Traffic-independent function of the Sar1p/COPII machinery in proteasomal sorting of the cystic fibrosis transmembrane conductance regulator

J Cell Biol. 2003 Jan 20;160(2):157-63. doi: 10.1083/jcb.200210086. Epub 2003 Jan 21.

Abstract

Newly synthesized proteins that do not fold correctly in the ER are targeted for ER-associated protein degradation (ERAD) through distinct sorting mechanisms; soluble ERAD substrates require ER-Golgi transport and retrieval for degradation, whereas transmembrane ERAD substrates are retained in the ER. Retained transmembrane proteins are often sequestered into specialized ER subdomains, but the relevance of such sequestration to proteasomal degradation has not been explored. We used the yeast Saccharomyces cerevisiae and a model ERAD substrate, the cystic fibrosis transmembrane conductance regulator (CFTR), to explore whether CFTR is sequestered before degradation, to identify the molecular machinery regulating sequestration, and to analyze the relationship between sequestration and degradation. We report that CFTR is sequestered into ER subdomains containing the chaperone Kar2p, and that sequestration and CFTR degradation are disrupted in sec12ts strain (mutant in guanine-nucleotide exchange factor for Sar1p), sec13ts strain (mutant in the Sec13p component of COPII), and sec23ts strain (mutant in the Sec23p component of COPII) grown at restrictive temperature. The function of the Sar1p/COPII machinery in CFTR sequestration and degradation is independent of its role in ER-Golgi traffic. We propose that Sar1p/COPII-mediated sorting of CFTR into ER subdomains is essential for its entry into the proteasomal degradation pathway. These findings reveal a new aspect of the degradative mechanism, and suggest functional crosstalk between the secretory and the degradative pathways.

MeSH terms

  • COP-Coated Vesicles / genetics
  • COP-Coated Vesicles / metabolism*
  • COP-Coated Vesicles / ultrastructure
  • Cell Compartmentation / physiology
  • Cell Membrane / metabolism*
  • Cell Membrane / ultrastructure
  • Cysteine Endopeptidases / metabolism*
  • Cysteine Endopeptidases / ultrastructure
  • Cystic Fibrosis Transmembrane Conductance Regulator / metabolism*
  • Endoplasmic Reticulum / metabolism
  • Endoplasmic Reticulum / ultrastructure
  • Eukaryotic Cells / metabolism*
  • Eukaryotic Cells / ultrastructure
  • Fungal Proteins / metabolism
  • GTPase-Activating Proteins
  • HSP70 Heat-Shock Proteins / metabolism
  • Membrane Proteins / genetics
  • Membrane Proteins / metabolism
  • Microscopy, Electron
  • Monomeric GTP-Binding Proteins / metabolism*
  • Multienzyme Complexes / metabolism*
  • Multienzyme Complexes / ultrastructure
  • Nuclear Pore Complex Proteins
  • Proteasome Endopeptidase Complex
  • Protein Structure, Tertiary / physiology
  • Protein Transport / physiology*
  • Recombinant Fusion Proteins / metabolism
  • Saccharomyces cerevisiae
  • Saccharomyces cerevisiae Proteins / genetics
  • Saccharomyces cerevisiae Proteins / metabolism*
  • Vesicular Transport Proteins

Substances

  • Fungal Proteins
  • GTPase-Activating Proteins
  • HSP70 Heat-Shock Proteins
  • KAR2 protein, yeast
  • Membrane Proteins
  • Multienzyme Complexes
  • Nuclear Pore Complex Proteins
  • Recombinant Fusion Proteins
  • SEC13 protein, S cerevisiae
  • SEC23 protein, S cerevisiae
  • Saccharomyces cerevisiae Proteins
  • Vesicular Transport Proteins
  • Cystic Fibrosis Transmembrane Conductance Regulator
  • Cysteine Endopeptidases
  • Proteasome Endopeptidase Complex
  • Monomeric GTP-Binding Proteins
  • SAR1 protein, S cerevisiae