Abstract
Brassinosteroid (BR) signaling has long been reported to have an effect on xylem development, but the detailed mechanism remains unclear, especially in tree species. In this study, we find PdC3H17, which was demonstrated to mediate xylem formation driven by auxin in our previous report, is also involved in BR-promoted xylem development. Y1H analysis, EMSA, and transcription activation assay confirmed that PdC3H17 was directly targeted by PdBES1, which is a key transcriptional regulator in BR signaling. Tissue specificity expression analysis and in situ assay revealed that PdC3H17 had an overlapping expression profile with PdBES1. Hormone treatment examinations verified that xylem phenotypes in PdC3H17 transgenic plants, which were readily apparent in normal condition, were attenuated by treatment with either brassinolide or the BR biosynthesis inhibitor propiconazole. The subsequent quantitative real-time polymerase chain reaction (qRT-PCR) analyses further revealed that BR converged with PdC3H17 to influence transcription of downstream xylem-related genes. Additionally, the enhancement of xylem differentiation by auxin in PdC3H17 overexpression plants was significantly attenuated compared with wild-type and dominant negative plants due to BR deficiency, which suggested that the BR- and auxin-responsive gene PdC3H17 acted as an mediation of these two hormones to facilitate xylem development. Taken together, our results demonstrate that BR signaling converges with auxin-mediated PdC3H17 to regulate xylem formation in Populus and thus provide insight into the regulation mechanism of BRs and the crosstalk with auxin signaling on xylem formation.
Highlights
Xylem is a highly specialized vascular tissue with the roles of transporting water and minerals and providing mechanical support for upright growth
In order to determine whether PdC3H17 could be a target of PdBES1, we first conducted PdC3H17 promoter sequence analysis; the result showed that multiple E-box elements (CANNTG), which are putative BES1 binding site, were found within -500 bp relative to the transcription start site (Supplementary Figure S1)
To further confirm this observation, we purified PdBES1 protein fused with MBP tag from E. coli and conducted EMSA analysis in vitro
Summary
Xylem is a highly specialized vascular tissue with the roles of transporting water and minerals and providing mechanical support for upright growth. As a key component in BR signaling, BES1 was revealed to function as a downstream target of GSK3s during xylem differentiation in a TDIF-dependent manner (Kondo et al, 2014), which indicated that BES1 was a shared common signaling component between TDIF signaling pathway and the BR signaling pathway in xylem development (Ryu et al, 2010; Guo et al, 2013; Kondo et al, 2014) Another similar result was obtained using VISUAL (Vascular cell Induction culture System Using Arabidopsis Leaves) system in Arabidopsis, indicating that BES1 and its homolog BZR1 redundantly promoted both phloem and xylem differentiation (Saito et al, 2018). These studies suggest that BR signaling plays an important role in secondary growth and wood formation
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