Selective autophagy regulates heat stress memory in Arabidopsis by NBR1-mediated targeting of HSP90.1 and ROF1

ABSTRACT

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

KEYWORDS:

• Arabidopsis thaliana
• heat stress
• HSFA2
• HSP90.1
• NBR1
• ROF1
• selective autophagy
• stress memory
• stress recovery

Acknowledgments

The pHSFA2:HSFA2-YFP seeds were kindly provided by Prof. Dr. Isabel Bäurle, University of Potsdam, Germany. We thank Dr. Karin Koehl and her team for the excellent plant care. We thank Josef Bergstein for photography service. This work was supported by funding from the Deutsche Forschungsgemeinschaft (DFG) to S.B. (SFB973). Additional financial support was provided by the MPI of Molecular Plant Physiology to SB, ASam, and ASk, the INRA Versailles to CMD, and by the US National Institute of Health NIGMS to RDV (RO1-GM124452).

Disclosure statement

The authors declare no competing financial interests.

Data availability

The mass spectrometry proteomics data were deposited in the ProteomeXchange Consortium via PRIDE [80] partner repository with the dataset identifier PXD015027.

Supplementary materials

Supplemental data for this article can be accessed here.

Additional information

Funding

This work was supported by the Deutsche Forschungsgemeinschaft [SFB 973].