Dataset Information

Identification, phylogeny, and transcript of chitinase family genes in sugarcane.

ABSTRACT: Chitinases are pathogensis-related proteins, which play an important role in plant defense mechanisms. The role of the sugarcane chitinase family genes remains unclear due to the highly heterozygous and aneuploidy chromosome genetic background of sugarcane. Ten differentially expressed chitinase genes (belonging to class I~VII) were obtained from RNA-seq analysis of both incompatible and compatible sugarcane genotypes during Sporisorium scitamineum challenge. Their structural properties and expression patterns were analyzed. Seven chitinases (ScChiI1, ScChiI2, ScChiI3, ScChiIII1, ScChiIII2, ScChiIV1 and ScChiVI1) showed more positive with early response and maintained increased transcripts in the incompatible interaction than those in the compatible one. Three (ScChiII1, ScChiV1 and ScChiVII1) seemed to have no significant difference in expression patterns between incompatible and compatible interactions. The ten chitinases were expressed differentially in response to hormone treatment as well as having distinct tissue specificity. ScChiI1, ScChiIV1 and ScChiVII1 were induced by various abiotic stresses (NaCl, CuCl2, PEG and 4?°C) and their involvement in plant immunity was demonstrated by over-expression in Nicotiana benthamiana. The results suggest that sugarcane chitinase family exhibit differential responses to biotic and abiotic stress, providing new insights into their function.

SUBMITTER: Su Y

PROVIDER: S-EPMC4451799 | biostudies-literature | 2015 Jun

REPOSITORIES: biostudies-literature

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Identification, phylogeny, and transcript of chitinase family genes in sugarcane.

Su Yachun Y Xu Liping L Wang Shanshan S Wang Zhuqing Z Yang Yuting Y Chen Yun Y Que Youxiong Y

Scientific reports 20150602

Chitinases are pathogensis-related proteins, which play an important role in plant defense mechanisms. The role of the sugarcane chitinase family genes remains unclear due to the highly heterozygous and aneuploidy chromosome genetic background of sugarcane. Ten differentially expressed chitinase genes (belonging to class I~VII) were obtained from RNA-seq analysis of both incompatible and compatible sugarcane genotypes during Sporisorium scitamineum challenge. Their structural properties and expr ...[more]

PMID: 26035173

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Project description:BackgroundInvertases (INVs) are key enzymes regulating sucrose metabolism and are here revealed to be involved in responses to environmental stress in plants. To date, individual members of the invertase gene family and their expression patterns are unknown in sugarcane due to its complex genome despite their significance in sucrose metabolism.ResultsIn this study, based on comparative genomics, eleven cDNA and twelve DNA sequences belonging to 14 non-redundant members of the invertase gene family were successfully cloned from sugarcane. A comprehensive analysis of the invertase gene family was carried out, including gene structures, phylogenetic relationships, functional domains, conserved motifs of proteins. The results revealed that the 14 invertase members from sugarcane could be clustered into three subfamilies, including 6 neutral/alkaline invertases (ShN/AINVs), and 8 acid invertases (ShAINVs). Faster divergence occurred in acid INVs than in neutral/alkaline INVs after the split of sugarcane and sorghum. At least a one-time gene duplication event was observed to have occurred in the four groups of acid INVs, whereas ShN/AINV1 and ShN/AINV2 in the β8 lineage were revealed to be the most recently duplicated genes among their paralogous genes in the β group of N/AINVs. Furthermore, comprehensive expression analysis of these genes was performed in sugarcane seedlings subjected to five abiotic stresses (drought, low temperature, glucose, fructose, and sucrose) using Quantitative Real-time PCR. The results suggested a functional divergence of INVs and their potential role in response to the five different treatments. Enzymatic activity in sugarcane seedlings was detected under five abiotic stresses treatments, and showed that the activities of all INVs were significantly inhibited in response to five different abiotic stresses, and that the neutral/alkaline INVs played a more prominent role in abiotic stresses than the acid INVs.ConclusionsIn this study, we determined the INV gene family members of sugarcane by PCR cloning using sorghum as a reference, providing the first study of the INV gene family in sugarcane. Combining existing INV gene data from 7 plants with a comparative approach including a series of comprehensive analyses to isolate and identify INV gene family members proved to be highly successful. Moreover, the expression levels of INV genes and the variation of enzymatic activities associated with drought, low temperature, glucose, fructose, and sucrose are reported in sugarcane for the first time. The results offered useful foundation and framework for future research for understanding the physiological roles of INVs for sucrose accumulation in sugarcane.

Project description:The oomycete Aphanomyces astaci is regarded as the causative agent of crayfish plague and represents an evident hazard for European crayfish species. Native crayfish populations infected with this pathogen suffer up to 100% mortality. The existence of multiple transmission paths necessitates the development of a reliable, robust and efficient test to detect the pathogen. Currently, A. astaci is diagnosed by a PCR-based assay that suffers from cross-reactivity to other species. We developed an alternative closed-tube assay for A. astaci, which achieves robustness through simultaneous amplification of multiple functionally constrained genes.Two novel constitutively expressed members of the glycosyl hydrolase (GH18) gene family of chitinases were isolated from the A. astaci strain Gb04. The primary amino acid sequence of these chitinase genes, termed CHI2 and CHI3, is composed of an N-terminal signal peptide directing the post-translational transport of the protein into the extracellular space, the catalytic GH18 domain, a proline-, serine-, and threonine-rich domain and a C-terminal cysteine-rich putative chitin-binding site. The A. astaci mycelium grown in a pepton-glucose medium showed significant temporal changes in steady-state CHI2 and CHI3 mRNA amounts indicating functional constraint. Their different temporal occurrence with maxima at 48 and 24 hours of incubation for CHI2 and CHI3, respectively, is in accordance with the multifunctionality of GH18 family members. To identify A. astaci-specific primer target sites in these novel genes, we determined the partial sequence homologs in the related oomycetes A. frigidophilus, A. invadans, A. helicoides, A. laevis, A. repetans, Achlya racemosa, Leptolegnia caudata, and Saprolegnia parasitica, as well as in the relevant fungi Fusarium solani and Trichosporon cutaneum. An A. astaci-specific primer pair targeting the novel genes CHI2 and CHI3 as well as CHI1 - a third GH18 family member - was multiplexed with primers targeting the 5.8S rRNA used as an endogenous control. A species was typed unambiguously as A. astaci if two peaks were concomitantly detected by melting curve analysis (MCA). For sensitive detection of the pathogen, but also for quantification of agent levels in susceptible crayfish and carrier crayfish, a TaqMan-probe based real-time PCR (qPCR) assay was developed. It targets the same chitinase genes and allows quantification down to 25 target sequences.The simultaneous qualitative detection of multiple sequences by qPCR/MCA represents a promising approach to detect species with elevated levels of genetic variation and/or limited available sequence information. The homogenous closed-tube format, reduced detection time, higher specificity, and the considerably reduced chance of false negative detection achieved by targeting multiple genes (CHI1, CHI2, CHI3, and the endogenous control) at least two of which are subject to high functional constraint, are the major advantages of this multiplex assay compared to other diagnostic methods. Sensitive quantification achieved with TaqMan qPCR facilitates to monitor infection status and pathogen distribution in different tissues and can help prevent disease transmission.

Project description:WRKY transcription factors (TFs) are essential players in different signaling cascades and regulatory networks involved in defense responses to various stressors. This study systematically analyzed and characterized WRKY family genes in the Saccharum spp. hybrid R570 and their expression in two sugarcane cultivars LCP85-384 (resistant to leaf scald) and ROC20 (susceptible to leaf scald) in response to bacterial pathogen infection and nitrogen implantation dosage. A total of 53 ShWRKY genes with 66 alleles were systematically identified in R570 based on the query sequence SsWRKY in S. spontaneum AP85-441. All ShRWKY alleles were further classified into four groups with 11 (16.7%) genes in group I, 36 (54.5%) genes in group II, 18 (27.3%) genes in group III, and 1 (1.5%) gene in group IV. Among them, 4 and 11 ShWRKY gene pairs displayed tandem and segmental duplication events, respectively. The ShWRKY genes exhibited conserved DNA-binding domains, which were accompanied by variations in introns, exons, and motifs. RT-qPCR analysis of two sugarcane cultivars triggered by Xanthomonas albilineans (Xa) revealed that four genes, ShWRKY13-2/39-1/49-3/125-3, exhibited significant upregulation in leaf scald-resistant LCP85-384. These WRKY genes were downregulated or unchanged in ROC20 at 24-72 h post-inoculation, suggesting that they play an important role in defense responses to Xa infection. Most of the 12 tested ShWRKYs, ShWRKY22-1/49-3/52-1 in particular, functioned as negative regulators in the two cultivars in response to a range of nitrogen (N) implantation doses. A total of 11 ShWRKY proteins were predicted to interact with each other. ShWRKY43 and ShWRKY49-3 are predicted to play core roles in the interaction network, as indicated by their interaction with six other ShWRKY proteins. Our results provide important candidate gene resources for the genetic improvement of sugarcane and lay the foundation for further functional characterization of ShWRKY genes in response to coupling effects of Xa infection and different N levels.

Dataset Information

Identification, phylogeny, and transcript of chitinase family genes in sugarcane.

Publications

Identification, phylogeny, and transcript of chitinase family genes in sugarcane.

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