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Ser360 and Ser364 in the Kinase Domain of Tomato SIMAPKKKα are Critical for Programmed Cell Death Associated with Plant Immunity

  • Hwang, In Sun (Department of Horticultural Biotechnology, Kyung Hee University) ;
  • Brady, Jen (Boyce Thompson Institute for Plant Research) ;
  • Martin, Gregory B. (Boyce Thompson Institute for Plant Research) ;
  • Oh, Chang-Sik (Department of Horticultural Biotechnology, Kyung Hee University)
  • Received : 2016.11.20
  • Accepted : 2017.01.22
  • Published : 2017.04.01

Abstract

$SIMAPKKK{\alpha}$, a tomato (Solanum lycopersicum) mitogen-activated protein kinase kinase kinase, is a positive regulator of Pto-mediated effector-triggered immunity, which elicits programmed cell death (PCD) in plants. In this study, we examined whether putative phosphorylation sites in the conserved activation segment of the $SIMAPKKK{\alpha}$ kinase domain are critical for eliciting PCD. Three amino acids, $threonine^{353}$, $serine^{360}$ ($Ser^{360}$), or $serine^{364}$ ($Ser^{364}$), in the conserved activation segment of $SIMAPKKK{\alpha}$ kinase domain were substituted to alanine (T353A, S360A, or S364A), and these variants were transiently expressed in tomato and Nicotiana benthamiana plants. Two alanine substitutions, S360A and S364A, completely abolished $SIMAPKKK{\alpha}$ PCD-eliciting activity in both plants, while T353A substitution did not affect its PCD-eliciting activity. $SIMAPKKK{\alpha}$ wild type and variant proteins accumulated to similar levels in plant leaves. However, $SIMAPKKK{\alpha}$ protein with the largest size was missed when either S360A or S364A substitutions were expressed, whereas proteins with the smaller masses were more accumulated than those of full-length of $SIMAPKKK{\alpha}$ and T353A. These results suggest that phosphorylation of $SIMAPKKK{\alpha}$ at $Ser^{360}$ and $Ser^{364}$ is critical for PCD elicitation in plants.

Acknowledgement

Grant : Center for Horticultural Seed Development

Supported by : Golden Seed Project, Ministry of Agriculture, Food and Rural Affairs (MAFRA), Ministry of Oceans and Fisheries (MOF), Rural Development Administration (RDA), and Korea Forest Service (KFS)

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