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Comparative molecular evolution of trichoderma chitinases in response to mycoparasitic interactions.

Ihrmark K, Asmail N, Ubhayasekera W, Melin P, Stenlid J, Karlsson M - Evol. Bioinform. Online (2010)

Bottom Line: Two of them, chi18-13 and chi18-17, are members of the B1/B2 chitinase subgroup that have expanded significantly in paralog number in mycoparasitic Hypocrea atroviridis and H. virens.Differences in amino acid diversity/conservation patterns between different Trichoderma clades are observed.These observations show that Trichoderma chitinases chi18-13 and chi18-15 evolve in a manner consistent with rapid co-evolutionary interactions and identifies putative target regions involved in determining substrate-specificity.

View Article: PubMed Central - PubMed

Affiliation: Department of Forest Mycology and Pathology, Swedish University of Agricultural Sciences, Box 7026, S-75007, Uppsala, Sweden.

ABSTRACT
Certain species of the fungal genus Trichoderma are potent mycoparasites and are used for biological control of fungal diseases on agricultural crops. In Trichoderma, whole-genome sequencing reveal between 20 and 36 different genes encoding chitinases, hydrolytic enzymes that are involved in the mycoparasitic attack. Sequences of Trichoderma chitinase genes chi18-5, chi18-13, chi18-15 and chi18-17, which all exhibit specific expression during mycoparasitism-related conditions, were determined from up to 13 different taxa and studied with regard to their evolutionary patterns. Two of them, chi18-13 and chi18-17, are members of the B1/B2 chitinase subgroup that have expanded significantly in paralog number in mycoparasitic Hypocrea atroviridis and H. virens. Chi18-13 contains two codons that evolve under positive selection and seven groups of co-evolving sites. Chi18-15 displays a unique codon-usage and contains five codons that evolve under positive selection and three groups of co-evolving sites. Regions of high amino acid variability are preferentially localized to substrate- or product side of the catalytic clefts. Differences in amino acid diversity/conservation patterns between different Trichoderma clades are observed. These observations show that Trichoderma chitinases chi18-13 and chi18-15 evolve in a manner consistent with rapid co-evolutionary interactions and identifies putative target regions involved in determining substrate-specificity.

No MeSH data available.


Related in: MedlinePlus

Reverse conservation analysis of chi18-13 orthologs and paralogs. A) Amino acid diversity was estimated using Rate4Site, based on a Clustal X alignment of chi18-13 Trichoderma orthologs and paralogs, and plotted as W mean scores. The y-axis represents arbitrary units (not shown) while a horizontal line indicates a 0.5 standard deviation cutoff. The x-axis represents residue position, asterisks (*) indicate positions of catalytic residues, diamonds (⋄) indicate substrate-interacting residues, boxed P indicate residues under strong (Bayes factor ≥50) positive selection, P indicate residues under weak (Bayes factor 10–49) positive selection, boxed C interconnected by horizontal lines indicate co-evolving residue groups and vertical dashed lines indicate identical residues. The position of the signal peptide, two proline-rich repeat units and regions with high amino acid diversity successfully visualised by homology modelling are indicated (Ib–IVb). B) Comparison of separate reverse conservation analyses on chi18-13 orthologs from H. minutispora, H. parapilulifera, H. pilulifera and H. atroviridis (dotted line) and T. ghanense, H. jecorina, T. brevicompactum, H. citrina, H. schweinitzii, H. virens, T. tomentosum, H. lixii and strain CBS816.68 (solid line). Arrows indicate regions with different W mean score distribution, magnifications illustrate residue S score distribution of the selected region.
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f3-ebo-2010-001: Reverse conservation analysis of chi18-13 orthologs and paralogs. A) Amino acid diversity was estimated using Rate4Site, based on a Clustal X alignment of chi18-13 Trichoderma orthologs and paralogs, and plotted as W mean scores. The y-axis represents arbitrary units (not shown) while a horizontal line indicates a 0.5 standard deviation cutoff. The x-axis represents residue position, asterisks (*) indicate positions of catalytic residues, diamonds (⋄) indicate substrate-interacting residues, boxed P indicate residues under strong (Bayes factor ≥50) positive selection, P indicate residues under weak (Bayes factor 10–49) positive selection, boxed C interconnected by horizontal lines indicate co-evolving residue groups and vertical dashed lines indicate identical residues. The position of the signal peptide, two proline-rich repeat units and regions with high amino acid diversity successfully visualised by homology modelling are indicated (Ib–IVb). B) Comparison of separate reverse conservation analyses on chi18-13 orthologs from H. minutispora, H. parapilulifera, H. pilulifera and H. atroviridis (dotted line) and T. ghanense, H. jecorina, T. brevicompactum, H. citrina, H. schweinitzii, H. virens, T. tomentosum, H. lixii and strain CBS816.68 (solid line). Arrows indicate regions with different W mean score distribution, magnifications illustrate residue S score distribution of the selected region.

Mentions: Amplification products and full-length sequences for the H. schweinitzii and H. virens chi18-13 orthologs were obtained, as were partial sequences presumably lacking the eight C-terminal residues from an additional nine species (Fig. S3). The H. virens chi18-13 gene was sequenced in the current work because the ortholog from the genome sequence (protein ID 25421) lacked 70 C-terminal residues compared to translated chi18-13 orthologs from other Trichoderma species. Additional sequences from H. jecorina and H. atroviridis were retrieved from the genome sequences, together with two paralogous sequences, protein ID 79492 from H. atroviridis (originally cloned as Ech30)10,14 and 58102 from H. virens. Two short proline-rich repeat regions in the C-terminal part (reference pos. 320–337 and 373–380 in H. atroviridis 45585) were removed from all species before the analysis, due to the highly variable number of repeats between species. The two paralogous sequences, 79492 and 58102, were 70 C-terminal residues shorter than the orthologs. Phylogenetic analysis confirmed the orthologous and paralogous status of the selected sequences (Fig. S3). Amino acid diversity was distributed amongst eight regions with W mean scores above the 0.5 standard deviation threshold from the RCA analysis (Fig. 3A). Four regions (Ib, IIb, IIIb and IVb) were visualized by the homology model of H. jecorina chi18-13 (Fig. 2B). One of the non-mapped high W score regions was associated with the signal peptide cleavage site (Fig. 3A). The eight predicted residues important for catalysis and substrate-binding by homology modelling (Table S2) were located in conserved regions with low W scores (Fig. 3A). Predicted substrate-binding residues (cd02877 in CDD) were associated with regions Ib and IIb of low amino acid conservation levels (Fig. 3A). More specifically, substrate-binding residues S74, S76 and T77 (reference H. atroviridis) were located in region Ib, which forms the entrance to the catalytic cleft (Fig. 2B). Substrate-binding residues G119, A120 and V121 (reference H. atroviridis) were located in region IIb, which forms a loop that protrudes into the catalytic centre of chi18-13 (Fig. 2B). Regions IIIb and IVb were located on the surface but were not a part of the catalytic cleft (Fig. 2B).


Comparative molecular evolution of trichoderma chitinases in response to mycoparasitic interactions.

Ihrmark K, Asmail N, Ubhayasekera W, Melin P, Stenlid J, Karlsson M - Evol. Bioinform. Online (2010)

Reverse conservation analysis of chi18-13 orthologs and paralogs. A) Amino acid diversity was estimated using Rate4Site, based on a Clustal X alignment of chi18-13 Trichoderma orthologs and paralogs, and plotted as W mean scores. The y-axis represents arbitrary units (not shown) while a horizontal line indicates a 0.5 standard deviation cutoff. The x-axis represents residue position, asterisks (*) indicate positions of catalytic residues, diamonds (⋄) indicate substrate-interacting residues, boxed P indicate residues under strong (Bayes factor ≥50) positive selection, P indicate residues under weak (Bayes factor 10–49) positive selection, boxed C interconnected by horizontal lines indicate co-evolving residue groups and vertical dashed lines indicate identical residues. The position of the signal peptide, two proline-rich repeat units and regions with high amino acid diversity successfully visualised by homology modelling are indicated (Ib–IVb). B) Comparison of separate reverse conservation analyses on chi18-13 orthologs from H. minutispora, H. parapilulifera, H. pilulifera and H. atroviridis (dotted line) and T. ghanense, H. jecorina, T. brevicompactum, H. citrina, H. schweinitzii, H. virens, T. tomentosum, H. lixii and strain CBS816.68 (solid line). Arrows indicate regions with different W mean score distribution, magnifications illustrate residue S score distribution of the selected region.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC2865166&req=5

f3-ebo-2010-001: Reverse conservation analysis of chi18-13 orthologs and paralogs. A) Amino acid diversity was estimated using Rate4Site, based on a Clustal X alignment of chi18-13 Trichoderma orthologs and paralogs, and plotted as W mean scores. The y-axis represents arbitrary units (not shown) while a horizontal line indicates a 0.5 standard deviation cutoff. The x-axis represents residue position, asterisks (*) indicate positions of catalytic residues, diamonds (⋄) indicate substrate-interacting residues, boxed P indicate residues under strong (Bayes factor ≥50) positive selection, P indicate residues under weak (Bayes factor 10–49) positive selection, boxed C interconnected by horizontal lines indicate co-evolving residue groups and vertical dashed lines indicate identical residues. The position of the signal peptide, two proline-rich repeat units and regions with high amino acid diversity successfully visualised by homology modelling are indicated (Ib–IVb). B) Comparison of separate reverse conservation analyses on chi18-13 orthologs from H. minutispora, H. parapilulifera, H. pilulifera and H. atroviridis (dotted line) and T. ghanense, H. jecorina, T. brevicompactum, H. citrina, H. schweinitzii, H. virens, T. tomentosum, H. lixii and strain CBS816.68 (solid line). Arrows indicate regions with different W mean score distribution, magnifications illustrate residue S score distribution of the selected region.
Mentions: Amplification products and full-length sequences for the H. schweinitzii and H. virens chi18-13 orthologs were obtained, as were partial sequences presumably lacking the eight C-terminal residues from an additional nine species (Fig. S3). The H. virens chi18-13 gene was sequenced in the current work because the ortholog from the genome sequence (protein ID 25421) lacked 70 C-terminal residues compared to translated chi18-13 orthologs from other Trichoderma species. Additional sequences from H. jecorina and H. atroviridis were retrieved from the genome sequences, together with two paralogous sequences, protein ID 79492 from H. atroviridis (originally cloned as Ech30)10,14 and 58102 from H. virens. Two short proline-rich repeat regions in the C-terminal part (reference pos. 320–337 and 373–380 in H. atroviridis 45585) were removed from all species before the analysis, due to the highly variable number of repeats between species. The two paralogous sequences, 79492 and 58102, were 70 C-terminal residues shorter than the orthologs. Phylogenetic analysis confirmed the orthologous and paralogous status of the selected sequences (Fig. S3). Amino acid diversity was distributed amongst eight regions with W mean scores above the 0.5 standard deviation threshold from the RCA analysis (Fig. 3A). Four regions (Ib, IIb, IIIb and IVb) were visualized by the homology model of H. jecorina chi18-13 (Fig. 2B). One of the non-mapped high W score regions was associated with the signal peptide cleavage site (Fig. 3A). The eight predicted residues important for catalysis and substrate-binding by homology modelling (Table S2) were located in conserved regions with low W scores (Fig. 3A). Predicted substrate-binding residues (cd02877 in CDD) were associated with regions Ib and IIb of low amino acid conservation levels (Fig. 3A). More specifically, substrate-binding residues S74, S76 and T77 (reference H. atroviridis) were located in region Ib, which forms the entrance to the catalytic cleft (Fig. 2B). Substrate-binding residues G119, A120 and V121 (reference H. atroviridis) were located in region IIb, which forms a loop that protrudes into the catalytic centre of chi18-13 (Fig. 2B). Regions IIIb and IVb were located on the surface but were not a part of the catalytic cleft (Fig. 2B).

Bottom Line: Two of them, chi18-13 and chi18-17, are members of the B1/B2 chitinase subgroup that have expanded significantly in paralog number in mycoparasitic Hypocrea atroviridis and H. virens.Differences in amino acid diversity/conservation patterns between different Trichoderma clades are observed.These observations show that Trichoderma chitinases chi18-13 and chi18-15 evolve in a manner consistent with rapid co-evolutionary interactions and identifies putative target regions involved in determining substrate-specificity.

View Article: PubMed Central - PubMed

Affiliation: Department of Forest Mycology and Pathology, Swedish University of Agricultural Sciences, Box 7026, S-75007, Uppsala, Sweden.

ABSTRACT
Certain species of the fungal genus Trichoderma are potent mycoparasites and are used for biological control of fungal diseases on agricultural crops. In Trichoderma, whole-genome sequencing reveal between 20 and 36 different genes encoding chitinases, hydrolytic enzymes that are involved in the mycoparasitic attack. Sequences of Trichoderma chitinase genes chi18-5, chi18-13, chi18-15 and chi18-17, which all exhibit specific expression during mycoparasitism-related conditions, were determined from up to 13 different taxa and studied with regard to their evolutionary patterns. Two of them, chi18-13 and chi18-17, are members of the B1/B2 chitinase subgroup that have expanded significantly in paralog number in mycoparasitic Hypocrea atroviridis and H. virens. Chi18-13 contains two codons that evolve under positive selection and seven groups of co-evolving sites. Chi18-15 displays a unique codon-usage and contains five codons that evolve under positive selection and three groups of co-evolving sites. Regions of high amino acid variability are preferentially localized to substrate- or product side of the catalytic clefts. Differences in amino acid diversity/conservation patterns between different Trichoderma clades are observed. These observations show that Trichoderma chitinases chi18-13 and chi18-15 evolve in a manner consistent with rapid co-evolutionary interactions and identifies putative target regions involved in determining substrate-specificity.

No MeSH data available.


Related in: MedlinePlus