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TAL effectors and activation of predicted host targets distinguish Asian from African strains of the rice pathogen Xanthomonas oryzae pv. oryzicola while strict conservation suggests universal importance of five TAL effectors.

Wilkins KE, Booher NJ, Wang L, Bogdanove AJ - Front Plant Sci (2015)

Bottom Line: By pathogen whole genome, single molecule, real-time sequencing and host RNA sequencing, we compared TAL effector content and rice transcriptional responses across 10 geographically diverse Xoc strains.Filtering with a classifier we developed previously decreases the number of predicted binding elements across the genome, suggesting that it may reduce false positives among upregulated genes.Applying this filter and eliminating genes for which upregulation did not strictly correlate with presence of the corresponding TAL effector, we generated testable numbers of candidate targets for four of the five strictly conserved TAL effectors.

View Article: PubMed Central - PubMed

Affiliation: Plant Pathology and Plant-Microbe Biology Section, School of Integrative Plant Science, Cornell University Ithaca, NY, USA ; Graduate Field of Computational Biology, Cornell University Ithaca, NY, USA.

ABSTRACT
Xanthomonas oryzae pv. oryzicola (Xoc) causes the increasingly important disease bacterial leaf streak of rice (BLS) in part by type III delivery of repeat-rich transcription activator-like (TAL) effectors to upregulate host susceptibility genes. By pathogen whole genome, single molecule, real-time sequencing and host RNA sequencing, we compared TAL effector content and rice transcriptional responses across 10 geographically diverse Xoc strains. TAL effector content is surprisingly conserved overall, yet distinguishes Asian from African isolates. Five TAL effectors are conserved across all strains. In a prior laboratory assay in rice cv. Nipponbare, only two contributed to virulence in strain BLS256 but the strict conservation indicates all five may be important, in different rice genotypes or in the field. Concatenated and aligned, TAL effector content across strains largely reflects relationships based on housekeeping genes, suggesting predominantly vertical transmission. Rice transcriptional responses did not reflect these relationships, and on average, only 28% of genes upregulated and 22% of genes downregulated by a strain are up- and down- regulated (respectively) by all strains. However, when only known TAL effector targets were considered, the relationships resembled those of the TAL effectors. Toward identifying new targets, we used the TAL effector-DNA recognition code to predict effector binding elements in promoters of genes upregulated by each strain, but found that for every strain, all upregulated genes had at least one. Filtering with a classifier we developed previously decreases the number of predicted binding elements across the genome, suggesting that it may reduce false positives among upregulated genes. Applying this filter and eliminating genes for which upregulation did not strictly correlate with presence of the corresponding TAL effector, we generated testable numbers of candidate targets for four of the five strictly conserved TAL effectors.

No MeSH data available.


Related in: MedlinePlus

TAL effector content of diverse Xoc strains in relation to BLS256, and shared upregulation of BLS256 TAL effector targets. Each column labeled in bold font represents a group of orthologous TAL effectors, identified by finding reciprocal best BLAST hits. Columns are labeled according to the BLS256 ortholog, or if absent, by the designation “non-BLS256 TAL,” numbered arbitrarily. In these columns, the number of BSRs by which each TAL effector in the group (by strain) differs from the reference TAL effector for that group is given. The reference is the BLS256 TAL effector if present and the TAL effector with the most conserved BSR sequence otherwise. Gray fill indicates at least one difference. An “X,” with black fill, indicates absence of an ortholog. “Ψ” indicates that the TAL effector sequence derives from a pseudogene. “–(AD)” indicates a C-terminus like that of BLS256 Tal2h, missing the activation domain. Columns labeled in regular font with a rice gene name (locus ID) show the fold upregulation of that gene by each strain, with black fill indicating genes that are not significantly upregulated. The genes are grouped by the BLS256 TAL effector that upregulates them, immediately to the right of the column for that TAL effector. Fold change values in red font mark instances in which the gene promoter has no predicted EBE that passes a machine learning filter for the BLS256 TAL effector ortholog present in the strain represented in that row.
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Figure 1: TAL effector content of diverse Xoc strains in relation to BLS256, and shared upregulation of BLS256 TAL effector targets. Each column labeled in bold font represents a group of orthologous TAL effectors, identified by finding reciprocal best BLAST hits. Columns are labeled according to the BLS256 ortholog, or if absent, by the designation “non-BLS256 TAL,” numbered arbitrarily. In these columns, the number of BSRs by which each TAL effector in the group (by strain) differs from the reference TAL effector for that group is given. The reference is the BLS256 TAL effector if present and the TAL effector with the most conserved BSR sequence otherwise. Gray fill indicates at least one difference. An “X,” with black fill, indicates absence of an ortholog. “Ψ” indicates that the TAL effector sequence derives from a pseudogene. “–(AD)” indicates a C-terminus like that of BLS256 Tal2h, missing the activation domain. Columns labeled in regular font with a rice gene name (locus ID) show the fold upregulation of that gene by each strain, with black fill indicating genes that are not significantly upregulated. The genes are grouped by the BLS256 TAL effector that upregulates them, immediately to the right of the column for that TAL effector. Fold change values in red font mark instances in which the gene promoter has no predicted EBE that passes a machine learning filter for the BLS256 TAL effector ortholog present in the strain represented in that row.

Mentions: To assess conservation of TAL effector sequences across strains, we grouped TAL effectors by apparent orthology (descent from the same ancestral gene with no duplication events; see Materials and Methods) (Figure 1). To ascertain potential variation in targeting specificity within groups, we then recorded the number of BSR differences among the TAL effectors in each group, using the BLS256 TAL effector in the group as a reference, if present, or the TAL effector that was conserved exactly in the most strains otherwise (Figure 1). For orthologs of the 13 BLS256 TAL effectors with one or more known targets (Cernadas et al., 2014), we attempted to infer function and targeting specificity by asking whether the ortholog is predicted to bind the promoter for each corresponding target, and whether each target was upregulated by the corresponding strain in our RNA-Seq experiment. Whenever a BLS256 TAL effector is perfectly conserved at the BSR level in another strain, that strain upregulated the known target(s) (Figure 1). For imperfect matches, results were mixed. With one exception discussed below, every BLS256 TAL effector ortholog with no more than six BSR differences has in the promoter of each corresponding BLS256 target gene a predicted EBE that passes a machine learning filter for predicting functionality (see Materials and Methods), and for 99% of these, the target is upregulated. The exception is the BLS256 Tal6 ortholog group. Neither Tal6 nor any of its orthologs has an EBE that passes the filter in the confirmed Tal6 target Os01g31220. The reason for this is not clear. Nonetheless, since target activation by BLS256 in all cases was shown to be TAL effector dependent, for any ortholog the presence of a predicted EBE and upregulation of the corresponding target is strong evidence of activation by that ortholog in each case. Where this is the case, the ortholog likely serves the same function as the BLS256 TAL effector, but we cannot exclude the possibility that non-identical orthologs target different genes distinct from the BLS256 TAL effector target(s).


TAL effectors and activation of predicted host targets distinguish Asian from African strains of the rice pathogen Xanthomonas oryzae pv. oryzicola while strict conservation suggests universal importance of five TAL effectors.

Wilkins KE, Booher NJ, Wang L, Bogdanove AJ - Front Plant Sci (2015)

TAL effector content of diverse Xoc strains in relation to BLS256, and shared upregulation of BLS256 TAL effector targets. Each column labeled in bold font represents a group of orthologous TAL effectors, identified by finding reciprocal best BLAST hits. Columns are labeled according to the BLS256 ortholog, or if absent, by the designation “non-BLS256 TAL,” numbered arbitrarily. In these columns, the number of BSRs by which each TAL effector in the group (by strain) differs from the reference TAL effector for that group is given. The reference is the BLS256 TAL effector if present and the TAL effector with the most conserved BSR sequence otherwise. Gray fill indicates at least one difference. An “X,” with black fill, indicates absence of an ortholog. “Ψ” indicates that the TAL effector sequence derives from a pseudogene. “–(AD)” indicates a C-terminus like that of BLS256 Tal2h, missing the activation domain. Columns labeled in regular font with a rice gene name (locus ID) show the fold upregulation of that gene by each strain, with black fill indicating genes that are not significantly upregulated. The genes are grouped by the BLS256 TAL effector that upregulates them, immediately to the right of the column for that TAL effector. Fold change values in red font mark instances in which the gene promoter has no predicted EBE that passes a machine learning filter for the BLS256 TAL effector ortholog present in the strain represented in that row.
© Copyright Policy
Related In: Results  -  Collection

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Figure 1: TAL effector content of diverse Xoc strains in relation to BLS256, and shared upregulation of BLS256 TAL effector targets. Each column labeled in bold font represents a group of orthologous TAL effectors, identified by finding reciprocal best BLAST hits. Columns are labeled according to the BLS256 ortholog, or if absent, by the designation “non-BLS256 TAL,” numbered arbitrarily. In these columns, the number of BSRs by which each TAL effector in the group (by strain) differs from the reference TAL effector for that group is given. The reference is the BLS256 TAL effector if present and the TAL effector with the most conserved BSR sequence otherwise. Gray fill indicates at least one difference. An “X,” with black fill, indicates absence of an ortholog. “Ψ” indicates that the TAL effector sequence derives from a pseudogene. “–(AD)” indicates a C-terminus like that of BLS256 Tal2h, missing the activation domain. Columns labeled in regular font with a rice gene name (locus ID) show the fold upregulation of that gene by each strain, with black fill indicating genes that are not significantly upregulated. The genes are grouped by the BLS256 TAL effector that upregulates them, immediately to the right of the column for that TAL effector. Fold change values in red font mark instances in which the gene promoter has no predicted EBE that passes a machine learning filter for the BLS256 TAL effector ortholog present in the strain represented in that row.
Mentions: To assess conservation of TAL effector sequences across strains, we grouped TAL effectors by apparent orthology (descent from the same ancestral gene with no duplication events; see Materials and Methods) (Figure 1). To ascertain potential variation in targeting specificity within groups, we then recorded the number of BSR differences among the TAL effectors in each group, using the BLS256 TAL effector in the group as a reference, if present, or the TAL effector that was conserved exactly in the most strains otherwise (Figure 1). For orthologs of the 13 BLS256 TAL effectors with one or more known targets (Cernadas et al., 2014), we attempted to infer function and targeting specificity by asking whether the ortholog is predicted to bind the promoter for each corresponding target, and whether each target was upregulated by the corresponding strain in our RNA-Seq experiment. Whenever a BLS256 TAL effector is perfectly conserved at the BSR level in another strain, that strain upregulated the known target(s) (Figure 1). For imperfect matches, results were mixed. With one exception discussed below, every BLS256 TAL effector ortholog with no more than six BSR differences has in the promoter of each corresponding BLS256 target gene a predicted EBE that passes a machine learning filter for predicting functionality (see Materials and Methods), and for 99% of these, the target is upregulated. The exception is the BLS256 Tal6 ortholog group. Neither Tal6 nor any of its orthologs has an EBE that passes the filter in the confirmed Tal6 target Os01g31220. The reason for this is not clear. Nonetheless, since target activation by BLS256 in all cases was shown to be TAL effector dependent, for any ortholog the presence of a predicted EBE and upregulation of the corresponding target is strong evidence of activation by that ortholog in each case. Where this is the case, the ortholog likely serves the same function as the BLS256 TAL effector, but we cannot exclude the possibility that non-identical orthologs target different genes distinct from the BLS256 TAL effector target(s).

Bottom Line: By pathogen whole genome, single molecule, real-time sequencing and host RNA sequencing, we compared TAL effector content and rice transcriptional responses across 10 geographically diverse Xoc strains.Filtering with a classifier we developed previously decreases the number of predicted binding elements across the genome, suggesting that it may reduce false positives among upregulated genes.Applying this filter and eliminating genes for which upregulation did not strictly correlate with presence of the corresponding TAL effector, we generated testable numbers of candidate targets for four of the five strictly conserved TAL effectors.

View Article: PubMed Central - PubMed

Affiliation: Plant Pathology and Plant-Microbe Biology Section, School of Integrative Plant Science, Cornell University Ithaca, NY, USA ; Graduate Field of Computational Biology, Cornell University Ithaca, NY, USA.

ABSTRACT
Xanthomonas oryzae pv. oryzicola (Xoc) causes the increasingly important disease bacterial leaf streak of rice (BLS) in part by type III delivery of repeat-rich transcription activator-like (TAL) effectors to upregulate host susceptibility genes. By pathogen whole genome, single molecule, real-time sequencing and host RNA sequencing, we compared TAL effector content and rice transcriptional responses across 10 geographically diverse Xoc strains. TAL effector content is surprisingly conserved overall, yet distinguishes Asian from African isolates. Five TAL effectors are conserved across all strains. In a prior laboratory assay in rice cv. Nipponbare, only two contributed to virulence in strain BLS256 but the strict conservation indicates all five may be important, in different rice genotypes or in the field. Concatenated and aligned, TAL effector content across strains largely reflects relationships based on housekeeping genes, suggesting predominantly vertical transmission. Rice transcriptional responses did not reflect these relationships, and on average, only 28% of genes upregulated and 22% of genes downregulated by a strain are up- and down- regulated (respectively) by all strains. However, when only known TAL effector targets were considered, the relationships resembled those of the TAL effectors. Toward identifying new targets, we used the TAL effector-DNA recognition code to predict effector binding elements in promoters of genes upregulated by each strain, but found that for every strain, all upregulated genes had at least one. Filtering with a classifier we developed previously decreases the number of predicted binding elements across the genome, suggesting that it may reduce false positives among upregulated genes. Applying this filter and eliminating genes for which upregulation did not strictly correlate with presence of the corresponding TAL effector, we generated testable numbers of candidate targets for four of the five strictly conserved TAL effectors.

No MeSH data available.


Related in: MedlinePlus