Calcium/Calmodulin Up-regulates a Cytoplasmic Receptor-like Kinase in Plants

Calcium/calmodulin-dependent kinases play an important role in protein phosphorylation in eukaryotes. However, not much is known about calcium/calmodulin-dependent protein phosphorylation and its role in signal transduction in plants. By using a protein-protein interaction-based approach, we have isolated a novel plant-specific calmodulin-binding receptor-like cytoplasmic kinase (CRCK1) from Arabidopsis thaliana, as well as its ortholog from Medicago sativa (alfalfa). CRCK1 does not show high homology to calcium/calmodulin-dependent protein kinases in animals. In contrast, it shows high homology in the kinase domain to serine/threonine receptor-like kinases in plants. However, it contains neither a transmembrane domain nor an extracellular domain. Calmodulin binds to CRCK1 in a calcium-dependent manner with an affinity of ∼20.5 nm. The calmodulin-binding site in CRCK1 is located in amino acids 160–183, which overlap subdomain II of the kinase domain. CRCK1 undergoes autophosphorylation in the presence of Mg2+ at the threonine residue(s). The Km and Vmax values of CRCK1 for ATP are 1 μm and 33.6 pmol/mg/min, respectively. Calcium/calmodulin stimulates the kinase activity of CRCK1, which increases the Vmax of CRCK1 ∼9-fold. The expression of CRCK1 is increased in response to stresses such as cold and salt and stress molecules such as abscisic acid and hydrogen peroxide. These results indicate the presence of a calcium/calmodulin-regulated receptor-like cytoplasmic kinase in plants. Furthermore, these results also suggest that calcium/calmodulin-regulated protein phosphorylation involving CRCK1 plays a role in stress signal transduction in plants. Calcium/calmodulin-dependent kinases play an important role in protein phosphorylation in eukaryotes. However, not much is known about calcium/calmodulin-dependent protein phosphorylation and its role in signal transduction in plants. By using a protein-protein interaction-based approach, we have isolated a novel plant-specific calmodulin-binding receptor-like cytoplasmic kinase (CRCK1) from Arabidopsis thaliana, as well as its ortholog from Medicago sativa (alfalfa). CRCK1 does not show high homology to calcium/calmodulin-dependent protein kinases in animals. In contrast, it shows high homology in the kinase domain to serine/threonine receptor-like kinases in plants. However, it contains neither a transmembrane domain nor an extracellular domain. Calmodulin binds to CRCK1 in a calcium-dependent manner with an affinity of ∼20.5 nm. The calmodulin-binding site in CRCK1 is located in amino acids 160–183, which overlap subdomain II of the kinase domain. CRCK1 undergoes autophosphorylation in the presence of Mg2+ at the threonine residue(s). The Km and Vmax values of CRCK1 for ATP are 1 μm and 33.6 pmol/mg/min, respectively. Calcium/calmodulin stimulates the kinase activity of CRCK1, which increases the Vmax of CRCK1 ∼9-fold. The expression of CRCK1 is increased in response to stresses such as cold and salt and stress molecules such as abscisic acid and hydrogen peroxide. These results indicate the presence of a calcium/calmodulin-regulated receptor-like cytoplasmic kinase in plants. Furthermore, these results also suggest that calcium/calmodulin-regulated protein phosphorylation involving CRCK1 plays a role in stress signal transduction in plants. Phosphorylation by protein kinase is one of the most common and important regulatory mechanisms in signal transduction in all organisms (1Hardie D.G. Annu. Rev. Plant Physiol. Plant Mol. Biol. 1999; 50: 97-131Crossref PubMed Scopus (259) Google Scholar). Since the first plant protein kinase sequences were reported in 1989, more than 1,000 have been reported in GenBank™. In particular, plants have a large number of receptor-like serine/threonine kinases (RLK) 1The abbreviations used are: RLK, receptor-like kinase; RLCK, receptor-like cytoplasmic kinase; CaM, calmodulin; CCaMK, chimeric calcium/calmodulin-dependent kinase; CRCK, calmodulin-regulated receptor-like cytoplasmic kinase; ABA, abscisic acid; T-DNA, transfer DNA. that resemble animal growth factor receptors, which can sense external signals and relay these signals by protein phosphorylation in plants (2Shiu S.H. Bleecker A.B. Science's STKE. 2001; (http:/www.stke.org/cgi/content/full/OC_sigtrans;2001/RE22)PubMed Google Scholar, 3Tichtinsky G. Vanoosthuyse V. Cock J.M. Gaude T. Trends Plant Sci. 2003; 8: 231-237Abstract Full Text Full Text PDF PubMed Scopus (74) Google Scholar, 4Becraft P.W. Annu. Rev. Cell Dev. Biol. 2002; 18: 163-192Crossref PubMed Scopus (189) Google Scholar). It is estimated that there are at least 600 RLK homologs, representing nearly 2.5% of the annotated protein-coding genes in Arabidopsis. Among them, 75% of the RLK family has a receptor configuration, with an extracellular domain, a transmembrane domain, and a kinase domain. These RLKs function in a wide range of signal response, such as BRI 1 for hormone perception (5Li J. Chory J. Cell. 1997; 90: 929-938Abstract Full Text Full Text PDF PubMed Scopus (913) Google Scholar, 6Scheer J.M. Ryan Jr., C.A. Proc. Natl. Acad. Sci. U. S. A. 2002; 99: 9585-9590Crossref PubMed Scopus (245) Google Scholar), SRK for pollen-pistil interaction (7Kachroo A. Nasrallah M.E. Nasrallah J.B. Plant Cell. 2002; 14: S227-238Crossref PubMed Scopus (80) Google Scholar), Xa21 for disease resistance (8Song W.Y. Wang G.L. Chen L.L. Kim H.S. Pi L.Y. Holsten T. Gardner J. Wang B. Zhai W.X. Zhu L.H. Fauquet C. Ronald P.C. Science. 1995; 270: 1804-1806Crossref PubMed Scopus (1772) Google Scholar), and CLAVATA1 for shoot apical meristem equilibrium (9Clark S.E. Williams R.W. Meyerowitz E.M. Cell. 1997; 89: 575-585Abstract Full Text Full Text PDF PubMed Scopus (1089) Google Scholar). Approximately 25% of the RLK family are cytoplasmic RLKs, containing only a kinase domain, and are thus named receptor-like cytoplasmic kinases (RLCKs). During the last decade, a few plant RLCKs, such as Pto and PBS1, have been characterized. In tomato, Pto confers resistance to the pathogen Pseudomonas syringae strains expressing avrPto (10Tang X. Frederick R.D. Zhou J. Halterman D.A. Jia Y. Martin G.B. Science. 1996; 274: 2060-2063Crossref PubMed Scopus (513) Google Scholar), and in Arabidopsis, PBS1 is required for specific resistance to P. syringae strains expressing avrPphB (11Swiderski M.R. Innes R.W. Plant J. 2001; 26: 101-112Crossref PubMed Google Scholar). However, the function of most plant RLCKs is not well understood. Calcium is a universal second messenger and acts as a mediator of stimulus-response coupling in the regulation of plant growth, development, and responses to environmental stimuli (12Poovaiah B.W. Reddy A.S. CRC Crit. Rev. Plant Sci. 1993; 12: 185-211Crossref PubMed Scopus (330) Google Scholar, 13Reddy A.S. Plant Sci. 2001; 160: 381-404Crossref PubMed Scopus (326) Google Scholar, 14Trewavas A.J. Malho R. Curr. Opin. Plant Biol. 1998; 1: 428-433Crossref PubMed Scopus (256) Google Scholar, 15Poovaiah B.W. Reddy A.S. CRC Crit. Rev. Plant Sci. 1987; 6: 47-103Crossref PubMed Scopus (283) Google Scholar, 16Yang T. Poovaiah B.W. Trends Plant Sci. 2003; 8: 505-512Abstract Full Text Full Text PDF PubMed Scopus (445) Google Scholar). Various stimuli, such as cold, salt, abscisic acid (ABA), and hydrogen peroxide, trigger changes in the cytosolic calcium concentration (17Knight H. Int. Rev. Cytol. 2000; 195: 269-324Crossref PubMed Google Scholar, 18Xiong L. Schumaker K.S. Zhu J.K. Plant Cell. 2002; 14: 165-183Crossref PubMed Scopus (185) Google Scholar, 19Pei Z.M. Murata Y. Benning G. Thomine S. Klusener B. Allen G.J. Grill E. Schroeder J.I. Nature. 2000; 406: 731-734Crossref PubMed Scopus (1675) Google Scholar, 20Price A.H. Taylor A. Ripley S.J. Griffiths A. Trewavas A.J. Knight M.R. Plant Cell. 1994; 6: 1301-1310Crossref PubMed Scopus (309) Google Scholar), which can be recognized by calcium receptors. Calmodulin (CaM), a small acidic protein with four EF-hand motifs, is one of the best characterized calcium receptors in eukaryotes. Upon calcium binding to the EF hands, CaM undergoes conformational changes, and the active Ca2+-CaM complex regulates the activity of downstream target proteins (13Reddy A.S. Plant Sci. 2001; 160: 381-404Crossref PubMed Scopus (326) Google Scholar, 16Yang T. Poovaiah B.W. Trends Plant Sci. 2003; 8: 505-512Abstract Full Text Full Text PDF PubMed Scopus (445) Google Scholar, 21Snedden W. Fromm H. New Physiologist. 2001; 151: 35-66Crossref Scopus (387) Google Scholar, 22Zielinski R.E. Annu. Rev. Plant Physiol. Plant Mol. Biol. 1998; 49: 697-725Crossref PubMed Scopus (397) Google Scholar). Ca2+/CaM-dependent protein kinases (CaM kinases) are the best characterized CaM-binding proteins in mammals and are major players in Ca2+/CaM-mediated signal transduction (23Hanson P.I. Schulman H. Annu. Rev. Biochem. 1992; 61: 559-601Crossref PubMed Scopus (664) Google Scholar). In plants, Ca2+/CaM-dependent protein phosphorylation Poovaiah B.W. Science. PubMed Scopus Google Scholar). However, only which show homology to CaM have been reported to there are a few reported CaM-binding protein kinases L. Y. Trends Plant Sci. 2003; 8: Full Text Full Text PDF PubMed Scopus Google Scholar). has reported that a chimeric Ca2+/CaM-dependent protein kinase autophosphorylation and Ca2+/CaM-dependent phosphorylation S. V. Poovaiah B.W. J. Biol. 1996; Full Text Full Text PDF PubMed Scopus Google Scholar, S. Poovaiah B.W. Proc. Natl. Acad. Sci. U. S. A. 1995; Scopus Google Scholar). is required for and in plants J. C. R. B. G. J.M. E. T. J. C. Science. PubMed Scopus Google Scholar). the activity of to be by CaM W. S. Biochem. J. 2003; PubMed Scopus Google Scholar). we the and of a novel plant-specific CRCK1, from Arabidopsis In the kinase domain, kinase has homology to receptor-like kinases than to CaM with subdomain II the kinase domain and the kinase Furthermore, the expression of CRCK1 in plants is by cold, salt, ABA, and hydrogen of A. and Medicago sativa expression were by using CaM as T. Poovaiah B.W. J. Biol. 2000; Full Text Full Text PDF PubMed Scopus Google Scholar). The were sequences and amino acid sequences were by using and The of and a Arabidopsis CRCK1 acids were from the The of CRCK1 by by using the and the sequences of the of for of were by from the with containing the for the of the expression The sequences of the by were the The were E. The were in at with a of CaM proteins were by and with CaM and in The were with the CaM and were to CaM CRCK1 to by acid the of the The protein an and with of CaM with in and of CaM and CaM in the were using a In to the CaM the used as a The from the of protein the specific The T. Poovaiah B.W. J. Biol. 2000; Full Text Full Text PDF PubMed Scopus Google Scholar). by using in the of and containing of CaM and of in and in a of were for 1 at The were by as T. Poovaiah B.W. J. Biol. 2000; Full Text Full Text PDF PubMed Scopus Google Scholar). Plant and were in a of and a at in a Plant stress were as T. Poovaiah B.W. J. Biol. 2002; Full Text Full Text PDF PubMed Scopus Google Scholar). In for cold stress plants were at for salt to the for and plants were with μm in with and the plants were with the were from the were at cold plants were and in and at used for protein and isolated from as T. Poovaiah B.W. J. Biol. 2000; Full Text Full Text PDF PubMed Scopus Google Scholar). were transfer to the were using and as T. Poovaiah B.W. J. Biol. 2000; Full Text Full Text PDF PubMed Scopus Google Scholar). to the amino acids of CRCK1, which does not show high homology to proteins in Arabidopsis, with were by The for and of were from and at and The as R. R.E. in New Scholar). The by in at and in and proteins were by plant with and with a The and proteins were from of plants as H. J. Plant Physiol. 2002; PubMed Scopus Google Scholar, M.R. C. PubMed Scopus Google Scholar). the at 1,000 for and the to at for The in of and by the of and of by at for The protein by the The proteins were and The to the by the using the The used as a of of with a in of were from the Arabidopsis The CRCK1 were by using and a as J.M. Kim Chen H. P. J. P. R. C. C. A. E. H. L. Y. H. E. C. R. P. L. T. E. A. Science. 2003; PubMed Scopus Google Scholar). The and the site The plants were by by using CRCK1 as a Furthermore, by using the CRCK1 not in in and Phosphorylation kinase as P. S. G. J. J. Biol. 1996; Full Text Full Text PDF PubMed Scopus Google with The autophosphorylation by using an that CRCK1, and of The phosphorylation by using CRCK1, of and μm ATP containing of The at for and using the ATP of protein with μm of ATP containing of and the by binding Poovaiah B.W. J. Biol. 2001; Full Text Full Text PDF PubMed Scopus Google Scholar). The by using acid as S. V. Poovaiah B.W. J. Biol. 1996; Full Text Full Text PDF PubMed Scopus Google Scholar). the CRCK1 from the and by at for and to using 1 and as a and were used as of the CRCK1 from plant protein as E. S. E. S. Y. J. B. P. J. Biol. 2002; Full Text Full Text PDF PubMed Scopus Google with plants were in an containing 1 1 and which at The were and and 1 1 of protein with of the with of protein were and the were for an at and with the The complex to autophosphorylation in the containing and of for at and the by The proteins were by the the were in and the were to an of A. and Medicago sativa were with were from These known CaM-binding such as G. Chen Y. T. H. Fromm H. J. Biol. 1993; Full Text PDF PubMed Google Scholar), W. Reddy A.S. Poovaiah B.W. Plant Mol. Biol. 1996; PubMed Scopus Google Scholar, A.S. X. J. Biol. 1996; Full Text Full Text PDF PubMed Scopus Google Scholar), S. Poovaiah B.W. Proc. Natl. Acad. Sci. U. S. A. 1995; Scopus Google Scholar), and T. Poovaiah B.W. J. Biol. 2000; Full Text Full Text PDF PubMed Scopus Google Scholar). from which for a with high homology to plant RLCK, and were named Arabidopsis CRCK1 and The sequences of Arabidopsis CRCK1 in the are the as of the and are and of the and sequences in the that is a of an of the site in the is that has by were by using that the is not Arabidopsis CRCK1 for a with amino has a of and a of for a with amino has a of and a of These and for amino acid that are have kinase with all of the serine/threonine that have homology to plant RLKs PBS1, BRI and than CaM kinases from plants and are plant-specific RLKs S.H. Bleecker A.B. Plant Physiol. 2003; PubMed Scopus Google Scholar). the homology is with Pto and with However, there is than homology with and a are a family of plant protein kinases containing a CaM domain to the kinase domain S.H. M.R. Chen J. Plant Physiol. 2002; PubMed Scopus Google Scholar). It is that and in the Arabidopsis show high homology to CRCK1 that are CRCK1 and are thus named and than the kinase domain in the of CRCK1 have homology to known in the In have extracellular domain transmembrane domain the protein that are cytoplasmic RLKs S.H. Bleecker A.B. Plant Physiol. 2003; PubMed Scopus Google Scholar). CaM-binding that are CaM-binding proteins from an E. containing the of Arabidopsis CRCK1 and the were to CaM binding CaM binds to the CRCK1 and not in the presence of However, in the presence of a calcium 1 not the CaM binding not that the CaM-binding of are using the to the of CRCK1 The CaM binding affinity of CRCK1 by CRCK1 a and with of CaM in the presence of The CaM binds to CRCK1 with a at the presence of a high affinity binding site in the of the the of CaM for CRCK1 estimated to be about nm. The binding of CaM to CRCK1 in the presence of The also that the of CRCK1 has a CaM-binding The CaM-binding of CRCK1 the characterized CaM-binding proteins have a a the amino acid in the CaM-binding of these proteins are not Cell. 2002; Full Text Full Text PDF PubMed Scopus Google Scholar, Trends Biochem. Sci. Full Text PDF PubMed Scopus Google Scholar). of the sequences that the amino acids subdomain II of the CRCK1 kinase domain the CaM-binding The amino acid an with a and a In particular, a in a of and is a common in known CaM Trends Biochem. Sci. Full Text PDF PubMed Scopus Google Scholar, T. G. Fromm H. Plant Physiol. 1995; PubMed Scopus Google Scholar). with to the CaM-binding acids with CaM The is of a complex with CaM plant CaM in the presence of not in the presence of the a CaM the representing the complex the of to CaM the CaM and the of the complex CaM as a of is These indicate that the CaM-binding site is located amino acids in the CaM-binding site is located subdomain II of the kinase domain, which contains a for ATP binding PubMed Scopus Google Scholar). of the CaM-binding site of CRCK1 with kinases that CRCK1 and its ortholog have amino acid Arabidopsis and a domain to the CaM-binding domain of CRCK1, that are also CaM-binding protein In contrast, the of Pto and PBS1 show homology to However, that the can an not that are also CaM-binding It be that with kinases in does not show high that CaM binding is to CaM the the kinase activity of CRCK1, CRCK1 with the in E. to by and by CRCK1 to in the presence of Mg2+ and the autophosphorylation about However, activity in the presence of CRCK1 with to The Km and Vmax values for by a are 1 μm and 33.6 pmol/mg/min, CRCK1 is a serine/threonine a acid The acid that CRCK1 at the threonine phosphorylation in and is a cytosolic protein kinase with binding in CRCK1 is a cytosolic and proteins and of the proteins from plants and of proteins from plants were to by using the CRCK1 complex from the plant and plants, and the were to autophosphorylation The the CaM with CRCK1 in CRCK1 from the proteins of and plants. The were to by using an The the role of in the kinase CRCK1 to autophosphorylation and phosphorylation in the presence of It that of CaM autophosphorylation and about a in kinase activity in the presence of CaM The Vmax of the in the presence of CaM increased from 33.6 to of in Km in the of of which is a kinase Biochem. J. 1994; PubMed Scopus Google Scholar, A.H. C. A. Knight M.R. Trewavas A.J. Plant Physiol. 1999; PubMed Scopus Google Scholar). Furthermore, the kinase activity by CaM with of the and the CaM These results suggest that regulates the kinase CaM regulates activity by with the the to the CaM-binding site acids to the in the presence of The kinase activity to concentration increased that CaM with amino acids of the kinase and increases its CRCK1 a CaM-binding in results that only a protein of the of CRCK1, recognized by the in the proteins of plants the of the and to the of the isolated from not The of recognized by in the plants in the plants that the plant In the CRCK1 protein in the not in the by that CRCK1 is a cytosolic that the of CRCK1 protein in the is than the protein be the of in the protein for protein and protein be the CRCK1 protein The also used for of plant CRCK1 from proteins of plants. The proteins were to an and a that the of CRCK1 However, in the plants. is a in the proteins from the plants. be a of The were also to an protein to CaM in in the with of also which CaM Arabidopsis has CaM genes that for T. Poovaiah B.W. Trends Plant Sci. 2003; 8: 505-512Abstract Full Text Full Text PDF PubMed Scopus (445) Google Scholar, 21Snedden W. Fromm H. New Physiologist. 2001; 151: 35-66Crossref Scopus (387) Google Scholar), which a homology from to However, CaM in the protein from plants Furthermore, CaM in the with that with CRCK1 in as in the with the for not of expression of CRCK1 by using the of CRCK1 as a of with expression in all not CRCK1 isolated from a the of cold CRCK1 expression in CRCK1 expression cold as well as increased expression of CRCK1 protein The expression of CRCK1 and proteins also increased salt, and and that kinase is in the transduction of cold, salt, and stress signals to and It is that the signal transduction involving cold and salt stress as well as signal molecules such as and stress such as common L. Schumaker K.S. Zhu J.K. Plant Cell. 2002; 14: 165-183Crossref PubMed Scopus (185) Google Scholar). RLCKs are a of kinases that kinase as RLKs the extracellular receptor (2Shiu S.H. Bleecker A.B. Science's STKE. 2001; (http:/www.stke.org/cgi/content/full/OC_sigtrans;2001/RE22)PubMed Google Scholar, S.H. Bleecker A.B. Plant Physiol. 2003; PubMed Scopus Google Scholar). than RLCKs are in the Arabidopsis the of most RLCKs are The results reported the and of a novel plant-specific CaM-binding protein CRCK1 from Arabidopsis that contains of have also isolated a CRCK1 ortholog from that is in plants. In there are CRCK1 homologs, and in the Arabidopsis that to a small of these proteins a CaM-binding that are CaM-binding CRCK1 also has homology to characterized RLCKs, Pto (10Tang X. Frederick R.D. Zhou J. Halterman D.A. Jia Y. Martin G.B. Science. 1996; 274: 2060-2063Crossref PubMed Scopus (513) Google and Arabidopsis PBS1 (11Swiderski M.R. Innes R.W. Plant J. 2001; 26: 101-112Crossref PubMed Google Scholar). Pto and PBS1 are in plant resistance to specific pathogen in plants. of the of Pto and PBS1 in the to the CaM-binding site of CRCK1 that can an a CaM-binding CaM binding also a CaM-binding site in that Pto and PBS1 are CaM-binding the and Pto and PBS1 and role in plant disease resistance be of to plant and CaM-binding RLKs from plants, have been reported A. R. B. Biochem. J. PubMed Scopus Google Scholar, V. G. C. Gaude T. Cock J.M. Plant Physiol. 2003; PubMed Scopus Google Scholar). and an extracellular domain, a transmembrane domain, and a kinase domain. The CaM-binding site is located the transmembrane domain in and in subdomain in However, the binding affinity is for and does not the kinase activity A. R. B. Biochem. J. PubMed Scopus Google Scholar, V. G. C. Gaude T. Cock J.M. Plant Physiol. 2003; PubMed Scopus Google Scholar). In contrast, has a high affinity for CRCK1 and the kinase activity of CRCK1 is binding to the CaM-binding and It is that the CaM-binding site in CRCK1 is located subdomain II of the kinase domain, which contains a for ATP binding In contrast, the CaM-binding in CaM kinases are to overlap the that are of the domain binding to the kinase a conformational that the domain and for activity Cell. 2002; Full Text Full Text PDF PubMed Scopus Google Scholar, Trends Cell Biol. 2000; Full Text Full Text PDF PubMed Scopus Google Scholar, Google Scholar). CRCK1 Ca2+/CaM-dependent changes in kinase the is In to the there are at least CaM that have been the conformational of CaM T. Poovaiah B.W. Trends Plant Sci. 2003; 8: 505-512Abstract Full Text Full Text PDF PubMed Scopus (445) Google Scholar, Cell. 2002; Full Text Full Text PDF PubMed Scopus Google Scholar). is active site as in the of of Upon a domain of factor undergoes a from the which a and to Cell. 2002; Full Text Full Text PDF PubMed Scopus Google Scholar, C. S. J. Y. S. A. Nature. 2002; PubMed Scopus Google Scholar). is CaM molecules with of a The EF to the and the EF are for to and coupling changes in and Cell. 2002; Full Text Full Text PDF PubMed Scopus Google Scholar, A. H. J. Nature. 2001; PubMed Scopus Google Scholar). It is that the role of CaM binding to CRCK1 is animal CaM kinases the CaM-binding site in CRCK1 is the domain of the CaM binding to CRCK1 increases the Vmax by the in Km values as in most CaM-binding and Biochem. J. 1994; PubMed Scopus Google reported a for the of kinase which is a CaM to the Vmax of the in Km is not well and a novel for the regulation of are to the of the regulation CRCK1 In plants, a of environmental and such as cold, salt, ABA, and hydrogen trigger changes in cytosolic (17Knight H. Int. Rev. Cytol. 2000; 195: 269-324Crossref PubMed Google Scholar, 18Xiong L. Schumaker K.S. Zhu J.K. Plant Cell. 2002; 14: 165-183Crossref PubMed Scopus (185) Google Scholar, 19Pei Z.M. Murata Y. Benning G. Thomine S. Klusener B. Allen G.J. Grill E. Schroeder J.I. Nature. 2000; 406: 731-734Crossref PubMed Scopus (1675) Google Scholar, 20Price A.H. Taylor A. Ripley S.J. Griffiths A. Trewavas A.J. Knight M.R. Plant Cell. 1994; 6: 1301-1310Crossref PubMed Scopus (309) Google Scholar). The cytosolic changes are by receptors such as CaM, which in the downstream signal and to responses in plants (13Reddy A.S. Plant Sci. 2001; 160: 381-404Crossref PubMed Scopus (326) Google Scholar, 16Yang T. Poovaiah B.W. Trends Plant Sci. 2003; 8: 505-512Abstract Full Text Full Text PDF PubMed Scopus (445) Google Scholar, 21Snedden W. Fromm H. New Physiologist. 2001; 151: 35-66Crossref Scopus (387) Google Scholar). The of protein phosphorylation has been recognized in (23Hanson P.I. Schulman H. Annu. Rev. Biochem. 1992; 61: 559-601Crossref PubMed Scopus (664) Google Scholar, Cell. 2002; Full Text Full Text PDF PubMed Scopus Google Scholar). the of the function of plant kinases has been by and L. Y. Trends Plant Sci. 2003; 8: Full Text Full Text PDF PubMed Scopus Google Scholar). of CRCK1 and its as is an important in the role of Ca2+/CaM-mediated phosphorylation in plants. the expression of CRCK1 is by cold and salt as well as the stress molecules and hydrogen that CRCK1 be in and stress signal transduction in plants. it is that CRCK1 to environmental signals in by the protein expression and by its kinase activity in response to changes in cytosolic It be that in to plants a of CaM genes that for CaM (13Reddy A.S. Plant Sci. 2001; 160: 381-404Crossref PubMed Scopus (326) Google Scholar, 16Yang T. Poovaiah B.W. Trends Plant Sci. 2003; 8: 505-512Abstract Full Text Full Text PDF PubMed Scopus (445) Google Scholar, 21Snedden W. Fromm H. New Physiologist. 2001; 151: 35-66Crossref Scopus (387) Google Scholar). Arabidopsis has CaM genes that for CaM The CaM in to and known in S.H. C. A. Fromm H. Biochem. J. 2000; PubMed Scopus Google Scholar, S.H. Kim Kim Kim S.E. J. Biol. 2002; Full Text Full Text PDF PubMed Scopus Google Scholar). In CaM and were and were in CaM binding and However, it is not known CRCK1 can be by all Arabidopsis CaM specific In CRCK1 be by to which can be by a from P. syringae avrPto that response in plants (10Tang X. Frederick R.D. Zhou J. Halterman D.A. Jia Y. Martin G.B. Science. 1996; 274: 2060-2063Crossref PubMed Scopus (513) Google Scholar). the of CRCK1 and its in the role of Ca2+/CaM-mediated stress in plants and plants to and to the for of A. and sativa and of for