Abstract
Euscaphis konishii is an evergreen plant that is widely planted as an industrial crop in Southern China. It produces red fruits with abundant secondary metabolites, giving E. konishii high medicinal and ornamental value. Auxin signaling mediated by members of the AUXIN RESPONSE FACTOR (ARF) and auxin/indole-3-acetic acid (Aux/IAA) protein families plays important roles during plant growth and development. Aux/IAA and ARF genes have been described in many plants but have not yet been described in E. konishii. In this study, we identified 34 EkIAA and 29 EkARF proteins encoded by the E. konishii genome through database searching using HMMER. We also performed a bioinformatic characterization of EkIAA and EkARF genes, including their phylogenetic relationships, gene structures, chromosomal distribution, and cis-element analysis, as well as conserved motifs in the proteins. Our results suggest that EkIAA and EkARF genes have been relatively conserved over evolutionary history. Furthermore, we conducted expression and co-expression analyses of EkIAA and EkARF genes in leaves, branches, and fruits, which identified a subset of seven EkARF genes as potential regulators of triterpenoids and anthocyanin biosynthesis. RT-qPCR, yeast one-hybrid, and transient expression analyses showed that EkARF5.1 can directly interact with auxin response elements and regulate downstream gene expression. Our results may pave the way to elucidating the function of EkIAA and EkARF gene families in E. konishii, laying a foundation for further research on high-yielding industrial products and E. konishii breeding.
Highlights
Plant secondary metabolites are important to plant development, but are significant as nutritional resources for humans, as sources for color and odorants, and as potential materials for drug discovery (Huang et al, 2020a; Bing et al, 2021)
We numbered the E. konishii IAA and AUXIN RESPONSE FACTOR (ARF) genes based on their homologs in Arabidopsis (Remington et al, 2004; Okushima et al, 2005); the full list is provided in Supplementary Table S1, along with their gene IDs, their coding sequences, genomic DNA and protein sequences, the lengths of the coding and protein sequences, and the predicted molecular weights and isoelectric points of the proteins
In Arabidopsis, ARF5 affects meristem development (Dastidar et al, 2019), while we showed here that EkARF5 is highly expressed during fruit maturation and ripening, likely reflecting changes in the promoter region associated with E. konishii and suggesting that the EkARF family underwent subfunctionalization during its evolutionary history
Summary
Plant secondary metabolites are important to plant development, but are significant as nutritional resources for humans, as sources for color and odorants, and as potential materials for drug discovery (Huang et al, 2020a; Bing et al, 2021). ARF DBDs bind to auxin response elements (AuxREs) located in the promoter region of auxinresponsive genes to activate or repress their transcription, depending on the type of MR (Tiwari et al, 2003; Guilfoyle and Hagen, 2007). Aux/IAAs are degraded via the 26S proteasome, alleviating the repressive effect of Aux/ IAAs on ARFs (Gray et al, 2001; Winkler et al, 2017; Roosjen et al, 2018) Both Aux/IAA and ARF proteins are encoded by large gene families with distinct expression patterns and regulatory mechanisms, contributing to the complexity of auxin signaling (Luo et al, 2018)
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