CmFUL1 was potentially involved in fruit elongation in melon

Abstract

Melon (Cucumis melo L.) is a fruit crop in the world; fruit size and fruit shape are major traits for melon quality. Fruit length is a crucial indicator affecting fruit size and shape, but few genes regulating this trait have been identified. Here, we identified the transcription factor CmFUL1 (FRUITFULL) as a candidate for regulating fruit length using genome-wide association analysis (GWAS) and phylogenetic analysis. CmFUL1 is mainly expressed during flower and ovary development by tissue-specific expression. Transcriptional analysis revealed that CmFUL1 expression levels exhibited a negative correlation with fruit length across diverse melon germplasm. Furthermore, functional characterization demonstrated that CmFUL1 acts as a negative regulator of fruit elongation, CR-Cmful1 mutants generated by CRISPR-Cas9 showing enhanced longitudinal fruits. This repressive role was evolutionarily conserved, as heterologous overexpression of CmFUL1 in tomato consistently inhibited fruit elongation. Collectively, the results suggested that CmFUL1 is a candidate gene involved in regulating fruit length in melon, and provided genetic resources for molecular breeding of melon.

Figure 1

Independent selection in fruit traits among the 1175 melon accessions. Ordinate axis indicates fruit traits. And abscissa axis indicates the chromosome 12. Horizontal lines indicate the genome-wide threshold of selection signals. P-value observations for these loci should be consistent with the Manhattan Plot values.

Figure 2

The expression pattern of candidate genes. The candidate gene on the region on the chromosome 12 from the Melon reference genome version 3.5.1.

Figure 3

Evolution and expression pattern of CmFUL1. (A) The phylogenetic tree of CmFUL1 and homologous genes in cucumber (Csa1p039910), Arabidopsis (AT5G60910), tobacco (Nitab4.50002449 g0010), tomato (Solyc03g114830), rice (Os03g54160), potato (PGSC0003DMC400042629), wax gourd (Bhi08M000888), watermelon (Cla97C04G078430), and +pumpkin (CmoCh13G01030, CmaCh13G010580). (B) The relative expression of CmFUL1 in tissues by RT-qPCR. CmACTIN1 was used as an internal control, and three biological replicates were used. (C) The gene structure of CmFUL1. (D) The K-domain of CmFUL1 orthologs in short melon (S), long melon (Long), tobacco (Nt), cucumber (Cs), Arabidopsis (At), and tomato (Solanum lycopersicum L.). (E) The protein structure of CmFUL1.

Figure 4

Correlation between fruit length and CmFUL1 expression. (A) The information of six accessions in the subspecies agrestis. (B) The fruit length of six accessions in the subspecies agrestis. (C) The expression level of CmFUL1 in the six species by qRT-PCR. CmACTIN1 was used as an internal control, and three biological replicates were used, error bars indicate the standard error of the mean. ^**P < 0.01.

Figure 5

The targets design, phenotype statistics of WT, and CR-Cmful1. (A) The gene structure and 16-bp deletion in the fourth exon of WT and CR-Cmful1, respectively. (B) Amino acids of CR-Cmful1 compared to WT. (C) The fruit phenotype of WT and CR-Cmful1. (D) The fruit length and width statistics of WT and CR-Cmful1. ^*** means P < 0.001, ns means no significant difference.

Figure 6

Functional validation of CmFUL1. (A) The fruit phenotype of CmFUL1 overexpression lines (CmFUL1-OE-1 and CmFUL1-OE-2). (B) qRT-PCR expression levels of CmFUL1 in CmFUL1-OE-1 and CmFUL1-OE-2 lines and MT. SlACTIN1 was used as an internal control, and three biological replicates were used. Error bars indicate the standard error of the mean. ^**P < 0.01, ^***P < 0.001. (C) The fruit length and fruit width of CmFUL1-OE lines and MT. The number of replicates was 11, 12, 15, and 11, 12, 14, respectively. Error bars indicate the standard error of the mean. ^***P < 0.001.