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BdCIPK31, a Calcineurin B-Like Protein-Interacting Protein Kinase, Regulates Plant Response to Drought and Salt Stress

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BdCIPK31, a Calcineurin B-Like Protein-Interacting Protein Kinase, Regulates Plant Response to Drought and Salt Stress

Qingchen Luo et al. Front Plant Sci.

Abstract

Calcineurin B-like protein interacting protein kinases (CIPKs) are vital elements in plant abiotic stress signaling pathways. However, the functional mechanism of CIPKs has not been understood clearly, especially in Brachypodium distachyon, a new monocot model plant. In this study, BdCIPK31, a CIPK gene from B. distachyon was characterized. BdCIPK31 was downregulated by polyethylene glycol, NaCl, H2O2, and abscisic acid (ABA) treatments. Transgenic tobacco plants overexpressing BdCIPK31 presented improved drought and salt tolerance, and displayed hypersensitive response to exogenous ABA. Further investigations revealed that BdCIPK31 functioned positively in ABA-mediated stomatal closure, and transgenic tobacco exhibited reduced water loss under dehydration conditions compared with the controls. BdCIPK31 also affected Na+/K+ homeostasis and root K+ loss, which contributed to maintain intracellular ion homeostasis under salt conditions. Moreover, the reactive oxygen species scavenging system and osmolyte accumulation were enhanced by BdCIPK31 overexpression, which were conducive for alleviating oxidative and osmotic damages. Additionally, overexpression of BdCIPK31 could elevate several stress-associated gene expressions under stress conditions. In conclusion, BdCIPK31 functions positively to drought and salt stress through ABA signaling pathway. Overexpressing BdCIPK31 functions in stomatal closure, ion homeostasis, ROS scavenging, osmolyte biosynthesis, and transcriptional regulation of stress-related genes.

Keywords: ABA; Brachypodium distachyon; CIPK; Nicotiana tabacum; drought stress; salt stress.

Figures

FIGURE 1
FIGURE 1
BdCIPK31-BdCBLs interaction and BdCIPK31 subcellular localization analysis. (A) Yeast-two-hybrid analysis of the interactions between BdCIPK31 and BdCBLs. The pGADT7-BdCIPK31 and pGBKT7-BdCBLs vectors were co-transformed into AH109. The transformants were screened by DDO/-Leu/-Trp and TDO/-Leu/-Trp/-His for 3 days. (B) Subcellular localization analysis of BdCIPK31::GFP. The left column shows the epidermal cells expressing the free GFP protein as controls. The right column shows the epidermal cells expressing the BdCIPK31::GFP fusion protein. Three independent biological replicates were performed and produced similar results.
FIGURE 2
FIGURE 2
Expression patterns analyses of BdCIPK31 gene. (A) Organ-specific expressions of BdCIPK31 in leaf, root, stem, and spike of 6-week-old plant. Ten-day-old seedlings were treated by: (B) no treatment control, (C) 20% PEG6000, (D) 200 mM NaCl, (E) 100 mM H2O2, and (F) 100 μM ABA treatment, respectively. The BdCIPK31 expression in the leaves of treated seedlings was detected. Data in this figure represent the mean ± SE of three independent replicates. Different letters represent significant difference in each condition (Duncan’s test, P < 0.05).
FIGURE 3
FIGURE 3
Analysis of root elongation and fresh weight in control and OE plants exposed to osmotic and salt stress. Seedlings grown on (A) 1/2 MS medium, (B,C) 1/2 MS medium containing (B) 150/(C) 300 mM mannitol, (D,E) 1/2 MS medium containing (D) 150/(E) 200 mM NaCl, (F) 1/2 MS medium containing 0.1 mM Tu, (G) 1/2 MS medium containing 300 mM mannitol and 0.1 mM Tu, and (H) 1/2 MS medium with 200 mM NaCl and 0.1 mM Tu for 14 days. (I) Statistical analysis of root length. (J) Statistical analysis of fresh weight. Data in (I,J) represent the mean ± SE from three independent replicates. Different letters represent significant difference in each condition (Duncan’s test, P < 0.05).
FIGURE 4
FIGURE 4
Analysis of the drought and salt tolerance in control plants and transgenic plants overexpressing BdCIPK31. (A,D) Four-week-old tobacco plants grown in normal condition. (B) Plants were subjected to drought for 35 days. (C) Plants in (B) were re-watered for 7 days. (E) Plants were subjected to high salinity for 7 days. (F) Statistical analysis of survival rates. (G) Chlorophyll content analysis of plants subjected to salt treatment. Data in (F,G) represent the means ± SE from three independent replicates. Different letters represent significant difference in each condition (Duncan’s test, P < 0.05).
FIGURE 5
FIGURE 5
Analyses of the plant sensitivity exogenous ABA, ABA content, and ABA synthesis-related gene expression and under drought and salt treatments. Seedlings grown on (A) 1/2 MS medium, (B,C) 1/2 MS medium with (B) 2 μM or (C) 5 μM ABA for 21 days. (D) Root length analysis of control and OE seedlings. (E) Endogenous ABA level of control and OE plants before and after treatments. (F,G) Expression levels of (F) NtNCED1 and (G) NtABA2 in control and OE plants before and after treatments. Data in (D–G) represent the means ± SE from three independent replicates. Different letters represent significant difference in each condition (Duncan’s test, P < 0.05).
FIGURE 6
FIGURE 6
Analyses of RWC, water loss, and stomatal aperture. (A) Water loss rate of control and OE leaves in 24 h. (B) RWC of control and OE plants after drought treatment. Data in (A,B) represent the means ± SE from three independent replicates. (C) Stomatal closure under water deficiency, ABA, and Tu treatments. (D) Statistical analysis of the stomatal width:length ratio. Data in (D) represent the means ± SE from three independent replicates. Different letters represent significant difference in each condition (Duncan’s test, P < 0.05).
FIGURE 7
FIGURE 7
Analyses of ion content, K+ efflux in seedlings under salt condition, and expression analyses of ion-transporter genes in plants. (A) Na+ and (B) K+ levels in shoots and roots of control and OE seedlings under salt stress. (C) Net K+ efflux in root tips of WT and OE seedlings at the addition of 150 mM NaCl. The insert shows the K+ efflux before NaCl addition. (D) Expression levels of TORK1 in the leaves of control and OE plants under drought treatment. (E–G) Expression levels of (E) NKT1, (F) NtSOS1, and (G) NtNHX2 in the control and OE young seedlings under salt stress. Data in (A,B,D–G) represent the means ± SE from three independent replicates. Different letters represent significant difference in each condition (Duncan’s test, P < 0.05).
FIGURE 8
FIGURE 8
Analyses of oxidative tolerance control and OE plants. (A) Leaf disks obtained from control and OE seedlings treated with methyl viologen (MV). (B) Seedlings grown on 1/2 MS containing 0, 2, or 5 μM MV. (C) Chlorophyll content in leaves treated with 20 or 50 μM MV. (D) Average fresh weight of seedlings treated by 2 or 5 μM MV. Data in (C,D) represent the means ± SE from three independent replicates. Different letters represent significant difference in each condition (Duncan’s test, P < 0.05).
FIGURE 9
FIGURE 9
Expression analyses of stress-related genes in plants under drought and salt treatments. Expression levels of (A) NtABF1, (B) NtABF2, (C) NtRD26, (D) NtDREB3, (E) NtERD10C, (F) NtERD10D, (G) NtLEA5, and (H) TobLTP1 were examined by qRT-PCR. Data represent the means ± SE from three independent replicates. Different letters represent significant difference in each condition (Duncan’s test, P < 0.05).

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References

    1. Ache P., Becker D., Ivashikina N., Dietrich P., Roelfsema M. R. G., Hedrich R. (2000). GORK, a delayed outward rectifier expressed in guard cells of Arabidopsis thaliana, is a K+-selective, K+-sensing ion channel. FEBS Lett. 486 93–98. 10.1016/s0014-5793(00)02248-1 - DOI - PubMed
    1. Asano T., Tanaka N., Yang G., Hayashi N., Komatsu S. (2005). Genome-wide identification of the rice calcium-dependent protein kinase and its closely related kinase gene families: comprehensive analysis of the CDPKs gene family in rice. Plant Cell Physiol. 46 356–366. 10.1093/pcp/pci035 - DOI - PubMed
    1. Bartels D., Sunkar R. (2005). Drought and salt tolerance in plants. Crit. Rev. Plant Sci. 24 23–58. 10.1080/07352680590910410 - DOI
    1. Batistic O., Kudla J. (2004). Integration and channeling of calcium signaling through the CBL calcium sensor/CIPK protein kinase network. Planta 219 915–924. 10.1007/s00425-004-1333-3 - DOI - PubMed
    1. Becker W., Heukelbach J., Kentrup H., Joost H. G. (1996). Molecular cloning and characterization of a novel mammalian protein kinase harboring a homology domain that defines a subfamily of serine/threonine kinases. Eur. J. Biochem. 235 736–743. - PubMed

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