Abstract
Main conclusion
Extensive characterization of the poplar GH9 gene family provides new insights into GH9 function and evolution in woody species, and may drive novel progress for molecular breeding in trees.
Abstract
In higher plants, endo-β-1,4-glucanases (cellulases) belonging to the glycosyl hydrolase family 9 (GH9) have roles in cell wall synthesis, remodeling and degradation. To increase the understanding of the GH9 family in perennial woody species, we conducted an extensive characterization of the GH9 family in the model tree species, Populus. We characterized 25 putative GH9 members in Populus with three subclasses (A, B, and C), using structures and bioinformatic analysis. Phylogenetic analyses of 114 GH9s from plant (dicot, monocot, and conifer) and bacterial species (outgroup) demonstrated that plant GH9s are monophyletic with respect to bacteria GH9s. Three subclasses, A, B, and C, of plant GH9 are formed before the divergence of angiosperms and gymnosperms. Chromosomal localization and duplications of GH9s in the Populus genome showed that eight paralogous pairs remained in conserved positions on segmental duplicated blocks, suggesting duplication of chromosomal segments has contributed to the family expansion. By examining tissue-specific expression profiles for all 25 members, we found that GH9 members exhibited distinct but partially overlapping expression patterns, while certain members have higher transcript abundance in mature or developing xylem. Based on our understanding of intraspecific variation and linkage disequilibrium of two KORRIGANs (PtoKOR1 and PtoKOR2) in natural population of Populus tomentosa, two non-synonymous SNPs in PtoKOR1 associated with fiber width and holocellulose content were obtained. Characterizations of the poplar GH9 family provide new insights into GH9 function and evolution in woody species, and may drive novel progress for molecular breeding in trees.
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Abbreviations
- CBM:
-
Carbohydrate-binding module
- CSC:
-
Cellulose synthase complex
- DBH:
-
Diameter at the breast height
- ESTs:
-
Expressed sequence tags
- FDR:
-
False discovery rate
- GH9:
-
Glycosyl hydrolase family 9
- GLM:
-
General linear model
- INDELs:
-
Insertions/deletions
- KOR1 :
-
KORRIGAN1
- LD:
-
Linkage disequilibrium
- LGs:
-
Linkage groups
- MFA:
-
Minor allele frequencies
- MLM:
-
Mixed linear model
- MY:
-
Million years
- NW:
-
Northwestern
- NE:
-
Northeastern
- RT-PCR:
-
Reverse transcription PCR
- RT-qPCR:
-
Real-time quantitative PCR
- r 2 :
-
The squared correlation of allele frequencies
- S:
-
Southern subset
- SNP:
-
Single-nucleotide polymorphisms
- T:
-
Divergence time
- TM:
-
Transmembrane domain
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Acknowledgments
This work was supported by grants from the Forestry Public Benefit Research Program (No. 201304102), the State Key Basic Research Program of China (No. 2012CB114506), and the Project of the National Natural Science Foundation of China (No. 31170622, 30872042).
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Q. Du and L. Wang contributed equally to this work
Electronic supplementary material
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425_2015_2271_MOESM1_ESM.doc
Fig. S1 Molecular characterization of PtoKOR1 and PtoKOR2. a Nucleotide and deduced amino acid sequences of PtoKOR1 and PtoKOR2. Numbers on the left refer to the positions of nucleotides or amino acid residues. b Gene structures of PtoKOR1 and PtoKOR2 (DOC 183 kb)
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Fig. S2 Protein sequence alignment of PtoKOR1 and PtoKOR2 with other plant PtoGH9s. Numbers on the left are the positions of the amino acids in each protein, with gaps (dashes) included to maximize alignments. Identical and similar amino acids are shaded in red and blue, respectively. represents the residues essential for catalytic activity identified in other plant PtoGH9s which are also conserved in PtoKOR1 and PtoKOR2 (D163/165, H504 and E561); * indicates the eight predicted glycosylation sites. The predicted transmembrane domain is overlined; the conserved polarized targeting signals are boxed. At: Arabidopsis thaliana, Os: Oryza sativa, Sl: Solanum lycopersicum (DOC 111 kb)
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Fig. S3 The decay of linkage disequilibrium within PtoKOR1 (a) and PtoKOR2 (b) in the natural population. Pairwise correlations between SNPs are plotted against the physical distance between the SNPs in base pairs. The curves describe the nonlinear regressions of r 2 (Er2) onto the physical distance in base pairs (DOC 756 kb)
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Fig. S4 Significant pairwise linkage disequilibrium (r 2 > 0.75, P < 0.001) between SNP markers in PtoKOR1. Four significant common SNP blocks are shown on a schematic of PtoKOR1 (DOC 52 kb)
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Fig. S5 The haplotype effect and protein structures for PtoKOR1 containing two significant haplotypes (T-C-T-A and A-T-A-G). a Haplotype effects on haplotype 1 (T-C-T-A) and haplotype 2 (A-T-A-G) found in PtoKOR1 controlling fiber width in Populus tomentosa natural populations. b Three-dimensional (3D) protein structures for PtoKOR1 containing two significant haplotypes (T-C-T-A and A-T-A-G) were predicted using SWISS-MODEL (http://swissmodel.expasy.org). Four significant fold architecture changes between these two protein structures are indicated with an arrow (DOC 459 kb)
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Sequence data from this article has been deposited in the GenBank Data Library under the accession Nos. HQ331247–HQ331273, HQ331276–HQ331300, and HQ380298–HQ380376. The phenotypic and genotypic data for SNP-trait association studies were provided as additional files (See Supplementary Information files S1 and S2). File S1 The phenotype data used in the SNP-traits association analysis in Populus tomentosa association population. File S2 The genotype data for SNPs in PtoKOR1 used in the SNP-traits association analysis in Populus tomentosa association population (XLSX 257 kb)
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Table S1 Geographical and meteorological parameters of three climatic distributional regions of Populus tomentosa in this study (DOC 46 kb)
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Table S3 The substitution rate ratios of nonsynonymous (dN or Ka) versus synonymous (dS or Ks) mutations estimated for paralogous PtrGH9 proteins. Synonymous (dS and Ks) and non-synonymous (dN and Ka) were used as parameters of substitution rates (DOC 35 kb)
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Table S6 Coding region nucleotide1 and amino acid2 sequence pairwise comparisons (% similarity) between poplar GH9 genes (DOC 107 kb)
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Table S8 Summary of nucleotide polymorphisms at the PtoKOR1 and PtoKOR2 loci, respectively. Regions containing indels are excluded from the calculation 14 (DOC 80 kb)
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Table S10 Validation of significant associations in PtoKOR1 using three climatic regions in Populus tomentosa natural populations (DOC 33 kb)
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Du, Q., Wang, L., Yang, X. et al. Populus endo-β-1,4-glucanases gene family: genomic organization, phylogenetic analysis, expression profiles and association mapping. Planta 241, 1417–1434 (2015). https://doi.org/10.1007/s00425-015-2271-y
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DOI: https://doi.org/10.1007/s00425-015-2271-y