Genome-Wide Analysis of Anthocyanin Biosynthesis Regulatory WD40 Gene FcTTG1 and Related Family in Ficus carica L.

Abstract

WD40 proteins serve as crucial regulators in a broad spectrum of plant developmental and physiological processes, including anthocyanin biosynthesis. However, in fig (Ficus carica L.), neither the WD40 family nor any member involved in anthocyanin biosynthesis has been elucidated. In the present study, 204 WD40 genes were identified from the fig genome and phylogenetically classified into 5 clusters and 12 subfamilies. Bioinformatics analysis prediction localized 109, 69, and 26 FcWD40 proteins to the cytoplasm, nucleus and other cellular compartments, respectively. RNA-seq data mining revealed 127 FcWD40s expressed at FPKM > 10 in fig fruit. Most of these genes demonstrated higher expression in the early stages of fruit development. FcWD40-97 was recruited according to three criteria: high expression in fig fruit, predicted nuclear localization, and closest clustering with TTG1s identified in other plants. FcWD40-97, encoding 339 amino acids including 5 WD-repeat motifs, showed 88.01 and 87.94% amino acid sequence similarity to apple and peach TTG1, respectively. The gene is located on fig chromosome 4, and is composed of 1 intron and 2 exons. Promoter analysis revealed multiple light-responsive elements, one salicylic acid-responsive element, three methyl jasmonate-responsive elements, and one MYB-binding site involved in flavonoid biosynthesis gene regulation. FcWD40-97 was in the FPKM > 100 expression level group in fig fruit, and higher expression was consistently found in the peel compared to the flesh at the same development stages. Expression level did not change significantly under light deprivation, whereas in leaves and roots, its expression was relatively low. Transient expression verified FcWD40-97’s localization to the nucleus. Yeast two-hybrid (Y2H) and biomolecular fluorescence complementation (BiFC) assays revealed that FcWD40-97 interacts with FcMYB114, FcMYB123, and FcbHLH42 proteins in vitro and in vivo, showing that FcWD40-97 functions as a member of the MYB–bHLH–WD40 (MBW) complex in anthocyanin-biosynthesis regulation in fig. We therefore renamed FcWD40-97 as FcTTG1. Our results provide the first systematic analysis of the FcWD40 family and identification of FcTTG1 in fig pigmentation.

The MYB transcription factor RcMYB1 plays a central role in rose anthocyanin biosynthesis

“…The WD40 protein interaction with MYB protein has also been reported in recent studies. For example, in woodland strawberry, FcTTG1 can interact with FcMYB114, FcMYB123, and FcbHLH42 [ 60 ]; in eggplant SmMYB86 interacts with SmTTG1 [ 61 ]; in Medicago truncatula MtWD40–1 interacts with MtPAR or MtLAP1 [ 62 ]; and in kiwifruit AcWDR1 interacts with AcMYBF110 [ 48 ].…”

Comprehensive analysis of the Spartina alterniflora WD40 gene family reveals the regulatory role of SaTTG1 in plant development

“…TRANSPARENT TESTA GLABRA 1 ( TTG1 ), an initially identified WD40 protein in Arabidopsis , serves as a component of the MYB-bHLH-WD40 complex, regulating anthocyanin biosynthesis in plants. This complex has been observed in diverse species such as rice ( Sun et al., 2022 ), blueberry fruits ( Zhao et al., 2019 ), Ficus carica ( Fan et al., 2022 ), Punica granatum ( Ben-Simhon et al., 2011 ), and Camellia sinensis ( Liu et al., 2018 ). Furthermore, the overexpression of TTG-like gene CsWD40 in tobacco significantly increased anthocyanin content in the transgenic plant’s petals ( Liu et al., 2018 ).…”

Genome-wide analysis of WD40 protein family and functional characterization of BvWD40-82 in sugar beet

“…For example, the reference plant Arabidopsis has 230 WD40 proteins (Li et al, 2014). Among the cash crops reported, there were 225 WD40 proteins in red sage (Salvia miltiorrhiza) (Liu et al, 2020), 187 WD40 proteins in Rosaceae (Rosa chinensis 'old blush') (Sun et al, 2020), 42 WD40 proteins in walnut (Juglans regia), and 204 WD40s in fig (Ficus carica) (Chen et al, 2022a;Fan et al, 2022). Furthermore, among the food crops, wheat (Triticum aestivum) has 743 WD40 proteins (Hu et al, 2018), potato (Solanum tuberosum) has 168 WD40 proteins (Tao et al, 2019), and rice (Oryza sativa) has 200 WD40 proteins (Ouyang et al, 2012).…”