Arabidopsis Ca2+-dependent protein kinase CPK3 mediates relationship of putative inositol triphosphate receptor with slow-type anion channel
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It has been suggested in Arabidopsis thaliana (L.) Heynh. cv. Columbia that, contrary to 30 μM abscisic acid (ABA), 20 μM ABA induces guard cell Ca2+ mobilization through activating phosphoinositide-specific phospholipase C (PI-PLC)-dependent inositol 1,4,5-triphosphate (IP3) production. Here, it was investigated whether Ca2+-dependent protein kinase, CPK3 or CPK6 would mediate ABA-induced stomatal closure downstream of IP3 production. In the knockout cpk3-1 mutant, the PLC inhibitor (U73122) adjusted 20 μM ABA-induced stomatal closure to the extent observed in the knockout cpk6-1 and cpk3-1cpk6-1 mutants and the wild type, whereas, in the wild type, the inhibitor of IP3-induced Ca2+ mobilization, xestospongin C (XeC), adjusted this closure to the extent observed in the cpk3-1 mutant. The Ca2+ buffer, EGTA and XeC positively interacted with the slow anion channel blocker, anthracene-9-carboxylic acid (9-AC) to inhibit 20 μM ABA-induced stomatal closure, which was suppressed in the dexamethasone-inducible AtPLC1 antisense transgene or the knockout cpk3-1, cpk6-1, cpk3-1cpk6-1 and NADPH oxidase atrbohD/F mutants. Discrete concentrations of 9-AC or another slow anion channel blocker (probenecid) negatively interacted with the Ca2+ buffer, BAPTA or the inhibitor of cyclic ADP-ribose-induced Ca2+ mobilization, ruthenium red, to inhibit 30 μM ABAinduced stomatal closure in the wild type but not in the cpk6-1, cpk3-1cpk6-1 and atrbohD/F mutants. Based on so far revealed features of the tested compounds and plant materials, interpretation of the results confirmed that guard cell ABA concentration discriminates between two Ca2+ mediations and outlined that one of them sequentially implicates CPK6, PLC1, a putative IP3 receptor homologue, CPK3, and the slow anion channel, whereas the other one excludes AtPLC1-dependent IP3 production and CPK3.
Additional key wordsabscisic acid concentration anion channel blockers Ca2+-dependent protein phosphorylation intracellular Ca2+ modulators phospholipase C stomatal closure
- AGI No.
Arabidopsis gene identifier number
Ca2+-dependent protein kinase
cystic fibrosis transmembrane conductance regulator
GTP-binding protein α-subunit
- G protein
multidrug resistance-associated protein
phosphoinositide-specific phospholipase C
rapid-type anion channel
slow-type anion channel
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This work was supported by an IFCPAR grant. The author gratefully thanks Prof. Chua (The Rockefeller University, New York, USA) for his gift of the dexamethasone-inducible AtPLC1 antisense transgene of A. thaliana and Prof. Schroeder (University of California, La Jolla, USA) for his gift of the cpk3-1, cpk6-1, cpk3-1cpk6-1 and atrbohD/F mutants of A. thaliana.
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