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A novel bioinformatics pipeline to discover genes related to arbuscular mycorrhizal symbiosis based on their evolutionary conservation pattern among higher plants.

Favre P, Bapaume L, Bossolini E, Delorenzi M, Falquet L, Reinhardt D - BMC Plant Biol. (2014)

Bottom Line: However, genes that are members of functionally redundant gene families, or genes that have a vital function and therefore result in lethal mutant phenotypes, are difficult to identify.As a result we present a list of yet uncharacterized proteins that show a strongly AM-related pattern of sequence conservation, indicating that the respective genes may have been under selection for a function in AM.This strategy can be applied to diverse other biological phenomena if species with established genome sequences fall into distinguished groups that differ in a defined functional trait of interest.

View Article: PubMed Central - PubMed

ABSTRACT

Background: Genes involved in arbuscular mycorrhizal (AM) symbiosis have been identified primarily by mutant screens, followed by identification of the mutated genes (forward genetics). In addition, a number of AM-related genes has been identified by their AM-related expression patterns, and their function has subsequently been elucidated by knock-down or knock-out approaches (reverse genetics). However, genes that are members of functionally redundant gene families, or genes that have a vital function and therefore result in lethal mutant phenotypes, are difficult to identify. If such genes are constitutively expressed and therefore escape differential expression analyses, they remain elusive. The goal of this study was to systematically search for AM-related genes with a bioinformatics strategy that is insensitive to these problems. The central element of our approach is based on the fact that many AM-related genes are conserved only among AM-competent species.

Results: Our approach involves genome-wide comparisons at the proteome level of AM-competent host species with non-mycorrhizal species. Using a clustering method we first established orthologous/paralogous relationships and subsequently identified protein clusters that contain members only of the AM-competent species. Proteins of these clusters were then analyzed in an extended set of 16 plant species and ranked based on their relatedness among AM-competent monocot and dicot species, relative to non-mycorrhizal species. In addition, we combined the information on the protein-coding sequence with gene expression data and with promoter analysis. As a result we present a list of yet uncharacterized proteins that show a strongly AM-related pattern of sequence conservation, indicating that the respective genes may have been under selection for a function in AM. Among the top candidates are three genes that encode a small family of similar receptor-like kinases that are related to the S-locus receptor kinases involved in sporophytic self-incompatibility.

Conclusions: We present a new systematic strategy of gene discovery based on conservation of the protein-coding sequence that complements classical forward and reverse genetics. This strategy can be applied to diverse other biological phenomena if species with established genome sequences fall into distinguished groups that differ in a defined functional trait of interest.

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Phylogenetic analysis of an α-glucosidase with high conservation ratio. The first hit in the list of predicted AM-related proteins was an α-glucosidase (AES81209, Additional file 12: Table S4). The S. lycopersicum sequence was used to retrieve the closest homologues in a wide range of species (Additional file 1: File S1) for phylogenetic analysis. (a) Tree as in Figure 1 with the first hit per species identified at NCBI by blastp against non-redundant protein database using the tomato α-glucosidase (Solyc03g094020.2.1) as a query. (b) Tree with all homologues of Solyc03g094020.2.1 from tomato (S. lycopersicum), grape vine (V. vinifera), poplar (P. trichocarpa), M. truncatula, and A. thaliana. Note the AM-related branch (Gluc_1) that has no homologue from A. thaliana. The closest Arabidopsis homologue (At_Gluc_1) clusters far away.
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Fig8: Phylogenetic analysis of an α-glucosidase with high conservation ratio. The first hit in the list of predicted AM-related proteins was an α-glucosidase (AES81209, Additional file 12: Table S4). The S. lycopersicum sequence was used to retrieve the closest homologues in a wide range of species (Additional file 1: File S1) for phylogenetic analysis. (a) Tree as in Figure 1 with the first hit per species identified at NCBI by blastp against non-redundant protein database using the tomato α-glucosidase (Solyc03g094020.2.1) as a query. (b) Tree with all homologues of Solyc03g094020.2.1 from tomato (S. lycopersicum), grape vine (V. vinifera), poplar (P. trichocarpa), M. truncatula, and A. thaliana. Note the AM-related branch (Gluc_1) that has no homologue from A. thaliana. The closest Arabidopsis homologue (At_Gluc_1) clusters far away.

Mentions: The goal of this study was to identify AM-related genes based on the conservation pattern of their orthologues between AM-competent and non-mycorrhizal plant species. This approach is particularly targeted to identify AM-related genes that are not induced during symbiosis, hence, we focused on the proteins identified by Task3 that are not induced at the gene level (Additional file 14: Table S6). To evaluate the efficiency of the approach, this list was ordered according to the ratios of E-values between the averaged Brassicaceae and AM-competent dicot plants, respectively (Additional file 14: Table S6). The list was sorted in descending order, since the highest values for the conservation ratios indicate the proteins that are conserved to a higher degree among AM-competent species than between AM-competent and non-mycorrhizal species. In this list, the first protein, a predicted α-glucosidase/ xylosidase, was chosen to evaluate its conservation pattern in detail. Indeed, a phylogenetic tree prepared as in Figure 1 shows an extremely skewed pattern of conservation with a clearly resolved common branch of the AM-competent monocots and dicots, including the basal lineage Amborella trichopoda, whereas all the non-mycorrhizal species, the Brassicaceae, and B. vulgaris as a representative of the Chenopodiaceae, form an outlier group (Figure 8a).Figure 8


A novel bioinformatics pipeline to discover genes related to arbuscular mycorrhizal symbiosis based on their evolutionary conservation pattern among higher plants.

Favre P, Bapaume L, Bossolini E, Delorenzi M, Falquet L, Reinhardt D - BMC Plant Biol. (2014)

Phylogenetic analysis of an α-glucosidase with high conservation ratio. The first hit in the list of predicted AM-related proteins was an α-glucosidase (AES81209, Additional file 12: Table S4). The S. lycopersicum sequence was used to retrieve the closest homologues in a wide range of species (Additional file 1: File S1) for phylogenetic analysis. (a) Tree as in Figure 1 with the first hit per species identified at NCBI by blastp against non-redundant protein database using the tomato α-glucosidase (Solyc03g094020.2.1) as a query. (b) Tree with all homologues of Solyc03g094020.2.1 from tomato (S. lycopersicum), grape vine (V. vinifera), poplar (P. trichocarpa), M. truncatula, and A. thaliana. Note the AM-related branch (Gluc_1) that has no homologue from A. thaliana. The closest Arabidopsis homologue (At_Gluc_1) clusters far away.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4274732&req=5

Fig8: Phylogenetic analysis of an α-glucosidase with high conservation ratio. The first hit in the list of predicted AM-related proteins was an α-glucosidase (AES81209, Additional file 12: Table S4). The S. lycopersicum sequence was used to retrieve the closest homologues in a wide range of species (Additional file 1: File S1) for phylogenetic analysis. (a) Tree as in Figure 1 with the first hit per species identified at NCBI by blastp against non-redundant protein database using the tomato α-glucosidase (Solyc03g094020.2.1) as a query. (b) Tree with all homologues of Solyc03g094020.2.1 from tomato (S. lycopersicum), grape vine (V. vinifera), poplar (P. trichocarpa), M. truncatula, and A. thaliana. Note the AM-related branch (Gluc_1) that has no homologue from A. thaliana. The closest Arabidopsis homologue (At_Gluc_1) clusters far away.
Mentions: The goal of this study was to identify AM-related genes based on the conservation pattern of their orthologues between AM-competent and non-mycorrhizal plant species. This approach is particularly targeted to identify AM-related genes that are not induced during symbiosis, hence, we focused on the proteins identified by Task3 that are not induced at the gene level (Additional file 14: Table S6). To evaluate the efficiency of the approach, this list was ordered according to the ratios of E-values between the averaged Brassicaceae and AM-competent dicot plants, respectively (Additional file 14: Table S6). The list was sorted in descending order, since the highest values for the conservation ratios indicate the proteins that are conserved to a higher degree among AM-competent species than between AM-competent and non-mycorrhizal species. In this list, the first protein, a predicted α-glucosidase/ xylosidase, was chosen to evaluate its conservation pattern in detail. Indeed, a phylogenetic tree prepared as in Figure 1 shows an extremely skewed pattern of conservation with a clearly resolved common branch of the AM-competent monocots and dicots, including the basal lineage Amborella trichopoda, whereas all the non-mycorrhizal species, the Brassicaceae, and B. vulgaris as a representative of the Chenopodiaceae, form an outlier group (Figure 8a).Figure 8

Bottom Line: However, genes that are members of functionally redundant gene families, or genes that have a vital function and therefore result in lethal mutant phenotypes, are difficult to identify.As a result we present a list of yet uncharacterized proteins that show a strongly AM-related pattern of sequence conservation, indicating that the respective genes may have been under selection for a function in AM.This strategy can be applied to diverse other biological phenomena if species with established genome sequences fall into distinguished groups that differ in a defined functional trait of interest.

View Article: PubMed Central - PubMed

ABSTRACT

Background: Genes involved in arbuscular mycorrhizal (AM) symbiosis have been identified primarily by mutant screens, followed by identification of the mutated genes (forward genetics). In addition, a number of AM-related genes has been identified by their AM-related expression patterns, and their function has subsequently been elucidated by knock-down or knock-out approaches (reverse genetics). However, genes that are members of functionally redundant gene families, or genes that have a vital function and therefore result in lethal mutant phenotypes, are difficult to identify. If such genes are constitutively expressed and therefore escape differential expression analyses, they remain elusive. The goal of this study was to systematically search for AM-related genes with a bioinformatics strategy that is insensitive to these problems. The central element of our approach is based on the fact that many AM-related genes are conserved only among AM-competent species.

Results: Our approach involves genome-wide comparisons at the proteome level of AM-competent host species with non-mycorrhizal species. Using a clustering method we first established orthologous/paralogous relationships and subsequently identified protein clusters that contain members only of the AM-competent species. Proteins of these clusters were then analyzed in an extended set of 16 plant species and ranked based on their relatedness among AM-competent monocot and dicot species, relative to non-mycorrhizal species. In addition, we combined the information on the protein-coding sequence with gene expression data and with promoter analysis. As a result we present a list of yet uncharacterized proteins that show a strongly AM-related pattern of sequence conservation, indicating that the respective genes may have been under selection for a function in AM. Among the top candidates are three genes that encode a small family of similar receptor-like kinases that are related to the S-locus receptor kinases involved in sporophytic self-incompatibility.

Conclusions: We present a new systematic strategy of gene discovery based on conservation of the protein-coding sequence that complements classical forward and reverse genetics. This strategy can be applied to diverse other biological phenomena if species with established genome sequences fall into distinguished groups that differ in a defined functional trait of interest.

Show MeSH