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Alfalfa (Medicago sativa L.) MsCML46 gene encoding calmodulin-like protein confers tolerance to abiotic stress in tobacco

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Abstract

Key message

MsCML46 enhances tolerance to abiotic stresses through alleviating osmotic stress and oxidative damage by regulating the expression of stress-related genes to optimize osmolytes levels and antioxidant enzyme activity in transgenic tobacco.

Abstract

Abiotic stresses are major environmental factors that constraint crop productivity worldwide. Various stimuli regulate intracellular calcium levels and calcium-mediated signal transduction, and cellular responses. Ca2+ signals are perceived by different Ca2+ receptors. Calmodulin-like protein (CML) is one of the best-characterized Ca2+ sensors which shares sequence similarity with highly conserved calmodulin (CaM) ubiquitously expressed in plants. Currently, the molecular and physiological functions of CMLs are largely unknown. In this study, the MsCML46 was characterized in alfalfa (Medicago sativa cv. Zhaodong) under freezing stress. Results showed that MsCML46 was localized to the cytoplasm of Arabidopsis, and its expression was strongly elevated by cold, drought, salt, saline-alkali, and ABA treatments. Overexpressing MsCML46 in tobacco enhanced tolerance to freezing, drought, and salt stresses as evidenced by improved contents of osmotic regulatory solutes and antioxidant enzyme activity but decreased reactive oxygen species (ROS) accumulation. Furthermore, cold, drought, and salt stresses increased the expression of stress-related genes in transgenic tobacco. MsCML46 binds free Ca2+ to promote signal transduction and maintain higher K+/Na+ ratio. In this way, it protects intracellular homeostasis under sodium ion toxicity. These results suggest that MsCML46 plays a crucial role in resisting abiotic stresses and can be exploited in genetic engineering for crops.

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Data availability

The MsCML46 sequence data that support the finding of this article is openly available in NCBI with GenBank accession number MT313689. Other data that support the findings of this study are available from the corresponding author upon reasonable request.

Abbreviations

ABA:

Abscisic acid

ANOVA:

Analysis of variance

CAT:

Catalase

DAB:

Diaminobenzidine

ECL:

Electrochemiluminescence

ERD:

Early response to dehydration protein

FW:

Fresh weight

HRP:

Horseradish peroxidase

LEA:

Late embryogenesis abundant protein

LTP:

Lipid transfer protein

MDA:

Malondialdehyde

NBT:

P-nitro blue tetrazolium chloride

NCBI:

National center for biotechnology information

ORF:

Open reading frame

P5CS:

Pyrroline-5-carboxylate synthetase

PEG:

Polyethylene glycol

PMSF:

Phenylmethylsulfonyl fluoride

POD:

Peroxidase

PVDF:

Polyvinylidene difluoride

RD29A:

Response to dehydration 29A

ROS:

Reactive oxygen species

SOD:

Superoxide dismutase

SRA:

Sequence read archive

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Acknowledgements

The project was sponsored by National Natural Science Foundation of China (31770575, 31972507), Science and Technology Major Project of Heilongjiang Province (GA19B103), Excellent Youth Project of Natural Science Foundation of Heilongjiang Province (YQ2020C033), Graduate Innovation Fund of Harbin Normal University (HSDSSCX2020-05), Shanghai Agricultural Science Committee Youth Talents Development Plan No. 2018 (1-34), Research Fund of Beijing Advanced Innovation Center for Food Nutrition and Human Health (20182018).

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  1. Binghao Du, Naiyu Chen and Lili Song are the co-first authors.

Authors and Affiliations

  1. Key Laboratory of Molecular and Cytogenetics, College of Life Science and Technology, Harbin Normal University, Harbin, 150025, Heilongjiang, China

    Binghao Du, Naiyu Chen, Lili Song, Dan Wang, Hongsheng Cai, Lin Yao & Changhong Guo

  2. Biotech Research Institute, Shanghai Academy of Agricultural Sciences, Shanghai, 201106, China

    Lili Song

  3. Beijing Advanced Innovation Center for Food Nutrition and Human Health, Beijing Technology and Business University (BTBU), Beijing, 100048, China

    Xiuting Li

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  1. Binghao Du
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Contributions

BD and LS conceived and designed the experiments. BD, NC and LS performed the experiments. BD analyzed the data and wrote the manuscript. DW, HC, LY, XL and CG supervised part of the work, contributed to the discussion and edited the manuscript. All authors read and approved the manuscript.

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Correspondence to Changhong Guo.

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The authors state that the planting materials were procured as per national regulations. All experimental materials were grown without causing any harm to the natural resource and biodiversity.

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Du, B., Chen, N., Song, L. et al. Alfalfa (Medicago sativa L.) MsCML46 gene encoding calmodulin-like protein confers tolerance to abiotic stress in tobacco. Plant Cell Rep 40, 1907–1922 (2021). https://doi.org/10.1007/s00299-021-02757-7

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  • DOI: https://doi.org/10.1007/s00299-021-02757-7

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