Regulation of CDPKs, including identification of PAL kinase, in biotically stressed cells of French bean

被引:30
作者
Allwood, EG [1 ]
Davies, DR [1 ]
Gerrish, C [1 ]
Bolwell, GP [1 ]
机构
[1] Univ London, Royal Holloway & Bedford New Coll, Sch Biol Sci, Div Biochem, Egham TW20 0EX, Surrey, England
关键词
calcium-dependent protein kinase; elicitor; oxidative burst; phenylalanine ammonia-lyase;
D O I
10.1023/A:1015502117870
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
Changes in protein kinase activity have been investigated during the early response of suspension cultured cells of French bean to fungal elicitor. One of the kinases activated has a known target, phenylalanine ammonia-lyase (PAL), which has an important role in plant defence responses, and was purified. Kinase acivity during purification was monitored for both the PAL-derived peptide and syntide-2, which it also phosphorylated. The kinase had an M-r of 55 000 on the basis of gel migration, Ca-45(2+) binding, autophosphorylation and phosphorylation of various substrates using in-gel assays. The kinase has been characterised with respect to kinetics and other properties in vitro and appears to be a CDPK. In-gel assays were also used to show that this kinase and a number of other CDPKs of similar M-r showed complex changes in elicitor-treated suspension-cultured cells of French bean. An activation was observed within 10 min and was maintained for up to 4 h. The time course of activation was different from MAP kinase and casein kinase assayed in the same extracts. However, at 5 min after addition of elicitor there is a transient inactivation of the CDPKs before activation. This inactivation can be mimicked by adding forskolin to the cells 30 min before elicitation, which brings about changes in the cellular pH. Forskolin potentiates the oxidative burst when elicitor is subsequently added while the CDPK cannot be activated by elicitor upon forskolin treatment. In contrast, intracellular acidification brought about by forskolin brings about slight activation of MAPkinase.
引用
收藏
页码:533 / 544
页数:12
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