Abstract
Jasmonates (JA) are oxylipin-derived phytohormones that trigger the production of specialized metabolites that often serve in defense against biotic stresses. In Medicago truncatula, a JA-induced endoplasmic reticulum-associated degradation (ERAD)-type machinery manages the production of bioactive triterpenes and thereby secures correct plant metabolism, growth, and development. This machinery involves the conserved RING membrane-anchor (RMA)-type E3 ubiquitin ligase MAKIBISHI1 (MKB1). Here, we discovered two additional members of this protein control apparatus via a yeast-based protein–protein interaction screen and characterized their function. First, a cognate E2 ubiquitin-conjugating enzyme was identified that interacts with MKB1 to deliver activated ubiquitin and to mediate its ubiquitination activity. Second, we identified a heat shock protein 40 (HSP40) that interacts with MKB1 to support its activity and was therefore designated MKB1-supporting HSP40 (MASH). MASH expression was found to be co-regulated with that of MKB1. The presence of MASH is critical for MKB1 and ERAD functioning because the dramatic morphological, transcriptional, and metabolic phenotype of MKB1 knock-down M. truncatula hairy roots was phenocopied by silencing of MASH. Interaction was also observed between the Arabidopsis thaliana (Arabidopsis) homologs of MASH and MKB1, suggesting that MASH represents an essential and plant-specific component of this vital and conserved eukaryotic protein quality control machinery.
Highlights
Jasmonates (JA) are oxylipin-derived phytohormones that trigger defense responses upon biotic stresses, confer tolerance to abiotic stresses, and regulate various developmental cues
In the model legume M. truncatula, it has been shown that the JA signaling machinery recruits the endoplasmic reticulum-associated degradation (ERAD) E3 ubiquitin ligase MKB1 to monitor TriterpeneMASH Supports Ubiquitin Ligase MAKIBISHI1 saponins (TSs) biosynthesis by controlling the stability of the rate-limiting enzyme hydroxy-3-methylglutaryl-CoA reductase (HMGR), as do different, but analogous, ERAD machineries to monitor sterol biosynthesis in yeast and animals (Pollier et al, 2013)
To increase our molecular understanding of this protein control apparatus, a protein– protein interaction screen was carried out that allowed to identify two hitherto unknown potential components of the MKB1dependent ERAD machinery: a canonical E2 UBC encoded by Medtr3g062450 and the JA-inducible heat shock protein 40 (HSP40) protein MKB1-supporting heat-shock protein 40 (MASH) encoded by Medtr3g100330
Summary
Jasmonates (JA) are oxylipin-derived phytohormones that trigger defense responses upon biotic stresses, confer tolerance to abiotic stresses, and regulate various developmental cues One of those defense responses consists of the elicitation of the biosynthesis of bio-active specialized metabolites (Goossens et al, 2016b; Wasternack and Strnad, 2019; Lacchini and Goossens, 2020). TS-specific 2,3-oxidosqualene cyclases (OSCs) cyclize 2,3-oxidosqualene, which can subsequently be followed by additional modifications, mainly oxidations, by cytochrome P450s (P450s), yielding a myriad of triterpene backbones or the sapogenins. These scaffolds can be decorated by UDP-dependent glycosyltransferases (UGTs), thereby further diversifying this class of specialized metabolites (Seki et al, 2015; Cárdenas et al, 2019)
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