Interference with plastome gene expression and Clp protease activity in Arabidopsis triggers a chloroplast unfolded protein response to restore protein homeostasis

PLoS Genet. 2017 Sep 22;13(9):e1007022. doi: 10.1371/journal.pgen.1007022. eCollection 2017 Sep.

Abstract

Disruption of protein homeostasis in chloroplasts impairs the correct functioning of essential metabolic pathways, including the methylerythritol 4-phosphate (MEP) pathway for the production of plastidial isoprenoids involved in photosynthesis and growth. We previously found that misfolded and aggregated forms of the first enzyme of the MEP pathway are degraded by the Clp protease with the involvement of Hsp70 and Hsp100/ClpC1 chaperones in Arabidopsis thaliana. By contrast, the combined unfolding and disaggregating actions of Hsp70 and Hsp100/ClpB3 chaperones allow solubilization and hence reactivation of the enzyme. The repair pathway is promoted when the levels of ClpB3 proteins increase upon reduction of Clp protease activity in mutants or wild-type plants treated with the chloroplast protein synthesis inhibitor lincomycin (LIN). Here we show that LIN treatment rapidly increases the levels of aggregated proteins in the chloroplast, unleashing a specific retrograde signaling pathway that up-regulates expression of ClpB3 and other nuclear genes encoding plastidial chaperones. As a consequence, folding capacity is increased to restore protein homeostasis. This sort of chloroplast unfolded protein response (cpUPR) mechanism appears to be mediated by the heat shock transcription factor HsfA2. Expression of HsfA2 and cpUPR-related target genes is independent of GUN1, a central integrator of retrograde signaling pathways. However, double mutants defective in both GUN1 and plastome gene expression (or Clp protease activity) are seedling lethal, confirming that the GUN1 protein is essential for protein homeostasis in chloroplasts.

MeSH terms

  • Arabidopsis / genetics
  • Arabidopsis Proteins / genetics*
  • Chloroplasts / genetics*
  • DNA-Binding Proteins / genetics*
  • Endopeptidase Clp / genetics
  • Gene Expression Regulation, Plant / drug effects
  • Genome, Plant
  • HSP70 Heat-Shock Proteins / biosynthesis
  • HSP70 Heat-Shock Proteins / genetics
  • Heat Shock Transcription Factors
  • Heat-Shock Proteins / genetics*
  • Lincomycin / pharmacology
  • Molecular Chaperones / genetics
  • Photosynthesis / genetics
  • Plant Proteins / genetics*
  • Plants, Genetically Modified
  • Seedlings / genetics
  • Signal Transduction
  • Transcription Factors / genetics*
  • Unfolded Protein Response / genetics

Substances

  • APG6 protein, Arabidopsis
  • Arabidopsis Proteins
  • DNA-Binding Proteins
  • GUN1 protein, Arabidopsis
  • HSFA2 protein, Arabidopsis
  • HSP70 Heat-Shock Proteins
  • Heat Shock Transcription Factors
  • Heat-Shock Proteins
  • Hsp100 protein, Arabidopsis
  • Molecular Chaperones
  • Plant Proteins
  • Transcription Factors
  • Lincomycin
  • Endopeptidase Clp

Grants and funding

This work was funded by grants from the Spanish Ministry of Economy and Competitiveness (BIO2015-71703-REDT and BIO2014-59092-P) and Generalitat de Catalunya (2014SGR-1434). We also acknowledge the financial support from the Severo Ochoa Programme for Centres of Excellence in R&D 2016-2019 (SEV‐2015‐0533) and the CERCA Programme of the Generalitat de Catalunya. EL was supported by the Mexican CoNaCyT (PhD fellowships 421688 and “beca complemento”). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.