ABSTRACT
Target leaf spot (TLS), which is caused by Corynespora cassiicola (C. cassiicola), is one of the most important diseases in cucumber (Cucumis sativus L.). Our previous research identified several C. cassiicola-responsive miRNAs in cucumber by high-throughput sequencing, including two known miRNAs and two novel miRNAs. The target genes of these miRNAs were related to secondary metabolism. In this study, we verified the interaction between these miRNAs and target genes by histochemical staining and fluorescence quantitative assays of GUS. We transiently expressed the candidate miRNAs and target genes in cucumber cotyledons to investigate the resistance to C. cassiicola. Transient expression of miR164d, miR396b, Novel-miR1, and Novel-miR7 in cucumber resulted in decreased resistance to C. cassiicola, while transient expression of NAC (inhibited by miR164d), APE (inhibited by miR396b), 4CL (inhibited by Novel-miR1), and PAL (inhibited by Novel-miR7) led to enhanced resistance to C. cassiicola. In addition, overexpression of 4CL and PAL downregulated lignin synthesis, and overexpression of Novel-miR1 and Novel-miR7 also downregulated lignin synthesis, indicating that the regulation of 4CL and PAL by Novel-miR1 and Novel-miR7 could affect lignin content. The tobacco rattle virus (TRV) induced short tandem target mimic (STTM)-miRNA silencing vector was successfully constructed, and target miRNAs were successfully silenced. The identification of disease resistance and lignin content showed that silencing candidate miRNAs could improve cucumber resistance to C. cassiicola.
ABSTRACT
Pathogen stress often significantly decreases cucumber production. However, knowledge regarding the molecular mechanism and signals of cucumber disease resistance is far from complete. Here, we report two translationally controlled tumor protein genes, CsTCTP1 and CsTCTP2, that are both negative modulators in the Cucumis sativus defense response to Sphaerotheca fuliginea. Subcellular localization analysis showed that CsTCTP1 and CsTCTP2 were both localized in the cytoplasm. Expression analysis indicated that the transcript levels of CsTCTP1 and CsTCTP2 were linked to the degree of cucumber resistance to S. fuliginea. Transient overexpression of either CsTCTP1 or CsTCTP2 in cucumber cotyledons impaired resistance to S. fuliginea, whereas silencing of either CsTCTP1 or CsTCTP2 enhanced cucumber resistance to S. fuliginea. The relationship of several defense-related genes and ABA and target of rapamycin (TOR) signaling pathway-related genes to the overexpressing and silencing of CsTCTP1/CsTCTP2 in non-infested cucumber plants was investigated. The results indicated that CsTCTP1 participates in the defense response to S. fuliginea by regulating the expression of certain defense-associated genes and/or ABA signaling pathway-associated genes, and CsTCTP2 participates through regulating the expression of TOR signaling pathway-associated genes. Our findings will guide enhancing the resistance of cucumber to powdery mildew.
