The deubiquitinating enzyme USP10 regulates the post-endocytic sorting of cystic fibrosis transmembrane conductance regulator in airway epithelial cells.
The cystic fibrosis transmembrane conductance regulator (CFTR), a member of the ABC transporter superfamily, is a cyclic AMP-regulated chloride channel and a regulator of other ion channels and transporters. In epithelial cells CFTR is rapidly endocytosed from the apical plasma membrane and efficiently recycles back to the plasma membrane. Because ubiquitination targets endocytosed CFTR for degradation in the lysosome, deubiquitinating enzymes (DUBs) are likely to facilitate CFTR recycling. Accordingly, the aim of this study was to identify DUBs that regulate the post-endocytic sorting of CFTR. Using an activity-based chemical screen to identify active DUBs in human airway epithelial cells, we demonstrated that Ubiquitin Specific Protease-10 (USP10) is located in early endosomes and regulates the deubiquitination of CFTR and its trafficking in the post-endocytic compartment. small interference RNA-mediated knockdown of USP10 increased the amount of ubiquitinated CFTR and its degradation in lysosomes, and reduced both apical membrane CFTR and CFTR-mediated chloride secretion. Moreover, a dominant negative USP10 (USP10-C424A) increased the amount of ubiquitinated CFTR and its degradation, whereas overexpression of wt-USP10 decreased the amount of ubiquitinated CFTR and increased the abundance of CFTR. These studies demonstrate a novel function for USP10 in facilitating the deubiquitination of CFTR in early endosomes and thereby enhancing the endocytic recycling of CFTR.
Bomberger
Jennifer M
JM
Department of Physiology, Dartmouth Medical School, Hanover, New Hampshire 03755, USA. jbomberger@dartmouth.edu
Barnaby
Roxanna L
RL
Stanton
Bruce A
BA
D002462:Cell Membrane / Q000378:metabolism; D002478:Cells, Cultured; D019005:Cystic Fibrosis Transmembrane Conductance Regulator / Q000235:genetics* / Q000502:physiology*; D004305:Dose-Response Relationship, Drug; D004705:Endocytosis; D011992:Endosomes / Q000378:metabolism; D004847:Epithelial Cells / Q000166:cytology; D006801:Humans; D008247:Lysosomes / Q000737:chemistry / Q000378:metabolism; D018613:Microscopy, Confocal / Q000379:methods; D021381:Protein Transport; D034741:RNA, Small Interfering / Q000378:metabolism; D014162:Transfection; D025801:Ubiquitin / Q000737:chemistry; D043222:Ubiquitin Thiolesterase / Q000235:genetics* / Q000502:physiology*
C505032:CFTR protein, human; D034741:RNA, Small Interfering; C431363:USP10 protein, human; D025801:Ubiquitin; D019005:Cystic Fibrosis Transmembrane Conductance Regulator; D043222:Ubiquitin Thiolesterase