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BcMAF2 activates BcTEM1 and represses flowering in Pak-choi (Brassica rapa ssp. chinensis)

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Abstract

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BcMAF2 plays a key role in flowering regulation by controlling BcTEM1, BcSOC1 and BCSPL15 in Pak-choi.

Abstract

Flowering is a key event in the life cycle of plants. Flowering time shows an extensive variation from different Pak-choi (Brassica rapa ssp. chinensis) cultivars. However, the regulation mechanism of flowering in Pak-choi remains rarely known. In this study, a systematic identification and functional analysis of a Pak-choi MADS Affecting Flowering (MAF) gene, BcMAF2, was carried out. BcMAF2 encoded a protein containing a conserved MADS-box domain, which was localized in the nucleus. QPCR analysis indicated that the expression of BcMAF2 was higher in the leaves and flowers. Overexpression of BcMAF2 in Arabidopsis showed that BcMAF2 repressed flowering, which was further confirmed by silencing endogenous BcMAF2 in Pak-choi. In addition, Tempranillo 1 (TEM1) expression was up-regulated and MAF2 expression was down-regulated in the BcMAF2-overexpressing Arabidopsis. The expression of BcMAF2 and BcTEM1 was down-regulated in BcMAF2-silencing Pak-choi plants. The yeast one-hybrid, dual luciferase and qPCR results revealed that BcMAF2 protein could directly bind to BcTEM1 promoter and activate its expression, which was not reported in Arabidopsis. Meanwhile, a self-inhibition was found in BcMAF2. Taken together, this work suggested that BcMAF2 could repress flowering by directly activating BcTEM1.

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Abbreviations

AP3:

APETELA 3

CDF:

CYCLING DOF FACTOR

CO:

CONSTANS

FKF1:

FLAVIN KELCH F BOX 1

FLC:

Flowering Locus C

FT:

Flowering Locus T

GI:

GIGANTEA

LFY:

LEAFY

MAF:

MADS Affecting Flowering

PDS:

Phytoene desaturase

SOC1:

Suppressor of Overexpression of Constans 1

SPL:

Squamosa Promoter binding protein-Like

TEM1:

Tempranillo 1

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Acknowledgements

We are grateful to Dr Isabelle Jupin for providing the plasmid pTY-S. This work was supported by grants from Major Program of National Key Research and Development of China (2017YFD0101803), the Fundamental Research Funds for the Central Universities (Y0201700179), National Vegetable Industry Technology System (CARS-23-A-06), Jiangsu Modern Agriculture (vegetable) Industrial Technology System (SXGC[2017]273) and China Postdoctoral Science Foundation (2018M640493).

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Authors and Affiliations

  1. State Key Laboratory of Crop Genetics and Germplasm Enhancement/Key Laboratory of Biology and Germplasm Enhancement of Horticultural Crops in East China, Ministry of Agriculture, College of Horticulture, Nanjing Agricultural University, Nanjing, 210095, China

    Feiyi Huang, Tongkun Liu, Weike Duan & Xilin Hou

  2. Institute of Vegetable Crops, Jiangsu Academy of Agricultural Sciences, Nanjing, 210014, China

    Jun Tang

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  1. Feiyi Huang
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Contributions

Performed the experiments and wrote the paper: FYH. Manuscript revision and approval: FYH, TKL, JT and WKD. Contributed to the interpretation of the results and coordinated the study: XLH. All authors read and approved the final manuscript.

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Correspondence to Xilin Hou.

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11103_2019_867_MOESM1_ESM.jpg

Fig. S1 (a) Identification of T3 transgenic Arabidopsis plants with PCR. PCR analysis of T3 transgenic Arabidopsis plants overexpressing empty vector (control) and BcMAF2 (#2, #5, #35, #55 and #65). The amplified fragments on the left were the BcMAF2 coding sequence without termination codon (576 bp), on the right were the BcMAF2 coding sequence without termination codon plus a part of GFP coding sequence (850 bp). (b) Western Blot detection of #35 and #55 plants. The GFP and Actin antibodies were used (above and below). (c) Fluorescence observation of #35 and #55 plants (JPG 875 KB)

Fig. S2 Expression analysis of AtGA3OX1 and AtGA3OX2 in the control, #35 and #55 plants (JPG 238 KB)

11103_2019_867_MOESM3_ESM.jpg

Fig. S3 The expression of BcTEM1 in different tissues of Pak-choi. Data shown were means ± SE of three independent experiments (JPG 125 KB)

11103_2019_867_MOESM4_ESM.jpg

Fig. S4 QPCR analysis of the transcript level of BcMAF2 during the process of vernalization in Pak-choi. Data shown were means ± SE of three independent experiments (JPG 142 KB)

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Huang, F., Liu, T., Tang, J. et al. BcMAF2 activates BcTEM1 and represses flowering in Pak-choi (Brassica rapa ssp. chinensis). Plant Mol Biol 100, 19–32 (2019). https://doi.org/10.1007/s11103-019-00867-1

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