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Ubiquitin‐specific peptidase 7 (USP7) and USP10 mediate deubiquitination of human NHE3 regulating its expression and activity
- Source :
- FASEB J
- Publication Year :
- 2020
- Publisher :
- Wiley, 2020.
-
Abstract
- Na(+)/H(+) exchanger NHE3 of human or primates differs from NHE3 of other animals by having a PY motif, which mediates interaction with the E3 ubiquitin (Ub) ligase Nedd4–2. Ub-conjugation of human NHE3 by Nedd4–2 regulates endocytosis of NHE3 but does not affect its cellular expression. Because Ub-conjugation is a reversible process, the aim of this study is to identify deubiquitinating enzyme (DUB) regulating the post-endosomal fate of human NHE3. Using an activity-based chemical screening, we identified ubiquitin specific protease-7 (USP7) and USP10 that bind NHE3. The roles of DUBs in regulation of NHE3 were studied by determining the effects of silencing of USP7 and USP10. Knockdown of USP7 or USP10 resulted in increased NHE3 ubiquitination and decreased NHE3 expression at the surface membrane and cellular level. The endocytic retrieval of NHE3 was promoted by depletion of USP7 or USP10, with increased association of NHE3 with Rab5a and Rab7. Inhibition of USP7 and USP10 by chemical inhibitors or knockdown had an additive effect on NHE3. In addition, NHE3 half-life was reduced accounting for decreased NHE3 protein abundance. NHE3 is inhibited by protein kinase A. Activation of PKA by forskolin decreased the binding of USP7 and USP10 to NHE3, while increasing ubiquitination of NHE3. Knockdown of USP10 had an additive effect on PKA-dependent inhibition of NHE3. These findings demonstrate that USP7 and USP10 are DUBs that regulate NHE3 ubiquitination and expression, and reveal a new mechanism of NHE3 inhibition involving DUBs.
- Subjects :
- 0301 basic medicine
Endocytic cycle
Endocytosis
Biochemistry
Article
Cell Line
Deubiquitinating enzyme
Ubiquitin-Specific Peptidase 7
03 medical and health sciences
0302 clinical medicine
Ubiquitin
Cell Line, Tumor
Genetics
Humans
Gene silencing
Protein kinase A
Molecular Biology
Gene knockdown
biology
Sodium-Hydrogen Exchanger 3
urogenital system
Chemistry
Ubiquitination
Cell biology
030104 developmental biology
biology.protein
Caco-2 Cells
Ubiquitin Thiolesterase
030217 neurology & neurosurgery
Biotechnology
Deubiquitination
Subjects
Details
- ISSN :
- 15306860 and 08926638
- Volume :
- 34
- Database :
- OpenAIRE
- Journal :
- The FASEB Journal
- Accession number :
- edsair.doi.dedup.....cdc76c0ce71bcc9cf7c283fa80e5148c