Salicylic Acid Signaling Controls the Maturation and Localization of the Arabidopsis Defense Protein ACCELERATED CELL DEATH6

Abstract

ACCELERATED CELL DEATH6 (ACD6) is a multipass membrane protein with an ankyrin domain that acts in a positive feedback loop with the defense signal salicylic acid (SA). This study implemented biochemical approaches to infer changes in ACD6 complexes and localization. In addition to forming endoplasmic reticulum (ER)- and plasma membrane (PM)-localized complexes, ACD6 forms soluble complexes, where it is bound to cytosolic HSP70, ubiquitinated, and degraded via the proteasome. Thus, ACD6 constitutively undergoes ER-associated degradation. During SA signaling, the soluble ACD6 pool decreases, whereas the PM pool increases. Similarly, ACD6-1, an activated version of ACD6 that induces SA, is present at low levels in the soluble fraction and high levels in the PM. However, ACD6 variants with amino acid substitutions in the ankyrin domain form aberrant, inactive complexes, are induced by a SA agonist, but show no PM localization. SA signaling also increases the PM pools of FLAGELLIN SENSING2 (FLS2) and BRI1-ASSOCIATED RECEPTOR KINASE 1 (BAK1). FLS2 forms complexes ACD6; both FLS2 and BAK1 require ACD6 for maximal accumulation at the PM in response to SA signaling. A plausible scenario is that SA increases the efficiency of productive folding and/or complex formation in the ER, such that ACD6, together with FLS2 and BAK1, reaches the cell surface to more effectively promote immune responses.

Salicylic Acid Regulates Arabidopsis Microbial Pattern Receptor Kinase Levels and Signaling

“…In acd6-1, levels of MPK3, MPK6, and phosphorylated MPK3 and MPK6 were elevated ( Figure 3C; Supplemental Figure 2; Zhang et al, 2014). High levels of MPK3 and MPK6 were anticipated, as BTH treatment of wild-type plants induces these proteins, but not their phosphorylated forms, to accumulate (Beckers et al, 2009).…”

Arabidopsis AZI1 family proteins mediate signal mobilization for systemic defence priming

“…To obtain microsomal fractions 66 , leaf tissue (B1 g of N. benthamiana leaves) was frozen and ground in liquid nitrogen to a fine powder and then thawed in two volumes of ice-cold extraction buffer (50 mM Tris HCl, pH 7.5; 0.33 M sucrose; 5 mM EDTA; 150 mM NaCl and 1 Â complete protease inhibitor cocktail from Roche). The crude extracts were filtered through two layers of Miracloth and centrifuged at 10,000 g for 10 min to get total protein extract (supernatant), which was further ultracentrifuged at 100,000 g for 60 min to get the microsomal membrane and soluble fractions, respectively.…”

“…Membrane association assays 66 performed with microsomal protein extracts from N. benthamiana leaves infiltrated with Agrobacterium harbouring the different constructs. Extracts were treated with 1.5 M NaCl, 2 M urea, 1% Triton X-100, 1% NP-40 þ 0.5% deoxycolate or 1% SDS, respectively, at 22°C for 2 h with gentle shaking and fractionated into soluble supernatant or membrane pellet fractions by ultracentrifugation.…”

A Moving View: Subcellular Trafficking Processes in Pattern Recognition Receptor–Triggered Plant Immunity

“…This is dependent on accelerated cell death 6 (ACD6), a membrane-associated protein involved in positive feedback regulation with SA. ACD6 forms complexes with AtFLS2 and ER chaperones, possibly to facilitate the folding and delivery of AtFLS2 under SA-inducing conditions (152).…”