Genomic Analysis of the Glutathione S-Transferase Family in Pear (Pyrus communis) and Functional Identification of PcGST57 in Anthocyanin Accumulation

Anthocyanin accumulation in vacuoles results in red coloration in pear peels. Glutathione S-transferase (GST) proteins have emerged as important regulators of anthocyanin accumulation. Here, a total of 57 PcGST genes were identified in the European pear ‘Bartlett’ (Pyrus communis) through comprehensive genomic analysis. Phylogenetic analysis showed that PcGST genes were divided into 10 subfamilies. The gene structure, chromosomal localization, collinearity relationship, cis-elements in the promoter region, and conserved motifs of PcGST genes were analyzed. Further research indicated that glutamic acid (Glu) can significantly improve anthocyanin accumulation in pear peels. RNA sequencing (RNA-seq) analysis showed that Glu induced the expression of most PcGST genes, among which PcGST57 was most significantly induced. Further phylogenetic analysis indicated that PcGST57 was closely related to GST genes identified in other species, which were involved in anthocyanin accumulation. Transcript analysis indicated that PcGST57 was expressed in various tissues, other than flesh, and associated with peel coloration at different developmental stages. Silencing of PcGST57 by virus-induced gene silencing (VIGS) inhibited the expression of PcGST57 and reduced the anthocyanin content in pear fruit. In contrast, overexpression of PcGST57 improved anthocyanin accumulation. Collectively, our results demonstrated that PcGST57 was involved in anthocyanin accumulation in pear and provided candidate genes for red pear breeding.

A NAC Transcription Factor TuNAC69 Contributes to ANK-NLR-WRKY NLR-Mediated Stripe Rust Resistance in the Diploid Wheat Triticum urartu

Stripe rust is one of the most devastating diseases in wheat. Nucleotide-binding site (NBS) and leucine-rich repeat (LRR) domain receptors (NLRs) recognize pathogenic effectors and trigger plant immunity. We previously identified a unique NLR protein YrU1 in the diploid wheat Triticum urartu, which contains an N-terminal ANK domain and a C-terminal WRKY domain and confers disease resistance to stripe rust fungus Puccinia striiformis f. sp. Tritici (Pst). However, how YrU1 functions in disease resistance is not clear. In this study, through the RNA-seq analysis, we found that the expression of a NAC member TuNAC69 was significantly up-regulated after inoculation with Pst in the presence of YrU1. TuNAC69 was mainly localized in the nucleus and showed transcriptional activation in yeast. Knockdown TuNAC69 in diploid wheat Triticum urartu PI428309 that contains YrU1 by virus-induced gene silencing reduced the resistance to stripe rust. In addition, overexpression of TuNAC69 in Arabidopsis enhanced the resistance to powdery mildew Golovinomyces cichoracearum. In summary, our study indicates that TuNAC69 participates in the immune response mediated by NLR protein YrU1, and likely plays an important role in disease resistance to other pathogens.

Pepper Fruit Elongation Is Controlled by Capsicum annuum Ovate Family Protein 20

Fruit shape is one of the most important quality traits of pepper (Capsicum spp.) and is used as a major attribute for the classification of fruit types. Wide natural variation in fruit shape exists among the major cultivated species Capsicum annuum, allowing the identification of several QTLs controlling the trait. However, to date, no genes underlying fruit shape QTLs have been conclusively identified, nor has their function been verified in pepper. We constructed a mapping population from a cross of round- and elongated-fruited C. annuum parents and identified a single major QTL on chromosome 10, termed fs10, explaining 68 and 70% of the phenotypic variation for fruit shape index and for distal fruit end angle, respectively. The QTL was mapped in several generations and was localized to a 5 Mbp region containing the ortholog of SlOFP20 that suppresses fruit elongation in tomato. Virus-induced gene silencing of the pepper ortholog CaOFP20 resulted in increased fruit elongation on two independent backgrounds. Furthermore, CaOFP20 exhibited differential expression in fs10 near-isogenic lines, as well as in an association panel of elongated- and round-fruited accessions. A 42-bp deletion in the upstream region of CaOFP20 was most strongly associated with fruit shape variation within the locus. Histological observations in ovaries and fruit pericarps indicated that fs10 exerts its effect on fruit elongation by controlling cell expansion and replication. Our results indicate that CaOFP20 functions as a suppressor of fruit elongation in C. annuum and is the most likely candidate gene underlying fs10.

Chitinase Chi 2 Positively Regulates Cucumber Resistance against Fusarium oxysporum f. sp. cucumerinum

Cucumber (Cucumis sativus L.) is an important vegetable crop worldwide, and Fusarium wilt (FW), caused by Fusarium oxysporum f. sp. cucumerinum (Foc), severely restricts cucumber growth and yield. Accumulating lines of evidence indicate that chitinases play important roles in attacking the invading fungal pathogens through catalyzing their cell wall degradation. Here, we identified the chitinase (Chi) genes in cucumber and further screened the FW-responsive genes via a comparative transcriptome analysis and found that six common genes were predominantly expressed in roots but also significantly upregulated after Foc infection. Expression verification further conformed that Chi2 and Chi14 were obviously induced by Foc as well as by hormone treatments, compared with the controls. The purified Chi2 and Chi14 proteins significantly affected the growth of Foc in vitro, compared with the controls. Knockdown of Chi2 in cucumber by virus-induced gene silencing (VIGS) increased susceptibility to FW, compared with the Chi14-silenced and control plants, and silencing of Chi2 drastically impaired gene activation in the jasmonic acid pathway, suggesting that the Chi2 gene might play positive roles in cucumber FW defense and, therefore, can provide a gene resource for developing cucumber-FW-resistance breeding programs.

Silencing of a Wheat Ortholog of Glucan Synthase-Like Gene Reduced Resistance to Blumeria graminis f. sp. tritici

Wheat powdery mildew, caused by the obligate biotrophic ascomycete fungal pathogen Blumeria graminis f. sp. tritici (Bgt), is a major threat to wheat production worldwide. It is known that Arabidopsis thaliana glucan synthase-like 5 (AtGSL5) improves the resistance of wheat to powdery mildew by increasing its anti-penetration abilities. However, the function of glucan synthase-like (GSL) orthologs in crop species remains largely unknown. In this study, TaGSL22, a novel functional ortholog of AtGSL5, was isolated as the only Bgt-induced GSL gene in wheat. Phylogenetic analysis indicated that TaGSL22 was conserved within the group of Gramineae and showed a closer relationship to GSL orthologs from monocots than to those from dicots. The TaGSL22 transcript was highest in the wheat leaves, followed by stems then roots. TaGSL22 was localized in the cell membrane and cytoplasm of wheat protoplasts, as predicted by transmembrane structure analysis. In addition, expression of TaGSL22 was induced by the plant hormones ethylene (ETH) and salicylic acid (SA), but down-regulated by jasmonate (JA) and abscisic acid (ABA). The transcript level of TaGSL22 was up-regulated in the incompatible interaction between Bgt and wheat, whereas it remained relatively unchanged in the compatible interaction. Knocking down of TaGSL22 by virus-induced gene silencing (VIGS) induced a higher infection type in the wheat–Bgt interaction. The TaGSL22-silenced plants exhibited reduced resistance to Bgt, accompanied by decreased callose accumulation. Our study shows a conserved function of GSL genes in plant immunity associated with penetration resistance, and it indicates that TaGSL22 can be used to improve papilla composition and enhance resistance to wheat powdery mildew.

Hairiness Gene Regulated Multicellular, Non-Glandular Trichome Formation in Pepper Species

Trichomes are unicellular or multicellular epidermal structures that play a defensive role against environmental stresses. Although unicellular trichomes have been extensively studied as a mechanistic model, the genes involved in multicellular trichome formation are not well understood. In this study, we first classified the trichome morphology structures in Capsicum species using 280 diverse peppers. We cloned a key gene (Hairiness) on chromosome 10, which mainly controlled the formation of multicellular non-glandular trichomes (types II, III, and V). Hairiness encodes a Cys2-His2 zinc-finger protein, and virus-induced gene silencing of the gene resulted in a hairless phenotype. Differential expression of Hairiness between the hairiness and hairless lines was due to variations in promoter sequences. Transgenic experiments verified the hypothesis that the promoter of Hairiness in the hairless line had extremely low activity causing a hairless phenotype. Hair controlled the formation of type I glandular trichomes in tomatoes, which was due to nucleotide differences. Taken together, our findings suggest that the regulation of multicellular trichome formation might have similar pathways, but the gene could perform slightly different functions in crops.