The E3 ubiquitin ligase ZNRF2 is a substrate of mTORC1 and regulates its activation by amino acids

Abstract

The mechanistic Target of Rapamycin complex 1 (mTORC1) senses intracellular amino acid levels through an intricate machinery, which includes the Rag GTPases, Ragulator and vacuolar ATPase (V-ATPase). The membrane-associated E3 ubiquitin ligase ZNRF2 is released into the cytosol upon its phosphorylation by Akt. In this study, we show that ZNRF2 interacts with mTOR on membranes, promoting the amino acid-stimulated translocation of mTORC1 to lysosomes and its activation in human cells. ZNRF2 also interacts with the V-ATPase and preserves lysosomal acidity. Moreover, knockdown of ZNRF2 decreases cell size and cell proliferation. Upon growth factor and amino acid stimulation, mTORC1 phosphorylates ZNRF2 on Ser145, and this phosphosite is dephosphorylated by protein phosphatase 6. Ser145 phosphorylation stimulates vesicle-to-cytosol translocation of ZNRF2 and forms a novel negative feedback on mTORC1. Our findings uncover ZNRF2 as a component of the amino acid sensing machinery that acts upstream of Rag-GTPases and the V-ATPase to activate mTORC1.DOI: http://dx.doi.org/10.7554/eLife.12278.001

A semi-supervised Bayesian approach for simultaneous protein sub-cellular localisation assignment and novelty detection

“…We observe a mixed steady-state distribution between the endosome and PM for this protein (gure 6, panel d). In support of this result, we nd that ZNRF2 has been detected on the endosomes, lysosomes, Golgi apparatus and PM according to the literature (Araki and Milbrandt, 2003;Hoxhaj et al, 2016). There is no information in regard to the sub-cellular location of ZNRF2 in the HPA Cell Atlas database.…”

“…This protein has been found to control the sub-cellular localisation and biological function of mTORC1, the V-ATPase and the Na+/K+-ATPase α1 (Hoxhaj et al, 2012(Hoxhaj et al, , 2016. ZNRF2 is membrane-associated but can be released into the cytosol upon phosphorylation by various kinases (Hoxhaj et al, 2016). We observe a mixed steady-state distribution between the endosome and PM for this protein (gure 6, panel d).…”

“…Notably, it has been demonstrated that KIF16B co-localises with the small GTPase Rab5, whose isoforms Rab5a and Rab5b we also identied as potentially localised to the endosome and PM in this dataset. ZNRF2 (Q8NHG8) is an E3 ubiquitin ligase which has been shown to regulate mTOR signalling as well as lysosomal acidity and homeostasis in mouse and human cells and has been detected at the endosomes, lysosomes, Golgi apparatus and PM according to the literature (Araki and Milbrandt, 2003;Hoxhaj et al, 2016). Ykt6 (O15498) is a SNARE (soluble N-ethylmaleimide-sensitive factor attachment protein receptor) protein that regulates a wide variety of intracellular tracking and membrane tethering and fusion processes.…”

“…Fourth, Q8NHG8 (ZNRF2) is an E3 ubiquitin ligase which has been shown to regulate mTOR signalling as well as lysosomal acidity and homeostasis in mouse and human cells (Hoxhaj et al, 2016). This protein has been found to control the sub-cellular localisation and biological function of mTORC1, the V-ATPase and the Na+/K+-ATPase α1 (Hoxhaj et al, 2012(Hoxhaj et al, , 2016. ZNRF2 is membrane-associated but can be released into the cytosol upon phosphorylation by various kinases (Hoxhaj et al, 2016).…”

ANKRD44 Gene Silencing: A Putative Role in Trastuzumab Resistance in Her2-Like Breast Cancer

“…Interestingly, the activation of the mTOR pathway and the overexpression of LDHB has already been described in the literature (48) and could be associated with an impairment of PP6 activity due to ANKRD44 silencing. In fact, PP6 activity is strictly associated with mTOR signaling pathway by regulating ZNRF2 and eIF2α factors, regulators of mTORC1 activity (49, 50). LDHB is extensively described in the literature for its role in promoting tumorigenesis in colorectal cancer (51) and also for its involvement in the metastatic profile (52).…”