Transcriptome Analysis Highlights the Molecular Mechanisms Associated with the Senescence of Florets in Gladiolus

Abstract

Gladiolus (Gladiolus spp.) florets exhibit low ethylene sensitivity. Accordingly, the wilting of their tepals is an ethylene-independent process. Both trehalose and cycloheximide can extend the vase life of gladiolus florets. Floral senescence is probably regulated by programmed cell death. However, senescence-related genes have not been thoroughly investigated, except in ethylene-sensitive species. In this study, we analyzed the expression of senescence-associated genes by conducting transcriptome (RNA-seq) analysis. First, we examined the effects of 0.1 M trehalose (Tre), 300 μM cycloheximide (CHI), and 50 μM chloramphenicol (CAP) treatments on postharvest quality and senescence-related gene expression in gladiolus ‘Fujinoyuki’ cut florets. The Tre and CHI treatments extended the vase life of gladiolus florets by about 1 day, i.e., 30% of Cont. Tepals were sampled at 0 days (0d) and 2 days (2d) post-treatment. The RNA-seq analysis of floret tissues generated 81,136 unique sequences. Moreover, 2,892, 4,670, and 57 differentially expressed genes were identified from the 0d_Control (Cont) vs 2d_Cont, 2d_Cont vs 2d_CHI, and 2d_Cont vs 2d_Tre comparisons, respectively. Gene Ontology (GO) analysis suggested that cysteine-type endopeptidase activity was significantly higher for 2d_Cont than for 0d. Additionally, the 0d vs 2d_Cont comparison showed the cell wall-related GO terms were more enriched for 2d_Cont. The Kyoto Encyclopedia of Genes and Genomes analysis revealed an increase in the expression of sucrose synthesis-related genes in the 2d_Cont samples. Among the genes involved in starch and sucrose metabolism, the genes mediating cell wall degradation were more actively expressed in the 2d_Cont samples than in the 2d_CHI samples. The fragments per kilobase per million reads (FPKM) values were used to select candidate senescence-related gene families, including the cysteine protease, invertase, peroxidase, pectinesterase, and transcription factor (NAC [no apical meristem (NAM), Arabidopsis transcription activation factor (ATAF), and cup-shaped cotyledon (CUC)] and WRKY) families. The expression levels of transcription factor genes, including NAC 048, 68, 073 and WRKY 6, 11, 24, were validated by qPCR. The expression of these NAC and WRKY transcription factor genes was upregulated by CHI, suggesting their involvement in senescence or side reactions in gladiolus tepals. This study revealed several candidate genes and associated GO terms for senescence of cut florets, but further study is needed, especially on key genes, including transcription factors.