Syntaxin 6 and CAL mediate the degradation of the cystic fibrosis transmembrane conductance regulator.
The PDZ domain-containing protein CAL mediates lysosomal trafficking and degradation of CFTR. Here we demonstrate the involvement of a CAL-binding SNARE protein syntaxin 6 (STX6) in this process. Overexpression of STX6, which colocalizes and coimmunoprecipitates with CAL, dramatically reduces the steady-state level and stability of CFTR. Conversely, overexpression of a STX6 dominant-negative mutant increases CFTR. Silencing endogenous STX6 increases CFTR but has no effect on DeltaTRL-CFTR, which cannot bind to CAL. Silencing CAL eliminates the effect of STX6 on CFTR. Both results suggest a dependence of CAL on STX6 function. Consistent with its Golgi localization, STX6 does not bind to ER-localized DeltaF508-CFTR. Silencing STX6 has no effect on DeltaF508-CFTR expression. However, overexpression of STX6 coimmunoprecipitates with and reduces temperature-rescued DeltaF508-CFTR that escapes ER degradation. Conversely, silencing STX6 enhances the effect of low temperature in rescuing DeltaF508-CFTR. Finally, in human bronchial epithelial cells, silencing endogenous STX6 leads to increases in protein levels and Cl(-) currents of both wild-type and temperature-rescued CFTR. We have identified STX6 as a new component of the CAL complex that regulates the abundance and function of CFTR at the post-ER level. Our results suggest a therapeutic role of STX6 in enhancing rescued DeltaF508-CFTR.
Department of Physiology, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA.
Jie
J
Cheng
Valeriu
V
Cebotaru
Liudmila
L
Cebotaru
William B
WB
Guggino
D048868:Adaptor Proteins, Signal Transducing; D002352:Carrier Proteins / Q000502:physiology*; D002460:Cell Line; D019005:Cystic Fibrosis Transmembrane Conductance Regulator / Q000235:genetics / Q000378:metabolism*; D005786:Gene Expression Regulation; D020868:Gene Silencing; D005799:Genes, Dominant; D000077614:Golgi Matrix Proteins; D006801:Humans; D008565:Membrane Proteins / Q000502:physiology*; D026901:Membrane Transport Proteins; D018613:Microscopy, Confocal / Q000379:methods; D008954:Models, Biological; D009154:Mutation; D010957:Plasmids / Q000378:metabolism; D050765:Qa-SNARE Proteins / Q000502:physiology*; D034741:RNA, Small Interfering / Q000378:metabolism
D048868:Adaptor Proteins, Signal Transducing; D002352:Carrier Proteins; C434484:GOPC protein, human; D000077614:Golgi Matrix Proteins; D008565:Membrane Proteins; D026901:Membrane Transport Proteins; D050765:Qa-SNARE Proteins; D034741:RNA, Small Interfering; C093349:cystic fibrosis transmembrane conductance regulator delta F508; D019005:Cystic Fibrosis Transmembrane Conductance Regulator