Transcriptome profiling reveals the mechanism of fruit navel development in melon (Cucumis melo L.)

Abstract

Background: Melon is an important horticultural crop cultivated extensively worldwide. The size of the fruit navel, the terminal region of melon fruits, significantly influences the appearance quality of the fruit. However, the regulatory factors and molecular mechanisms governing the fruit navel development remain poorly understood in melon.

Results: In this study, the regulators and mechanisms underlying fruit navel development were investigated through phenotypic analysis, RNA sequencing (RNA-seq) and RT-qPCR methods. The inbred line 'T03' and a big fruit navel mutant 'BFN' of melon were used as experimental materials. RNA-seq analysis identified 116 differentially expressed genes (DEGs), including 54 up-regulated and 62 down-regulated genes, in both the green bud (GB) and ovary at anthesis (OA) stages of the 'BFN' melon compared to the 'T03' melon. Functional enrichment analysis revealed that these 116 DEGs were significantly associated with "Sesquiterpenoid and triterpenoid biosynthesis", "Circadian rhythm-plant", "Galactose metabolism" and "Biosynthesis of various alkaloids" pathways. There were three (AP2/ERF, MYB and C2H2 types) and eight (AP2/ERF, MADS-box, homeobox domain and bZIP types) transcription factors presented in up-regulated and down-regulated DEGs, and their putative target genes were predicted. Based on KEGG and expression analyses, two terpene cyclase/mutase genes (MELO3 C001812 and MELO3 C004329) were identified as being involved in the "Sesquiterpenoid and triterpenoid biosynthesis" pathway, and their transcripts were significantly downregulated in all detected development stages (EGB, GB, GYB, YB and OA) of 'BFN' fruits compared with 'T03' fruits.

Conclusions: The findings of this study elucidate a fundamental regulatory mechanism underlying fruit navel formation, and identify two key negative regulators, MELO3C001812 and MELO3C004329, involved in the development of the fruit navel in melon.

Keywords: Expression analysis; Fruit navel development; Melon; Transcriptome analysis.

Fig. 1

Phenotypic traits of fruit navel in ‘T03’ and ‘BFN’ fruits. A Comparison of fruit navels in ‘T03’ and ‘BFN’ fruits at the early green bud stage, green bud stage, green-yellow bud stage, yellow bud stage, ovary at anthesis, fruit at 3 days after pollination (DAP) and 7 DAP, fruit at veraison, and mature fruit stages. White frames represent the fruit navel tissues used for RNA-sequencing. B Diameter of fruit navel in ‘T03’ and ‘BFN’ fruits. Error bars represent ± SD. Asterisks indicate significant differences in the diameter of fruit navel in ‘BFN’ fruits relative to ‘T03’ fruits (Student t-test: *, P < 0.05; **, P < 0.01)

Fig. 2

Transcriptomic characteristics of fruit navel in ‘T03’ and ‘BFN’ melon. A Principal component analysis (PCA) of GB-T03, GB-BFN, OA-T03 and OA-BFN samples. B, C Venn diagrams of significantly upregulated (B) and downregulated (C) genes in ‘BFN’ fruits compared with ‘T03’ fruits at GB and OA stages

Fig. 3

RT-qPCR validation of RNA-seq data. Three upregulated and five downregulated DEGs identified by RNA-seq were randomly selected for RT-qPCR analyses. The expression trends of the selected DEGs detected by RT-qPCR were consistent with those obtained from RNA-seq. Error bars represent ± SD. Asterisks indicate significant differences in gene expression levels compared with the GB-T03 sample (Student t-test: *, P < 0.05; **, P < 0.01)

Fig. 4

Expression pattern of 116 DEGs between ‘BFN’ and ‘T03’ fruit navels. A total of 54 and 62 genes were significantly upregulated and downregulated, respectively, in both the GB and OA phases of ‘BFN’ fruits compared with ‘T03’ fruits. The color blocks represent the relative expression levels of genes, with high and low expression shown in red and blue color, respectively

Fig. 5

Significantly enriched (P < 0.05) KEGG terms in upregulated (A), downregulated (B), and all 116 DEGs (C) in the fruit navel of ‘BFN’ fruits compared with ‘T03’ fruits

Fig. 6

Identification of key regulators involved in fruit navel formation. A Analysis of transcription factors in up-regulated and down-regulated DEGs. Orange and green color represent up-regulated and down-regulated transcription factors, respectively. B Promoter analyses of DEGs involved in the “Sesquiterpenoid and triterpenoid biosynthesis”, “Galactose metabolism” and “Circadian rhythm—plant” pathways

Fig. 7

Expression analyses of five putative enzyme-coding genes identified in the “Sesquiterpenoid and triterpenoid biosynthesis”, “Galactose metabolism” and “Circadian rhythm—plant” pathways. RT-qPCR assays were conducted at the early green bud (EGB), green bud (GB), green yellow bud (GYB), yellow bud (YB) and ovary at anthesis (OA) stages. Error bars represent ± SD. Letters above the columns indicate significant differences in transcript levels at P < 0.05 (Duncan’s test)