ABSTRACT
Ras GTPase-activating protein-binding proteins 1 and 2 (G3BP1 and G3BP2, respectively) are widely recognized as core components of stress granules (SGs). We report that G3BPs reside at the cytoplasmic surface of lysosomes. They act in a non-redundant manner to anchor the tuberous sclerosis complex (TSC) protein complex to lysosomes and suppress activation of the metabolic master regulator mechanistic target of rapamycin complex 1 (mTORC1) by amino acids and insulin. Like the TSC complex, G3BP1 deficiency elicits phenotypes related to mTORC1 hyperactivity. In the context of tumors, low G3BP1 levels enhance mTORC1-driven breast cancer cell motility and correlate with adverse outcomes in patients. Furthermore, G3bp1 inhibition in zebrafish disturbs neuronal development and function, leading to white matter heterotopia and neuronal hyperactivity. Thus, G3BPs are not only core components of SGs but also a key element of lysosomal TSC-mTORC1 signaling.
Subject(s)
Adaptor Proteins, Signal Transducing/metabolism , DNA Helicases/metabolism , Lysosomes/metabolism , Mechanistic Target of Rapamycin Complex 1/metabolism , Poly-ADP-Ribose Binding Proteins/metabolism , RNA Helicases/metabolism , RNA Recognition Motif Proteins/metabolism , RNA-Binding Proteins/metabolism , Signal Transduction , Tuberous Sclerosis/metabolism , Amino Acid Sequence , Animals , Breast Neoplasms/metabolism , Breast Neoplasms/pathology , Cell Line, Tumor , Cell Movement/drug effects , Cytoplasmic Granules/drug effects , Cytoplasmic Granules/metabolism , DNA Helicases/chemistry , Evolution, Molecular , Female , Humans , Insulin/pharmacology , Lysosomal Membrane Proteins/metabolism , Lysosomes/drug effects , Neurons/drug effects , Neurons/metabolism , Phenotype , Poly-ADP-Ribose Binding Proteins/chemistry , RNA Helicases/chemistry , RNA Recognition Motif Proteins/chemistry , Rats, Wistar , Signal Transduction/drug effects , Zebrafish/metabolismABSTRACT
Wnt/ß-catenin signaling controls development and adult tissue homeostasis by regulating cell proliferation and cell fate decisions. Wnt binding to its receptors Frizzled (FZD) and low-density lipoprotein-related 6 (LRP6) at the cell surface initiates a signaling cascade that leads to the transcription of Wnt target genes. Upon Wnt binding, the receptors assemble into large complexes called signalosomes that provide a platform for interactions with downstream effector proteins. The molecular basis of signalosome formation and regulation remains elusive, largely due to the lack of tools to analyze its endogenous components. Here, we use internally tagged Wnt3a proteins to isolate and characterize activated, endogenous Wnt receptor complexes by mass spectrometry-based proteomics. We identify the single-span membrane protein TMEM59 as an interactor of FZD and LRP6 and a positive regulator of Wnt signaling. Mechanistically, TMEM59 promotes the formation of multimeric Wnt-FZD assemblies via intramembrane interactions. Subsequently, these Wnt-FZD-TMEM59 clusters merge with LRP6 to form mature Wnt signalosomes. We conclude that the assembly of multiprotein Wnt signalosomes proceeds along well-ordered steps that involve regulated intramembrane interactions.
