Pepper Heat Shock Protein 70a Interacts with the Type III Effector AvrBsT and Triggers Plant Cell Death and Immunity  

Abstract

Heat shock proteins (HSPs) function as molecular chaperones and are essential for the maintenance and/or restoration of protein homeostasis. The genus Xanthomonas type III effector protein AvrBsT induces hypersensitive cell death in pepper (Capsicum annuum). Here, we report the identification of the pepper CaHSP70a as an AvrBsT-interacting protein. Bimolecular fluorescence complementation and coimmunoprecipitation assays confirm the specific interaction between CaHSP70a and AvrBsT in planta. The CaHSP70a peptide-binding domain is essential for its interaction with AvrBsT. Heat stress (37°C) and Xanthomonas campestris pv vesicatoria (Xcv) infection distinctly induce CaHSP70a in pepper leaves. Cytoplasmic CaHSP70a proteins significantly accumulate in pepper leaves to induce the hypersensitive cell death response by Xcv (avrBsT) infection. Transient CaHSP70a overexpression induces hypersensitive cell death under heat stress, which is accompanied by strong induction of defense-and cell death-related genes. The CaHSP70a peptide-binding domain and ATPase-binding domain are required to trigger cell death under heat stress. Transient coexpression of CaHSP70a and avrBsT leads to cytoplasmic localization of the CaHSP70a-AvrBsT complex and significantly enhances avrBsT-triggered cell death in Nicotiana benthamiana. CaHSP70a silencing in pepper enhances Xcv growth but disrupts the reactive oxygen species burst and cell death response during Xcv infection. Expression of some defense marker genes is significantly reduced in CaHSP70a-silenced leaves, with lower levels of the defense hormones salicylic acid and jasmonic acid. Together, these results suggest that CaHSP70a interacts with the type III effector AvrBsT and is required for cell death and immunity in plants.

An Improved Brome mosaic virus Silencing Vector: Greater Insert Stability and More Extensive VIGS

“…It was demonstrated that transient overexpression of a cytoplasmic pepper (Capsicum annuum) HSP70 (HSP70a) during heat stress or Xanthamonas campestris pv vesicatoria infection caused cell death in pepper leaves (Kim and Hwang, 2015). Chen et al (2008) did not observe increased expression for any of the six HSP70 gene family members after infecting N. benthamiana plants with a PVX VIGS vector harboring a fragment of NbHSP70c-1.…”

Molecular functions of Xanthomonas type III effector AvrBsT and its plant interactors in cell death and defense signaling

“…HSP70 is involved in protein translocation, folding, synthesis, and macromolecular assemblies (Hartl and HayerHartl 2002;Mayer and Bukau 2005). Pepper heat shock protein 70a (CaHSP70a) physically interacts with AvrBsT in yeast and in planta (Kim and Hwang 2015b). Agrobacterium-mediated transient expression assays and Co-IP assays in N. benthamiana showed that AvrBsT directly binds to CaHSP70a in planta (Kim and Hwang 2015b).…”

“…Pepper heat shock protein 70a (CaHSP70a) physically interacts with AvrBsT in yeast and in planta (Kim and Hwang 2015b). Agrobacterium-mediated transient expression assays and Co-IP assays in N. benthamiana showed that AvrBsT directly binds to CaHSP70a in planta (Kim and Hwang 2015b).…”

“…A DNA-binding domain (BD) is fused with host interactors, and an activation domain (AD) is fused with AvrBsT to verify the interaction between AvrBsT and host interactors. Several AvrBsT-plant interactors have been identified from pepper and Arabidopsis cDNA libraries using avrBsT cDNA as bait, including pepper arginine decarboxylase1 (CaADC1) (Kim et al 2013b), pepper aldehyde dehydrogenase1 (CaALDH1) (Kim and Hwang 2015a), pepper heat shock protein 70a (CaHSP70a) (Kim and Hwang 2015b), pepper suppressor of the G2 allele of skp1 (CaSGT1) (Kim et al 2014b), pepper SNF1-related kinase1 (SnRK1) (Szczesny et al 2010), and Arabidopsis acetylated interacting protein1 (ACIP1) (Cheong et al 2014) (Table 1). Bimolecular fluorescence complementation (BiFC) assays (Hu et al 2002;Walter et al 2004) and coimmunoprecipitation (Co-IP) assays (Phizicky and Fields 1995) provide in planta verification that AvrBsT physically interacts with its plant interactors.…”

Varied expression pattern of the small heat shock protein gene encoding HSP17.7 against UVA, UVB, Cu²⁺and Zn²⁺stresses in sunflower

“…These proteins protect cells against harmful effects of stress conditions. Heat shock proteins (HSPs), also known as molecular chaperones, are known to belong among these stress proteins while they are present in all living organisms (Kim & Hwang 2015). Besides their role in protecting cells against environmental stress conditions, HSPs are essential components of plant cells and they play an important role in protein stabilisation and cellular functions related with plant growth (Pratt et al 2001;Koo et al 2015).…”