standard databases, KEGG and BioCyc [26,27]. For a pathway to be active, enzymes in it must be expressed in detectable quantities. Although gene-expression does not always directly correlate with protein abundances, transcription data is clearly suggestive of whether or not a protein is present in detectable quantities. Moderate correlation between expression levels and protein abundances has been reported for bacterial systems [25,28]. 338 pathways are identified for Msm that combines knowledge of experimentally known pathways from literature along with those inferred from genome sequence analysis. Genes corresponding to enzymes in expected pathways including central carbon metabolism, amino acid biosynthesis, purine and pyrimidine biosynthesis, fatty acid metabolism, mycolic acid biosynthesis are all expressed, as expected. Figure 3a and b shows gene expression pattern corresponding to enzymes in some pathways (data for all 338 pathways is given in Additional file 2), which illustrates that many pathways including those of central carbon metabolism, as expected, appear active owing to expression of all required genes. However expression levels vary from low to high, which is quite understandable owing to their individual biochemical properties. In all, 257 pathways can be considered to be active in the condition studied (for example, the first and second row in Figure 3b). There are about 14 pathways in which the genes show no expression (for example, last row in Figure 3b), and about 57 pathways where few genes in them are expressed whereas 75 pathways had most of the genes expressed. The latter have implications of ease of adaptability (discussed in a later section).Figure 3


Identifying feasible metabolic routes in Mycobacterium smegmatis and possible alterations under diverse nutrient conditions.

Baloni P, Padiadpu J, Singh A, Gupta KR, Chandra N - BMC Microbiol. (2014)

Representation of the feasible pathways identified from transcriptomics and phenotypic analysis. (a) Selected metabolic pathways with enzymes vertically ordered from bottom to top. They are colored in grey scale based on the extent of normalized gene expression. Multiple boxes present in a single layer depict the alternate genes encoding similar enzymes. (b) Representative pathways with the level of gene expression in reference condition (c) Schematic illustrating alternate enzymes being highly expressed in glycolysis, highlighted in orange (d) Multiple sequence alignment of tpi (triose phosphate isomerase) for Mtb and two Msm proteins. Sequence identity and conservation along with catalytic site residues (*) are shown.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4248442&req=5

Fig3: Representation of the feasible pathways identified from transcriptomics and phenotypic analysis. (a) Selected metabolic pathways with enzymes vertically ordered from bottom to top. They are colored in grey scale based on the extent of normalized gene expression. Multiple boxes present in a single layer depict the alternate genes encoding similar enzymes. (b) Representative pathways with the level of gene expression in reference condition (c) Schematic illustrating alternate enzymes being highly expressed in glycolysis, highlighted in orange (d) Multiple sequence alignment of tpi (triose phosphate isomerase) for Mtb and two Msm proteins. Sequence identity and conservation along with catalytic site residues (*) are shown.
Mentions: In order to identify metabolic pathways active in Msm in log phase cultures in the reference condition, we map inferred enzyme abundances from gene expression values of individual genes, for all pathways in the organism listed in standard databases, KEGG and BioCyc [26,27]. For a pathway to be active, enzymes in it must be expressed in detectable quantities. Although gene-expression does not always directly correlate with protein abundances, transcription data is clearly suggestive of whether or not a protein is present in detectable quantities. Moderate correlation between expression levels and protein abundances has been reported for bacterial systems [25,28]. 338 pathways are identified for Msm that combines knowledge of experimentally known pathways from literature along with those inferred from genome sequence analysis. Genes corresponding to enzymes in expected pathways including central carbon metabolism, amino acid biosynthesis, purine and pyrimidine biosynthesis, fatty acid metabolism, mycolic acid biosynthesis are all expressed, as expected. Figure 3a and b shows gene expression pattern corresponding to enzymes in some pathways (data for all 338 pathways is given in Additional file 2), which illustrates that many pathways including those of central carbon metabolism, as expected, appear active owing to expression of all required genes. However expression levels vary from low to high, which is quite understandable owing to their individual biochemical properties. In all, 257 pathways can be considered to be active in the condition studied (for example, the first and second row in Figure 3b). There are about 14 pathways in which the genes show no expression (for example, last row in Figure 3b), and about 57 pathways where few genes in them are expressed whereas 75 pathways had most of the genes expressed. The latter have implications of ease of adaptability (discussed in a later section).Figure 3

Bottom Line: Next, we probe cellular metabolism with an array of alternative carbon and nitrogen sources and identify those that are utilized and favour growth as well as those that do not support growth.We obtain mechanistic insights that explain observed phenotypic behaviour by studying gene-expression profiles and pathways inferred from the genome sequence.Comparison of transcriptome and phenome analysis of Msm and Mtb provides a rationale for understanding commonalities in metabolic adaptability.

View Article: PubMed Central - PubMed

Affiliation: Molecular Biophysics Unit, IISc, Bangalore, 560012, India. baloni@mbu.iisc.ernet.in.

ABSTRACT

Background: Many studies on M. tuberculosis have emerged from using M. smegmatis MC(2)155 (Msm), since they share significant similarities and yet Msm is non-pathogenic and faster growing. Although several individual molecules have been studied from Msm, many questions remain open about its metabolism as a whole and its capability to be versatile. Adaptability and versatility are emergent properties of a system, warranting a molecular systems perspective to understand them.

Results: We identify feasible metabolic pathways in Msm in reference condition with transcriptome, phenotypic microarray, along with functional annotation of the genome. Together with transcriptome data, specific genes from a set of alternatives have been mapped onto different pathways. About 257 metabolic pathways can be considered to be feasible in Msm. Next, we probe cellular metabolism with an array of alternative carbon and nitrogen sources and identify those that are utilized and favour growth as well as those that do not support growth. In all, about 135 points in the entire metabolic map are probed. Analyzing growth patterns under these conditions, lead us to hypothesize different pathways that can become active in various conditions and possible alternate routes that may be induced, thus explaining the observed physiological adaptations.

Conclusions: The study provides the first detailed analysis of feasible pathways towards adaptability. We obtain mechanistic insights that explain observed phenotypic behaviour by studying gene-expression profiles and pathways inferred from the genome sequence. Comparison of transcriptome and phenome analysis of Msm and Mtb provides a rationale for understanding commonalities in metabolic adaptability.

Show MeSH
Related in: MedlinePlus