Smoothened (Smo), a GPCR family protein, plays a critical role in the reception and transduction of Hedgehog (Hh) signal. Smo is phosphorylated and activated on the cell surface; however, it is unknown whether Smo can be intracellularly activated. Here, we demonstrate that inactivation of the ESCRT-III causes dramatic accumulation of Smo in the ESCRT-III/MVB compartment, and subsequent activation of Hh signaling. In contrast, inactivation of ESCRTs 0–II induces mild Smo accumulation in the ESCRT-III/MVB compartment. We provide evidence that Kurtz (Krz), the Drosophila β-arrestin2, acts in parallel with the ESCRTs 0-II pathway to sort Smo to the multivesicular bodies and lysosome-mediated degradation. Additionally, upon inactivation of ESCRT-III, all active and inactive forms of Smo are accumulated. Endogenous Smo accumulated upon ESCRT-III inactivation is highly activated, which is induced by phosphorylation but not sumoylation. Taken together, our findings demonstrate a model for intracellular activation of Smo, raising the possibility for tissue overgrowth caused by an excessive amount, rather than mutation of Smo.
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This study was supported by National Institutes of Health (grant R01 GM079684 and pilot grant P30 CA177558) to J.J. We also thank the Shared Resource Facilities of the University of Kentucky Markey Cancer Center (P30 CA177558), and acknowledge the Imaging Core of the COBRE (P20 GM121327).
Supplementary information available online at http://jcs.biologists.org/lookup/doi/10.1242/jcs.211367.supplemental.
Jiang, Kai; Liu, Yajuan; Zhang, Jie; and Jia, Jianhang, "An Intracellular Activation of Smoothened that is Independent of Hedgehog Stimulation in Drosophila" (2018). Markey Cancer Center Faculty Publications. 117.