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Arabidopsis cargo receptor NBR1 mediates selective autophagy of defective proteins.

ABSTRACT: Aggrephagy, a type of selective autophagy that sequesters protein aggregates for degradation in the vacuole, is an important protein quality control mechanism, particularly during cell stress. In mammalian cells, aggrephagy and several other forms of selective autophagy are mediated by dedicated cargo receptors such as NEIGHBOR OF BRCA1 (NBR1). Although plant NBR1 homologs have been linked to selective autophagy during biotic stress, it remains unclear how they impact selective autophagy under non-stressed and abiotic stress conditions. Through microscopic and biochemical analysis of nbr1 mutants expressing autophagy markers and an aggregation-prone reporter, we tested the connection between NBR1 and aggrephagy in Arabidopsis. Although NBR1 is not essential for general autophagy, or for the selective clearance of peroxisomes, mitochondria, or the ER, we found that NBR1 is required for the heat-induced formation of autophagic vesicles. Moreover, cytoplasmic puncta containing aggregation-prone proteins, which were rarely observed in wild-type plants, were found to accumulate in nbr1 mutants under both control and heat stress conditions. Given that NBR1 co-localizes with these cytoplasmic puncta, we propose that Arabidopsis NBR1 is a plant aggrephagy receptor essential for maintaining proteostasis under both heat stress and non-stress conditions.

SUBMITTER: Jung H

PROVIDER: S-EPMC6913707 | biostudies-literature | 2020 Jan

REPOSITORIES: biostudies-literature

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Arabidopsis cargo receptor NBR1 mediates selective autophagy of defective proteins.

Jung Hyera H Lee Han Nim HN Marshall Richard S RS Lomax Aaron W AW Yoon Min Ji MJ Kim Jimi J Kim Jeong Hun JH Vierstra Richard D RD Chung Taijoon T

Journal of experimental botany 20200101 1

Aggrephagy, a type of selective autophagy that sequesters protein aggregates for degradation in the vacuole, is an important protein quality control mechanism, particularly during cell stress. In mammalian cells, aggrephagy and several other forms of selective autophagy are mediated by dedicated cargo receptors such as NEIGHBOR OF BRCA1 (NBR1). Although plant NBR1 homologs have been linked to selective autophagy during biotic stress, it remains unclear how they impact selective autophagy under n ...[more]

PMID: 31494674

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Project description:Autophagy involvement in plant response to nitrogen, carbon and sulfur starvation was already reported, however the mechanisms responsible for its regulation and selectivity in such conditions were not yet investigated. We observed increased amounts of NBR1 transcript in plants exposed to sulfur deficit as compared to the control plants grown in nutrient sufficient conditions. This observation prompted us to investigate the role of this selective autophagy cargo receptor in plant response to sulfur deficit. Transcriptome analysis of the wild type and NBR overexpressing plants revealed differences in gene expression changes in response to sulfur deficit. Moreover, NBR1 overexpressors have significantly shorter roots than WT, when grown in nutrient deficient conditions in the presence of TOR kinase inhibitors, namely in the conditions leading to autophagy induction. Besides, NBR1 overexpression promoted stomata closure while NBR1 depletion stomata opening. Surprisingly, all lines had more closed stomata when grown in sulfur deficient than sulfur optimal conditions, what indicates that this effect is independent from NBR1. Similarly, ABA-dependent stomatal closure was independent from NBR1 and growth conditions. Cysteine also promoted stomatal closure in NBR1-independent way in plants grown in the optimal medium but in contrast, reduced the number of open stomata in plants from sulfur deficient medium. Interaction network analysis of the proteins co-purifying with NBR1 revealed links with proteins involved in degradation systems, and endosomal trafficking and a surprizing connection with nuclear transport. In addition, several proteins co-purifying with NBR1 were found only in sulfur deficient conditions. One of them, ribosomal protein S6 (RPS6) was further confirmed as a direct NBR1 interactor. Localization of the RPS6 interaction sites in NBR1 indicated that ubiquitin binding domain of NBR1 is not required for this interaction what means that it is rather not the classical “ubiquitinated target - autophagy receptor” interaction.

Project description:In nature, plants are constantly exposed to many transient, but recurring, stresses. Thus, to complete their life cycles, plants require a dynamic balance between capacities to recover following cessation of stress and maintenance of stress memory. Recently, we uncovered a new functional role for macroautophagy/autophagy in regulating recovery from heat stress (HS) and resetting cellular memory of HS in Arabidopsis thaliana. Here, we demonstrated that NBR1 (next to BRCA1 gene 1) plays a crucial role as a receptor for selective autophagy during recovery from HS. Immunoblot analysis and confocal microscopy revealed that levels of the NBR1 protein, NBR1-labeled puncta, and NBR1 activity are all higher during the HS recovery phase than before. Co-immunoprecipitation analysis of proteins interacting with NBR1 and comparative proteomic analysis of an nbr1-null mutant and wild-type plants identified 58 proteins as potential novel targets of NBR1. Cellular, biochemical and functional genetic studies confirmed that NBR1 interacts with HSP90.1 (heat shock protein 90.1) and ROF1 (rotamase FKBP 1), a member of the FKBP family, and mediates their degradation by autophagy, which represses the response to HS by attenuating the expression of HSP genes regulated by the HSFA2 transcription factor. Accordingly, loss-of-function mutation of NBR1 resulted in a stronger HS memory phenotype. Together, our results provide new insights into the mechanistic principles by which autophagy regulates plant response to recurrent HS.Abbreviations: AIM: Atg8-interacting motif; ATG: autophagy-related; BiFC: bimolecular fluorescence complementation; ConA: concanamycinA; CoIP: co-immunoprecipitation; DMSO: dimethyl sulfoxide; FKBP: FK506-binding protein; FBPASE: fructose 1,6-bisphosphatase; GFP: green fluorescent protein; HS: heat stress; HSF: heat shock factor; HSFA2: heat shock factor A2; HSP: heat shock protein; HSP90: heat shock protein 90; LC-MS/MS: Liquid chromatography-tandem mass spectrometry; 3-MA: 3-methyladenine; NBR1: next-to-BRCA1; PQC: protein quality control; RFP: red fluorescent protein; ROF1: rotamase FKBP1; TF: transcription factor; TUB: tubulin; UBA: ubiquitin-associated; YFP: yellow fluorescent protein.

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Arabidopsis cargo receptor NBR1 mediates selective autophagy of defective proteins.

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Arabidopsis cargo receptor NBR1 mediates selective autophagy of defective proteins.

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