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Expression of chickpea CIPK25 enhances root growth and tolerance to dehydration and salt stress in transgenic tobacco.

Meena MK, Ghawana S, Dwivedi V, Roy A, Chattopadhyay D - Front Plant Sci (2015)

Bottom Line: Expression of CaCIPK25 and its high active form differentially increased salt and water-deficit tolerance demonstrated by improved growth and reduced leaf chlorosis suggesting that the kinase activity of CaCIPK25 was required for these functions.Expressions of the abiotic stress marker genes were enhanced in the CaCIPK25-expressing tobacco plants.Our results suggested that CaCIPK25 functions in root development and abiotic stress tolerance.

View Article: PubMed Central - PubMed

Affiliation: National Institute of Plant Genome Research New Delhi, India.

ABSTRACT
Calcium signaling plays an important role in adaptation and developmental processes in plants and animals. A class of calcium sensors, known as Calcineurin B-like (CBL) proteins sense specific temporal changes in cytosolic Ca(2+) concentration and regulate activities of a group of ser/thr protein kinases called CBL-interacting protein kinases (CIPKs). Although a number of CIPKs have been shown to play crucial roles in the regulation of stress signaling, no study on the function of CIPK25 or its orthologs has been reported so far. In the present study, an ortholog of Arabidopsis CIPK25 was cloned from chickpea (Cicer arietinum). CaCIPK25 gene expression in chickpea increased upon salt, dehydration, and different hormonal treatments. CaCIPK25 gene showed differential tissue-specific expression. 5'-upstream activation sequence (5'-UAS) of the gene and its different truncated versions were fused to a reporter gene and studied in Arabidopsis to identify promoter regions directing its tissue-specific expression. Replacement of a conserved threonine residue with an aspartic acid at its catalytic site increased the kinase activity of CaCIPK25 by 2.5-fold. Transgenic tobacco plants overexpressing full-length and the high active versions of CaCIPK25 displayed a differential germination period and longer root length in comparison to the control plants. Expression of CaCIPK25 and its high active form differentially increased salt and water-deficit tolerance demonstrated by improved growth and reduced leaf chlorosis suggesting that the kinase activity of CaCIPK25 was required for these functions. Expressions of the abiotic stress marker genes were enhanced in the CaCIPK25-expressing tobacco plants. Our results suggested that CaCIPK25 functions in root development and abiotic stress tolerance.

No MeSH data available.


Related in: MedlinePlus

Expression analysis of abiotic stress marker genes in CaCIPK25 overexpressing plants. Expression of known abiotic stress marker genes determined by qRT-PCR in 20-day-old control, CaCIPK25− and CaCIPK25T/D− overexpressing tobacco plants in response to treatments mentioned. Y-axis described the fold change of expression. Tobacco actin gene was used as internal control. * indicates statistically significant difference (p < 0.05) from the control sample.
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Figure 9: Expression analysis of abiotic stress marker genes in CaCIPK25 overexpressing plants. Expression of known abiotic stress marker genes determined by qRT-PCR in 20-day-old control, CaCIPK25− and CaCIPK25T/D− overexpressing tobacco plants in response to treatments mentioned. Y-axis described the fold change of expression. Tobacco actin gene was used as internal control. * indicates statistically significant difference (p < 0.05) from the control sample.

Mentions: Expression levels of the known genes related to abiotic stress signaling and tolerance were assessed in the transgenic plants under control and stress treatments. Five genes, namely NtERD10B (GenBank:AB049336), NtERD10C (AB049337), NtDREB1 (EU727155), NtDREB2 (EU727156), NtAPX1 (U15933.1), and were selected on the basis of their reported enhanced expression in transgenic plants showing tolerance to such stresses (Shukla et al., 2006; Tripathi et al., 2009; Bao et al., 2015; Zhou et al., 2015). NtERD10B and NtERD10C encode dehydrins, NtDREB1 and NtDREB2 encode dehydration responsive element (DRE)/C-repeat element (CRE) binding proteins and are transcription factors, and NtAPX1 encode ascorbate peroxidase. Only NtERD10B and NtERD10C showed more than 2-fold increase in expression level with respect to the vector-control plants only in the CaCIPK25T/D-overexpressing line in the control condition. Three other genes showed less than 2-fold increase in this line. Expression of all the five genes was not significantly increased in the CaCIPK25-overexpressing plants. Upon treatment with 20% PEG or 250 mM sodium chloride, expression of these genes increased several folds in the vector-transformed plants. In the CaCIPK25- or CaCIPK25T/D-expressing tobacco lines, increase in expression level of all the five genes was more than 2-fold as compared to the vector-transformed plants when exposed to PEG and sodium chloride. Further, expression of all the five genes were always higher in the CaCIPK25T/D lines than that in the CaCIPK25 lines, suggesting that the higher kinase activity of the protein resulted in higher expression of stress-related genes and, thereby, further enhanced tolerance level (Figure 9).


Expression of chickpea CIPK25 enhances root growth and tolerance to dehydration and salt stress in transgenic tobacco.

Meena MK, Ghawana S, Dwivedi V, Roy A, Chattopadhyay D - Front Plant Sci (2015)

Expression analysis of abiotic stress marker genes in CaCIPK25 overexpressing plants. Expression of known abiotic stress marker genes determined by qRT-PCR in 20-day-old control, CaCIPK25− and CaCIPK25T/D− overexpressing tobacco plants in response to treatments mentioned. Y-axis described the fold change of expression. Tobacco actin gene was used as internal control. * indicates statistically significant difference (p < 0.05) from the control sample.
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Figure 9: Expression analysis of abiotic stress marker genes in CaCIPK25 overexpressing plants. Expression of known abiotic stress marker genes determined by qRT-PCR in 20-day-old control, CaCIPK25− and CaCIPK25T/D− overexpressing tobacco plants in response to treatments mentioned. Y-axis described the fold change of expression. Tobacco actin gene was used as internal control. * indicates statistically significant difference (p < 0.05) from the control sample.
Mentions: Expression levels of the known genes related to abiotic stress signaling and tolerance were assessed in the transgenic plants under control and stress treatments. Five genes, namely NtERD10B (GenBank:AB049336), NtERD10C (AB049337), NtDREB1 (EU727155), NtDREB2 (EU727156), NtAPX1 (U15933.1), and were selected on the basis of their reported enhanced expression in transgenic plants showing tolerance to such stresses (Shukla et al., 2006; Tripathi et al., 2009; Bao et al., 2015; Zhou et al., 2015). NtERD10B and NtERD10C encode dehydrins, NtDREB1 and NtDREB2 encode dehydration responsive element (DRE)/C-repeat element (CRE) binding proteins and are transcription factors, and NtAPX1 encode ascorbate peroxidase. Only NtERD10B and NtERD10C showed more than 2-fold increase in expression level with respect to the vector-control plants only in the CaCIPK25T/D-overexpressing line in the control condition. Three other genes showed less than 2-fold increase in this line. Expression of all the five genes was not significantly increased in the CaCIPK25-overexpressing plants. Upon treatment with 20% PEG or 250 mM sodium chloride, expression of these genes increased several folds in the vector-transformed plants. In the CaCIPK25- or CaCIPK25T/D-expressing tobacco lines, increase in expression level of all the five genes was more than 2-fold as compared to the vector-transformed plants when exposed to PEG and sodium chloride. Further, expression of all the five genes were always higher in the CaCIPK25T/D lines than that in the CaCIPK25 lines, suggesting that the higher kinase activity of the protein resulted in higher expression of stress-related genes and, thereby, further enhanced tolerance level (Figure 9).

Bottom Line: Expression of CaCIPK25 and its high active form differentially increased salt and water-deficit tolerance demonstrated by improved growth and reduced leaf chlorosis suggesting that the kinase activity of CaCIPK25 was required for these functions.Expressions of the abiotic stress marker genes were enhanced in the CaCIPK25-expressing tobacco plants.Our results suggested that CaCIPK25 functions in root development and abiotic stress tolerance.

View Article: PubMed Central - PubMed

Affiliation: National Institute of Plant Genome Research New Delhi, India.

ABSTRACT
Calcium signaling plays an important role in adaptation and developmental processes in plants and animals. A class of calcium sensors, known as Calcineurin B-like (CBL) proteins sense specific temporal changes in cytosolic Ca(2+) concentration and regulate activities of a group of ser/thr protein kinases called CBL-interacting protein kinases (CIPKs). Although a number of CIPKs have been shown to play crucial roles in the regulation of stress signaling, no study on the function of CIPK25 or its orthologs has been reported so far. In the present study, an ortholog of Arabidopsis CIPK25 was cloned from chickpea (Cicer arietinum). CaCIPK25 gene expression in chickpea increased upon salt, dehydration, and different hormonal treatments. CaCIPK25 gene showed differential tissue-specific expression. 5'-upstream activation sequence (5'-UAS) of the gene and its different truncated versions were fused to a reporter gene and studied in Arabidopsis to identify promoter regions directing its tissue-specific expression. Replacement of a conserved threonine residue with an aspartic acid at its catalytic site increased the kinase activity of CaCIPK25 by 2.5-fold. Transgenic tobacco plants overexpressing full-length and the high active versions of CaCIPK25 displayed a differential germination period and longer root length in comparison to the control plants. Expression of CaCIPK25 and its high active form differentially increased salt and water-deficit tolerance demonstrated by improved growth and reduced leaf chlorosis suggesting that the kinase activity of CaCIPK25 was required for these functions. Expressions of the abiotic stress marker genes were enhanced in the CaCIPK25-expressing tobacco plants. Our results suggested that CaCIPK25 functions in root development and abiotic stress tolerance.

No MeSH data available.


Related in: MedlinePlus