CELL CYCLE SEITCH 52 regulates tillering by interacting with LATERAL SUPPRESSOR in non-heading Chinese cabbage
Introduction
Tillering contributes greatly to plant morphogenesis [1] and plays a decisive role in crop yield, including rice [2], wheat [3], pea [4], and sorghum [5]. Tillering is mainly determined by the axillary meristem (AM), which is formed in leaf axils and may continue to grow and develop until tillering or dormancy [6,7]. Therefore, the whole tillering process is divided into two parts: axillary bud formation and outgrowth [8,9]. Tillering, as a complex process of plant growth and development, is the result of many factors, including environmental conditions [10], various hormones [11] and inheritance [12].
The GRAS [GAI (gibberellin-insensitive), RGA (repressor of GA1−3 mutant), and SCR (scarecrow)] family is a class of plant-specific transcription factors involved in many processes, such as growth and development [13], signal transmission [14,15], stress response [16]. OsMOC1, as a member of the GRAS family, is the first gene found to play an important role in tillering formation and outgrowth in rice, and its mutant shows an obvious monoculm phenomenon [17]. LS in tomato and LAS in Arabidopsis, which are the orthologues of MOC1, show significant tillering inhibition in mutants [18,19]. Recent studies in rice have shown that MOC1 functions as a coactivator of MOC3 [20], which can promote the expression of FLORAL ORGAN NUMBER 1 (FON1) [21] in the presence of MOC3 and further promote tillering. BRITTLE CULM 12 (BC12) negatively regulates the expression of MOC1, resulting in the inhibition of tillering in rice [22]. The degradation of MOC1 is inhibited by the combination with SLENDER RICE 1 (SLR1), but SLR1 can be degraded after gibberellin treatment [23]. Through the verification of interaction in vivo and in vitro, TILLERING AND DWARF 1 (TAD1) [24] and TILLER ENHANCER (TE) [25], the orthologues of CCS52, act as the coactivators of anaphase-promoting complex or cyclosomen (APC/C) [26], playing an important role in the degradation of MOC1, which results in the reduction of tillers in rice. In Arabidopsis, LAS activates REVOLUTA (REV) [27] and AUXIN RESISTANT 1 (AXR1) [8] during axillary meristem formation and lateral bud growth, respectively, to regulate shoot branching. However, there is no report on the interacting proteins of LAS or LS in dicots.
NHCC (Brassica rapa spp. chinensis), originating from China, is a leafy vegetable. Although research on tillering in rice [28], wheat [29], maize [30] and other economic crops has made great progress, it is still in the initial stage in NHCC. In our study, the expression of BcLAS was extremely abundant in leaf axils. Here, we used the ‘Maertou’ cultivar (tillering type), tillering at the vegetative growth stage, and the ‘Suzhouqing’ cultivar (common type), nontillering at the vegetative growth stage, to identify the function of BcLAS and BcCCS52. We found that when the expression of BcLAS was silenced, the tiller number of ‘Maertou’ decreased significantly at the vegetative growth stage. The overexpression of BcLAS obviously promoted tillering in ‘Suzhouqing’. Using yeast two-hybrid, bimolecular fluorescence complementation (BiFC) and luciferase complementation assays, BcCCS52 was determined to be an interacting protein of BcLAS. Silencing the expression of BcCCS52 in ‘Suzhouqing’ could promote tillering. Overexpression of BcCCS52 in Arabidopsis resulted in an obvious monoculm phenomenon. Our findings provide new insights into the tillering regulation mechanism during the vegetative growth of NHCCs.
Section snippets
Plant materials and growth conditions
For most NHCC cultivars, axillary buds enter dormancy after forming in axils until the transition from vegetative growth to reproductive growth, such as in ‘Suzhouqing’, but axillary buds will continue to grow at the vegetative growth stage for multiceps cultivars, such as in ‘Maertou’.
The B. rapa ssp. chinensis var. communis Tesnet Lee, cv. ‘Suzhouqing’ (common type), B. rapa ssp. chinensis var. multiceps, cv. ‘Maertou’ (tillering type), Arabidopsis and tobacco were grown in pots containing a
The characteristics of BcLAS
Previous studies found that OsMOC1 and AtLAS are irreplaceable in tillering regulation, which prompted us to verify whether there are genes with the same function in NHCC. A neighbour-joining phylogenetic tree (Fig. 1A) comprising the transcription factors of GRAS family in Arabidopsis and NHCC was constructed. Phylogenetic analysis showed that BraC06g00460.1 is an orthologue of AtLAS. BcLAS encodes 442 amino acids, which is 82.59 % consistent with AtLAS encoding 445 amino acids. We cloned BcLAS
Discussion
Tillering determines the number of leaves and inflorescences to some extent, so it is regarded as an important factor of yield in some plants. In our study about the tillering of NHCC, we found that BcLAS, as the orthologue of AtLAS belonging to the GRAS family, is an important positive regulator of tillering in the vegetative stage of NHCC and interacts with BcCCS52.
In the process of tillering, genetic factors have a major influence on the formation and outgrowth of axillary buds [38,39]. The
Author contributions
Mingliang Guo planned and designed the research. Mingliang Guo and Yan Long performed experiments. Mingliang Guo, Lanlan Xu, Wei Zhang, Tongkun Liu and Changwei Zhang analyzed the data. Mingliang Guo wrote the manuscript. Ying Li and Xilin Hou revised the manuscript.
Declaration of Competing Interest
The authors report no declarations of interest.
Acknowledgements
This study was supported by the National Natural Science Foundation of China (31872106), the National Key Research and Development Program (2018YFD1000800 and 2017YFD0101803), and National Vegetable Industry Technology System (CARS-23-A-06).
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