c-Cbl in GPCR Resensitization and Recycling

See allHide authors and affiliations

Science Signaling  19 Jul 2011:
Vol. 4, Issue 182, pp. ec200
DOI: 10.1126/scisignal.4182ec200

As a family, G protein–coupled receptors (GPCRs) are involved in transducing signals relating to biological processes ranging from development to chemokine signaling to neurotransmission. Appropriate trafficking of such receptors after internalization, whether the receptors are degraded or recycled to the plasma membrane, is an important aspect of the spatial and temporal control of GPCR signaling. Baldys and Raymond showed that the recycling of the serotonin (5-HT) 5-hydroxytryptamine 2A receptor (5-HT2AR) involved the C terminus of c-Cbl, an E3 ubiquitin ligase known for its role in the endocytosis and downstream trafficking of receptor tyrosine kinases. Coimmunoprecipitation experiments with lysates from human embryonic kidney 293 (HEK293) cells transiently transfected with v5 epitope–tagged 5-HT2AR (5-HT2AR-v5) revealed an interaction between endogenous c-Cbl and 5-HT2AR-v5. In contrast, coimmunoprecipitation of 5-HT2AR-v5 and c-Cbl was reduced in HEK293 cells cotransfected with 5-HT2AR-v5 and green fluorescent protein (GFP)–tagged c-Cbl C-terminal truncation mutants that either had or lacked ubiquitin ligase activity, suggesting that the role of c-Cbl in trafficking of this receptor was independent from its ubiquitin ligase activity. An internalization assay with HEK293 cells transiently transfected with 5-HT2AR-v5 and treated with serotonin indicated that the internalized receptor localized to early and recycling endosomes but not lysosomes. Small-interfering RNA (siRNA)–mediated knockdown of c-Cbl reduced the recycling of 5-HT2AR-v5 to the plasma membrane and caused the accumulation of the receptor in endosomes, in comparison to HEK293 cells transiently transfected with control siRNA. Similarly, cells cotransfected with 5-HT2AR-v5 and either of the two C-terminal c-Cbl mutants exhibited reduced recycling of 5-HT2AR-v5, as well as a reduction in serotonin-mediated Ca2+ mobilization—a measurement of 5-HT2AR resensitization. Thus, c-Cbl, through a ubiquitin ligase-independent activity, appears to mediate efficient endosomal recycling and resensitization of 5-HT2AR. It will be interesting to determine whether this pathway functions in the trafficking and resensitization of other classes of GPCRs.

A. Baldys, J. R. Raymond, Role of c-Cbl carboxyl terminus in serotonin 5-HT2A receptor recycling and resensitization. J. Biol. Chem. 286, 24656–24665 (2011). [PubMed]

Stay Connected to Science Signaling