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Planta

, Volume 248, Issue 3, pp 571–578 | Cite as

Casein kinase 2 α and β subunits inversely modulate ABA signal output in Arabidopsis protoplasts

  • Yukari Nagatoshi
  • Miki Fujita
  • Yasunari Fujita
Original Article

Abstract

Main conclusion

Our transient gene expression analyses in Arabidopsis protoplasts support the view that CK2αs and CK2βs positively and negatively modulate ABRE-dependent gene expression, respectively.

The phytohormone abscisic acid (ABA) regulates the expression of thousands of genes via ABA-responsive elements (ABREs), and has a crucial role in abiotic stress response. Casein kinase II (CK2), a conserved Ser/Thr protein kinase in eukaryotes, is essential for plant viability. Although the CK2 has been known as a tetrameric holoenzyme comprised of two catalytic α and two regulatory β subunits, each of the two types of subunits has been proposed to have independent functions. The Arabidopsis genome encodes four α subunits (CK2α1, CK2α2, CK2α3, CK2α4) and four β subunits (CK2β1, CK2β2, CK2β3, CK2β4). There is a growing body of evidence linking CK2 to ABA signaling and abiotic stress responses. However, the roles of each CK2 subunit in ABA signaling remain largely elusive. Using the transient expression system with the core ABA signaling components in Arabidopsis leaf mesophyll protoplasts, we show here that CK2α1 and CK2α2 (CK2α1/2) positively modulate ABRE-dependent gene expression as ABA signal output in ABA signaling, whereas all four CK2βs negatively modulate the ABRE-dependent gene expression mediated by subclass III SnRK2–AREB/ABF pathway and by CK2α1/2. These data indicate that CK2α1/2 and CK2βs positively and negatively modulate ABA signal output, respectively, suggesting that the quantitative balance of CK2 subunits determines the ABA signal output in plants. Given that CK2s act as pleiotropic enzymes involved in multiple developmental and stress–responsive processes, our findings suggest that CK2 subunits may be involved in integration and coordination of ABA-dependent and -independent signaling.

Keywords

ABA-responsive-elements Abscisic acid signaling Arabidopsis thaliana AREB/ABFs Protoplast transient expression system SnRK2 

Abbreviations

ABF

ABRE-binding factor

ABRE

ABA-responsive element

AREB

ABRE-binding protein

CK2

Casein kinase II

CK2s

Casein kinase II subunits

PP2C

Group-A protein phosphatase 2C

PYR1/PYL/RCAR

Pyrabactin resistance1/PYR1-like/regulatory components of ABA receptor

SnRK2

Subclass III sucrose nonfermenting 1 related protein kinase 2

Notes

Acknowledgements

We thank U. Mitsuyasu, T. Yoshida, E. Ohgawara, K. Mogami, M. Kishimoto, N. Takano, R. Motohashi, I. Saito, and M. Ikegami for excellent technical support, M. Toyoshima for skillful editorial assistance, and T. Ogata for critical reading of the manuscript.

Funding

This research was supported in part by the Ministry of Education, Culture, Sports, Science and Technology (MEXT) of Japan Grants-in-Aid for Scientific Research (C) (nos. 24510312 and 16K07412 to YF) and the Ministry of Agriculture, Forestry and Fisheries (MAFF) of Japan (to YN, YF).

Compliance with ethical standards

Conflict of interest

The authors declare no conflicts of interest.

Supplementary material

425_2018_2919_MOESM1_ESM.pdf (556 kb)
Suppl. Table S1 Oligonucleotide primers used in this study. Suppl. Fig. S1 Phylogenetic tree of CK2αs and CK2βs in Arabidopsis. The neighbor-joining phylogenetic tree (Saitou and Nei 1987) was created using MEGA7 (Kumar et al. 2016). The optimal tree with the sum of branch length = 1.25324778 is shown. Bootstrap values (1000 replicates) are shown next to the branches (Felsenstein 1985). Evolutionary distances were computed using the p-distance method (Nei and Kumar 2000); units represent the number of amino acid differences per site. The subcellular localization of each CK2 is based on previous reports (Salinas et al. 2006; Perales et al. 2006; Portoles and Mas 2010: Mulekar and Huq 2015). CK2αs and CK2βs are shaded in pink and in yellow, respectively. Suppl. Fig. S2 Alignment of CK2 amino acid sequences in Arabidopsis. Comparison of amino acid sequences of CK2αs (a) and CK2βs (b). Conservation ratio at each site is shown by shading (black, 100%; gray, 75%). Red bars mark reported characteristic domains. Red arrow indicates a signal peptide cleavage site detected by SignalP 4.1 Server (Petersen et al. 2011). Suppl. Fig. S3 Reconstitution of ABA signaling pathway by co-transfection of AREB1, ABI1 and PYL1. Protoplasts were isolated from WT leaves. RD29B-GUS (5.0 μg of plasmid per transformation) and pBI35SΩ-ELUC (1.0 μg per transfection) were used as the ABA-responsive reporter and internal control, respectively. Each transfection used 2.5 μg of effector plasmid, except for ABI1, which used 1.5 μg per transfection. Total amounts of effector DNA were 6.5 μg, which include effector plasmids alone or combined with the vector control plasmid pSKX for transient expression analysis. ‘Relative activity’ indicates combined expression relative to the value obtained from the vector control. After transfection, protoplasts were incubated for 14-18 h under dark conditions without ABA (open bars) or with 2.0 μM ABA (filled bars). Error bars indicate SD (n = 4). Experiments were performed at least three times, and a representative result is shown (PDF 555 kb)

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Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2018

Authors and Affiliations

  • Yukari Nagatoshi
    • 1
  • Miki Fujita
    • 2
  • Yasunari Fujita
    • 1
    • 3
  1. 1.Biological Resources and Post-harvest DivisionJapan International Research Center for Agricultural Sciences (JIRCAS)TsukubaJapan
  2. 2.RIKEN Center for Sustainable Resource ScienceTsukubaJapan
  3. 3.Graduate School of Life and Environmental SciencesUniversity of TsukubaTsukubaJapan

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