Small RNA sequencing analysis provides novel insights into microRNA-mediated regulation of defense responses in chickpea against Fusarium wilt infection.

Small RNA sequencing analysis in two chickpea genotypes, JG 62 (Fusarium wilt-susceptible) and WR 315 (Fusarium wilt-resistant), under Fusarium wilt stress led to identification of 544 miRNAs which included 406 known and 138 novel miRNAs. A total of 115 miRNAs showed differential expression in both the genotypes across different combinations. A miRNA, Car-miR398 targeted copper chaperone for superoxide dismutase (CCS) that, in turn, regulated superoxide dismutase (SOD) activity during chickpea-Foc interaction. Fusarium wilt (FW) of chickpea (Cicer arietinum L.) caused by Fusarium oxysporum f. sp. ciceris (Foc) is a destructive soil-borne disease that severely reduces the chickpea yield and quality globally. In the present study, we have investigated microRNAs and the microRNA/target gene crosstalk involved in chickpea resistance to FW. The control and stress samples from two genotypes, JG 62 (FW-susceptible) and WR 315 (FW-resistant), collected at 10 days post-inoculation (dpi), were selected for small RNA sequencing. A total of 12 libraries were constructed and sequenced using Illumina HiSeq 2500 platform. The sequencing and in silico analyses revealed the identification of 544 miRNAs which included 406 known and 138 novel miRNAs. A total of 50 miRNAs were physically co-localized with Foc-resistance QTLs present on chromosome 2 (also known as Foc hotspot). A total of 115 miRNAs showed differential expression in both the genotypes across different combinations. Prediction and functional annotation of miRNA targets revealed their role in transcription regulation, disease resistance, defense response, metabolism, etc. Ten miRNAs and their targets were validated using poly(A)-based qRT-PCR in two genotypes grown under lab and field conditions. Many miRNAs and their targets showed genotype-specific expression. The expression profiling also highlighted, both, similar and different expression patterns for the same sets of miRNA and mRNA at different stages of Foc infection. A high correlation in expression patterns of the miRNAs and their targets in lab- and field-grown plant samples was observed. Interestingly, Car-miR398 targeted copper chaperone for superoxide dismutase (CCS) that, in turn, regulated superoxide dismutase (SOD) activity during chickpea-Foc interaction. The cleavage site in targets was mapped for three miRNAs by analyzing publicly available degradome data for chickpea. The study, for the first time, provides novel insights into microRNA-mediated regulation of resistance and susceptibility mechanisms in chickpea against FW and opens up avenues for the development of the wilt-resistant cultivars in chickpea.

Priyadarshini P ，Kalwan G ，Kohli D ，Kumar D ，Bharadwaj C ，Gaikwad K ，Jain PK ... -  《-》

The transcriptional response to yellow and wilt disease, caused by race 6 of Fusarium oxysporum f. sp. Ciceris in two contrasting chickpea cultivars.

Chickpea (Cicer arietinum L.) ranks as the third most crucial grain legume worldwide. Fusarium wilt (Fusarium oxysporum f. sp. ciceri (Foc)) is a devastating fungal disease that prevents the maximum potential for chickpea production. To identify genes and pathways involved in resistance to race 6 of Foc, this study utilized transcriptome sequencing of two chickpea cultivars: resistant (Ana) and susceptible (Hashem) to Foc race 6. Illumina sequencing of the root samples yielded 133.5 million raw reads, with about 90% of the clean reads mapped to the chickpea reference genome. The analysis revealed that 548 genes (332 upregulated and 216 downregulated) in the resistant genotype (Ana) and 1115 genes (595 upregulated and 520 downregulated) in the susceptible genotype (Hashem) were differentially expressed under Fusarium wilt (FW) disease stress caused by Foc race 6. The expression patterns of some differentially expressed genes (DEGs) were validated using quantitative real-time PCR. A total of 131 genes were exclusively upregulated under FW stress in the resistant cultivar, including several genes involved in sensing (e.g., CaNLR-RPM1, CaLYK5-RLK, CaPR5-RLK, CaLRR-RLK, and CaRLP-EIX2), signaling (e.g., CaPP7, CaEPS1, CaSTY13, and CaPR-1), transcription regulation (e.g., CaMYBs, CaGLK, CaERFs, CaZAT11-like, and CaNAC6) and cell wall integrity (e.g., CaPGI2-like, CaEXLs, CaCSLD and CaCYP73A100-like). The achieved results could provide insights into the molecular mechanism underlying resistance to FW and could be valuable for breeding programs aimed at developing FW-resistant chickpea varieties.

Faramarzpour A ，Dezhsetan S ，Hassaneian Khoshro H ，Mirdar Mansuri R ，Pouralibaba HR ，Shobbar ZS ... -  《BMC GENOMICS》

Erratum: Eyestalk Ablation to Increase Ovarian Maturation in Mud Crabs.

《Jove-Journal of Visualized Experiments》

Chickpea defense against dual stresses of salt and Fusarium wilt is enhanced through selected bHLH transcription factors carrying the bHLH-MYC_N domain.

The plant transcriptome varies between combined stresses and single stresses, and is regulated differentially by transcription factors. Therefore, understanding the complexities of plant interactions with pathogens in stressed soils is always a challenge. In chickpea, 197 CabHLH genes were newly identified. Expression of 28 defense-associated CabHLHs [individual and combined stresses of Fusarium oxysporum f. sp. ciceris (Foc) and salt (NaCl) in three chickpea cultivars (JG-315: wilt resistant, JG-36: wilt tolerant, and JG-62: wilt susceptible) in Trichoderma asperellum T42 primed and non-primed conditions] revealed upregulation of most CabHLHs at 12 h post-stress in individual stresses but decreased significantly in the combined stress (Foc and salt). However, T42 priming stimulated the transcript accumulation of most CabHLHs even earlier (6 h). Three genes (CabHLH119, 158, and 184 carrying an additional domain bHLH-MYC_N) and two additional genes (CabHLH69 and 172) belonging to the subfamilies IIIde and IIIf were upregulated significantly in all three cultivars under individual and combined stresses, and upregulated further when primed with T42. Expression of the three bHLH-MYC_N domain containing genes, and defense activities (PAL, PO activities, phenylpropanoid accumulation) in the combined stress correlated very strongly. Protein-protein interactome studies further strengthened the claim that the three bHLH-MYC_N domain carrying CabHLHs, is likely to regulate the defense signaling in chickpea under stress as they could form complexes either directly or indirectly with cis-elements of promoters of some important defense genes. The results thus showed the significance of the IIIde and IIIf subfamily genes, particularly those carrying the bHLH-MYC_N domain, in mitigating combined stresses.

Rai N ，Rai SP ，Sarma BK 《-》

Discarded sequencing reads uncover natural variation in pest resistance in Thlaspi arvense.

Understanding the genomic basis of natural variation in plant pest resistance is an important goal in plant science, but it usually requires large and labor-intensive phenotyping experiments. Here, we explored the possibility that non-target reads from plant DNA sequencing can serve as phenotyping proxies for addressing such questions. We used data from a whole-genome and -epigenome sequencing study of 207 natural lines of field pennycress (Thlaspi arvense) that were grown in a common environment and spontaneously colonized by aphids, mildew, and other microbes. We found that the numbers of non-target reads assigned to the pest species differed between populations, had significant SNP-based heritability, and were associated with climate of origin and baseline glucosinolate contents. Specifically, pennycress lines from cold and thermally fluctuating habitats, presumably less favorable to aphids, showed higher aphid DNA load, i.e., decreased aphid resistance. Genome-wide association analyses identified genetic variants at known defense genes but also novel genomic regions associated with variation in aphid and mildew DNA load. Moreover, we found several differentially methylated regions associated with pathogen loads, in particular differential methylation at transposons and hypomethylation in the promoter of a gene involved in stomatal closure, likely induced by pathogens. Our study provides first insights into the defense mechanisms of Thlaspi arvense, a rising crop and model species, and demonstrates that non-target whole-genome sequencing reads, usually discarded, can be leveraged to estimate intensities of plant biotic interactions. With rapidly increasing numbers of large sequencing datasets worldwide, this approach should have broad application in fundamental and applied research.

Galanti D ，Jung JH ，Müller C ，Bossdorf O ... -  《eLife》