The Endosomal Sorting Complex Required for Transport Pathway Mediates Chemokine Receptor CXCR4-promoted Lysosomal Degradation of the Mammalian Target of Rapamycin Antagonist DEPTOR

Rita Verma,Adriano Marchese

doi:10.1074/jbc.m114.606699

Rita Verma, Adriano Marchese

Open Access

https://doi.org/10.1074/jbc.m114.606699

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Abstract

G protein-coupled receptor (GPCR) signaling mediates many cellular functions, including cell survival, proliferation, and cell motility. Many of these processes are mediated by GPCR-promoted activation of Akt signaling by mammalian target of rapamycin complex 2 (mTORC2) and the phosphatidylinositol 3-kinase (PI3K)/phosphoinositide-dependent kinase 1 (PDK1) pathway. However, the molecular mechanisms by which GPCRs govern Akt activation by these kinases remain poorly understood. Here, we show that the endosomal sorting complex required for transport (ESCRT) pathway mediates Akt signaling promoted by the chemokine receptor CXCR4. Pharmacological inhibition of heterotrimeric G protein Gαi or PI3K signaling and siRNA targeting ESCRTs blocks CXCR4-promoted degradation of DEPTOR, an endogenous antagonist of mTORC2 activity. Depletion of ESCRTs by siRNA leads to increased levels of DEPTOR and attenuated CXCR4-promoted Akt activation and signaling, consistent with decreased mTORC2 activity. In addition, ESCRTs likely have a broad role in Akt signaling because ESCRT depletion also attenuates receptor tyrosine kinase-promoted Akt activation and signaling. Our data reveal a novel role for the ESCRT pathway in promoting intracellular signaling, which may begin to identify the signal transduction pathways that are important in the physiological roles of ESCRTs and Akt.

Highlights

Akt signaling is activated by G protein-coupled receptors (GPCRs) via the phosphatidylinositol 3kinase (PI3K)-mammalian target of rapamycin complex 2 (mTORC2) signaling cascade, but mechanistic insight is lacking
Depletion of endosomal sorting complex required for transport (ESCRT) leads to elevated levels of DEPTOR (Fig. 2D), and because DEPTOR is a natural endogenous antagonist of mTOR kinase activity [17], this likely negatively impacts mTORC2 activity and thereby inhibits Akt activation and signaling promoted by GPCRs (Figs. 4, A–D, and 8A) and receptor tyrosine kinase (RTK) (Fig. 8, B and C)
Acute CXC chemokine receptor 4 (CXCR4) (Ͻ30 min) activation promotes DEPTOR degradation, and this requires G protein and PI3K signaling (Fig. 10A). This suggests that Akt signaling is linked to DEPTOR degradation, but whether this is linked to activation of mTORC2 remains to be clearly defined

Summary

Background

Akt signaling is activated by G protein-coupled receptors (GPCRs) via the PI3K-mTORC2 signaling cascade, but mechanistic insight is lacking. Results: GPCR signaling promotes lysosomal and ESCRT-dependent degradation of DEPTOR, an antagonist of mTORC2, that regulates Akt activation and signaling. G protein-coupled receptor (GPCR) signaling mediates many cellular functions, including cell survival, proliferation, and cell motility Many of these processes are mediated by GPCR-promoted activation of Akt signaling by mammalian target of rapamycin complex 2 (mTORC2) and the phosphatidylinositol 3kinase (PI3K)/phosphoinositide-dependent kinase 1 (PDK1) pathway. Pharmacological inhibition of heterotrimeric G protein G␣i or PI3K signaling and siRNA targeting ESCRTs blocks CXCR4-promoted degradation of DEPTOR, an endogenous antagonist of mTORC2 activity. Glycogen synthase kinase 3␤ (GSK-3␤), which serves as an endogenous antagonist of the canonical Wnt/␤-catenin signaling pathway, is targeted into ILVs by ESCRTs. GSK-3␤ is normally active and free in the cytoplasm to phosphorylate ␤-catenin, thereby promoting its proteasomal degradation (34 –36). Our findings indicate that ESCRTs regulate receptorpromoted DEPTOR degradation and Akt signaling

EXPERIMENTAL PROCEDURES

RESULTS

47.8 DEPTOR

50 IP: myc RAPTOR

DISCUSSION