Cysteine string protein promotes proteasomal degradation of the cystic fibrosis transmembrane conductance regulator (CFTR) by increasing its interaction with the C terminus of Hsp70-interacting protein and promoting CFTR ubiquitylation.
Cysteine string protein (Csp) is a J-domain-containing protein whose overexpression blocks the exit of cystic fibrosis transmembrane conductance regulator (CFTR) from the endoplasmic reticulum (ER). Another method of blocking ER exit, the overexpression of Sar1-GTP, however, yielded twice as much immature CFTR compared with Csp overexpression. This finding suggested that Csp not only inhibits CFTR ER exit but also facilitates the degradation of immature CFTR. This was confirmed by treatment with a proteasome inhibitor, which returned the level of immature CFTR to that found in cells expressing Sar1-GTP only. CspH43Q, which does not interact with Hsc70/Hsp70 efficiently, did not promote CFTR degradation, suggesting that the pro-degradative effect of Csp requires Hsc70/Hsp70 binding/activation. In agreement with this, Csp overexpression increased the amount of Hsc70/Hsp70 co-immunoprecipitated with CFTR, whereas overexpression of CspH43Q did not. The Hsc70/Hsp70 binding partner C terminus of Hsp70-interacting protein (CHIP) can target CFTR for proteasome-mediated degradation. Csp overexpression also increased the amount of CHIP co-immunoprecipitated with CFTR. In addition, CHIP interacted directly with Csp, which was confirmed by in vitro binding experiments. Csp overexpression also increased CFTR ubiquitylation and reduced the half-life of immature CFTR. These findings indicate that Csp not only regulates the exit of CFTR from the ER, but that this action is accompanied by Hsc70/Hsp70 and CHIP-mediated CFTR degradation.
Schmidt
Béla Z
BZ
Department of Cell Biology and Physiology, University of Pittsburgh, Pittsburgh, Pennsylvania 15261.
Watts
Rebecca J
RJ
Aridor
Meir
M
Frizzell
Raymond A
RA
D019005:Cystic Fibrosis Transmembrane Conductance Regulator / Q000235:genetics / Q000378:metabolism*; D004721:Endoplasmic Reticulum / Q000235:genetics / Q000378:metabolism*; D050883:HSC70 Heat-Shock Proteins / Q000235:genetics / Q000378:metabolism*; D050956:HSP40 Heat-Shock Proteins / Q000235:genetics / Q000378:metabolism*; D006367:HeLa Cells; D006801:Humans; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D020559:Monomeric GTP-Binding Proteins / Q000235:genetics / Q000378:metabolism; D011480:Protease Inhibitors / Q000494:pharmacology; D046988:Proteasome Endopeptidase Complex / Q000235:genetics / Q000378:metabolism*; D061988:Proteasome Inhibitors; D017434:Protein Structure, Tertiary / Q000502:physiology; D044767:Ubiquitin-Protein Ligases / Q000235:genetics / Q000378:metabolism; D054875:Ubiquitination / Q000187:drug effects / Q000502:physiology*
C505032:CFTR protein, human; D050883:HSC70 Heat-Shock Proteins; D050956:HSP40 Heat-Shock Proteins; C494371:HSPA8 protein, human; D008565:Membrane Proteins; D011480:Protease Inhibitors; D061988:Proteasome Inhibitors; C088460:cysteine string protein; D019005:Cystic Fibrosis Transmembrane Conductance Regulator; C467728:STUB1 protein, human; D044767:Ubiquitin-Protein Ligases; D046988:Proteasome Endopeptidase Complex; C493083:SAR1A protein, human; D020559:Monomeric GTP-Binding Proteins