A Cdc48 "Retrochaperone" Function Is Required for the Solubility of Retrotranslocated, Integral Membrane Endoplasmic Reticulum-associated Degradation (ERAD-M) Substrates

J Biol Chem. 2017 Feb 24;292(8):3112-3128. doi: 10.1074/jbc.M116.770610. Epub 2017 Jan 11.


A surprising feature of endoplasmic reticulum (ER)-associated degradation (ERAD) is the movement, or retrotranslocation, of ubiquitinated substrates from the ER lumen or membrane to the cytosol where they are degraded by the 26S proteasome. Multispanning ER membrane proteins, called ERAD-M substrates, are retrotranslocated to the cytosol as full-length intermediates during ERAD, and we have investigated how they maintain substrate solubility. Using an in vivo assay, we show that retrotranslocated ERAD-M substrates are moved to the cytoplasm as part of the normal ERAD pathway, where they are part of a solely proteinaceous complex. Using proteomics and direct biochemical confirmation, we found that Cdc48 serves as a critical "retrochaperone" for these ERAD-M substrates. Cdc48 binding to retrotranslocated, ubiquitinated ERAD-M substrates is required for their solubility; removal of the polyubiquitin chains or competition for binding by addition of free polyubiquitin liberated Cdc48 from retrotranslocated proteins and rendered them insoluble. All components of the canonical Cdc48 complex Cdc48-Npl4-Ufd1 were present in solubilized ERAD-M substrates. This function of the complex was observed for both HRD and DOA pathway substrates. Thus, in addition to the long known ATP-dependent extraction of ERAD substrates during retrotranslocation, the Cdc48 complex is generally and critically needed for the solubility of retrotranslocated ERAD-M intermediates.

Keywords: Cdc48; E3 ubiquitin ligase; Hmg2; Npl4; Retrotranslocation; Ufd1; endoplasmic reticulum (ER); endoplasmic-reticulum-associated protein degradation (ERAD); ubiquitin; ubiquitin ligase.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adenosine Triphosphatases / metabolism*
  • Cell Cycle Proteins / metabolism*
  • Endoplasmic Reticulum / metabolism
  • Endoplasmic Reticulum-Associated Degradation*
  • Hydroxymethylglutaryl CoA Reductases / metabolism
  • Saccharomyces cerevisiae / cytology
  • Saccharomyces cerevisiae / metabolism*
  • Saccharomyces cerevisiae Proteins / metabolism
  • Solubility
  • Ubiquitin / metabolism
  • Ubiquitination
  • Valosin Containing Protein


  • Cell Cycle Proteins
  • Saccharomyces cerevisiae Proteins
  • Ubiquitin
  • HMG2 protein, S cerevisiae
  • Hydroxymethylglutaryl CoA Reductases
  • Adenosine Triphosphatases
  • CDC48 protein, S cerevisiae
  • Valosin Containing Protein