RAPID AND TRANSIENT ACTIVATION OF A MYELIN BASIC-PROTEIN KINASE IN TOBACCO-LEAVES TREATED WITH HARPIN FROM ERWINIA-AMYLOVORA

Harpins are bacterial protein elicitors that induce hypersensitive res ponse-like necrosis when infiltrated into nonhost plants such as tobac co (Nicotiana tabacum L.) (Z.-M. Wei, R.J. Laby, C.H. Zumoff, D.W. Bau er, S.Y. He, A. Collmer, S.V. Beer [1992] Science 257: 85-88). Activit y of a 49-kD Mg2+-dependent and Ca2+-independent kinase in tobacco lea ves increased 50-fold 15 min after infiltration of harpin from Erwinia amylovora (harpin(Ea)). Much less pronounced and more transient activ ation was detected in water-infiltrated leaves. Biochemical characteri stics of the harpin(Ea)-activated protein kinase (HAPK) activity are c onsistent with those of the mitogen-activated protein kinase family. H APK is cytosolic and phosphorylates myelin basic protein on serine/thr eonine residues. Treatment with a protein tyrosine phosphatase complet ely eliminated HAPK activity, suggesting that tyrosine phosphorylation is required for posttranslational activation. Sustained HAPK activati on after cycloheximide treatment implies that HAPK may be negatively r egulated by a translation-dependent mechanism. The extracellular Ca2chelater EGTA or the protein kinase inhibitor K252a, infiltrated in pl anta together with harpin(Ea), partially blocked HAPK activation. The Ca2+-channel blocker La3+ had no effect on HAPK activation, suggesting that phosphorylation events precede and/or do not depend on the entry of extracellular Ca2+ into the cell. These results suggest that early signal transduction events during harpin(Ea) induced hypersensitive r esponse elicitation depend in part on the activation of HAPK.