Ragulator-Rag Complex Targets mTORC1 to the Lysosomal Surface and Is Necessary for Its Activation by Amino Acids

“For example, in Ragulator null cells, Akt is slightly activated, almost certainly because the well-known inhibitory signal from mTORC1 to PI3K is absent.”

“the Lysosomal Surface and Is Necessary for Its Activation by Amino Acids”

“Because mTORC1 pathway activity is high in cells expressing RagB (Sancak et al., 2008), the reduced Ragulator-Rag interaction in these cells may reflect a compensatory mechanism to reduce mTORC1 activity.”

“the Lysosomal Surface and Is Necessary for Its Activation by Amino Acids”

“the Lysosomal Surface and Is Necessary for Its Activation by Amino Acids”

“On the other hand, mice lacking the core mTORC1 component raptor die earlier (before embryonic day 6.5) than p14 and p18 null mice (Guertin et al., 2006).”

“the Lysosomal Surface and Is Necessary for Its Activation by Amino Acids”

“HEK293T, cells the mTORC1 pathway is strongly regulated by serum and insulin (Sancak et al., 2007).”

“As Akt suppresses MAPK signaling by phosphorylating and inhibiting Raf (Zimmermann and Moelling, 1999), it is conceivable that the activation of Akt that occurs in Ragulator null cells could account, at least in part, for the inhibition of MAPK signaling that has been observed in these cells.”

“…02139, USA 4Department of Oncogene Research, Research Institute for Microbial Diseases, Osaka University, 3-1 Yamadaoka, Suita, Osaka 565-0871, Japan 5Howard Hughes Medical Institute 6These authors contributed equally to this work *Correspondence: sabatini@wi.mit.edu DOI 10.1016/j.cell.2010.02.024”

“Previous work indicates that these three small proteins interact with each other, localize to endosomes and lysosomes, and play positive roles in the MAPK pathway (Lunin et al., 2004; Nada et al., 2009; Schaeffer et al., 1998; Teis et al., 2006; Teis et al., 2002; Wunderlich et al., 2001).”

“the Lysosomal Surface and Is Necessary for Its Activation by Amino Acids”

“Because the Rag GTPases interact with Ragulator and given the function of p18 in localizing MP1 and p14 to lysosomes (Nada et al., 2009), it seemed possible that the Ragulator is necessary for localizing the Rag proteins to the lysosomal surface.”

“RagA and RagB, like RagC and RagD, are highly similar to each other and are functionally redundant (Hirose et al., 1998; Sancak et al., 2008; Schürmann et al., 1995; Sekiguchi et al., 2001).”

“Furthermore, mice engineered to lack either p14 or p18 die around embryonic day 7–8 and exhibit severe growth retardation (Nada et al., 2009; Teis et al., 2006).”

“…02139, USA 4Department of Oncogene Research, Research Institute for Microbial Diseases, Osaka University, 3-1 Yamadaoka, Suita, Osaka 565-0871, Japan 5Howard Hughes Medical Institute 6These authors contributed equally to this work *Correspondence: sabatini@wi.mit.edu DOI 10.1016/j.cell.2010.02.024”

“Previous work indicates that these three small proteins interact with each other, localize to endosomes and lysosomes, and play positive roles in the MAPK pathway (Lunin et al., 2004; Nada et al., 2009; Schaeffer et al., 1998; Teis et al., 2006; Teis et al., 2002; Wunderlich et al., 2001).”

“For technical reasons (Buerger et al., 2006; Sancak et al., 2008), it has not been possible to determine the intracellular localization of endogenous Rheb, and work using overexpressed GFP-tagged Rheb1 has placed it on various endomembrane compartments, including endosomes and lysosomes (Buerger et al.”

“TORC1 and the Gtr proteins are located on the surface of the vacuole (Berchtold and Walther, 2009; Binda et al., 2009), the yeast equivalent of lysosomes, but, unlike in mammals, yeast TORC1 does not leave the vacuolar surface upon amino acid deprivation although amino acids do control the interaction of TORC1 with Gt…”

“As a control, we generated a raptor fusion protein that lacks the CAAX box of the Rheb1 targeting signal (raptor-Rheb15DCAAX) and so cannot associate with membranes (Buerger et al., 2006; Clark et al., 1997; Takahashi et al., 2005).”

“In particular, they do not share any sequence similarity with the Ego1p or Ego3p, proteins, which interact with Gtr1p and Gtr2p (Dubouloz et al., 2005; Gao and Kaiser, 2006), the orthologs of the Rag proteins in budding yeast (Gao and Kaiser, 2006; Schürmann et al., 1995).”

“mTORC1 by amino acids and, in cells lacking TSC2, the mTORC1 pathway is sensitive to amino acid starvation but resistant to growth factor withdrawal (Roccio et al., 2006; Smith et al., 2005).”

“, 2005) and can localize to late endosomes/lysosomes (Saito et al., 2005; Sancak et al., 2008), is consistent with a model in which amino acids induce mTORC1 to associate with the endomembrane system of the cell and thus allow it to encounter its activator Rheb.”

“In particular, they do not share any sequence similarity with the Ego1p or Ego3p, proteins, which interact with Gtr1p and Gtr2p (Dubouloz et al., 2005; Gao and Kaiser, 2006), the orthologs of the Rag proteins in budding yeast (Gao and Kaiser, 2006; Schürmann et al., 1995).”

“Previous studies suggest that MP1-p14-p18 complex plays an adaptor role in the MAP Kinase (MAPK) pathway (reviewed in (Dard and Matthias, 2006)) and our current findings do not contradict these results.”

“, 2008), it has not been possible to determine the intracellular localization of endogenous Rheb, and work using overexpressed GFP-tagged Rheb1 has placed it on various endomembrane compartments, including endosomes and lysosomes (Buerger et al., 2006; Saito et al., 2005; Sancak et al., 2008; Takahashi et al., 2005).”

“…02139, USA 4Department of Oncogene Research, Research Institute for Microbial Diseases, Osaka University, 3-1 Yamadaoka, Suita, Osaka 565-0871, Japan 5Howard Hughes Medical Institute 6These authors contributed equally to this work *Correspondence: sabatini@wi.mit.edu DOI 10.1016/j.cell.2010.02.024”

“Previous work indicates that these three small proteins interact with each other, localize to endosomes and lysosomes, and play positive roles in the MAPK pathway (Lunin et al., 2004; Nada et al., 2009; Schaeffer et al., 1998; Teis et al., 2006; Teis et al., 2002; Wunderlich et al., 2001).”

“…evidence is needed to support this connection because the YY1 response element was not identified as a motif enriched in the promoters of mTORC1-regulated genes (Düvel et al., 2010) and little endogenous mTORC1 is found in the nucleus (Sancak et al., 2010; Sancak et al., 2008; Zoncu et al., 2011).”

“The Rags are obligate heterodimers of RagA or RagB with RagC or RagD and are tethered to the lysosomal membrane through their association with the pentameric Ragulator complex comprising MP1, p14, p18, HBXIP, and C7ORF59 (Sancak et al., 2010; Bar-Peled et al., 2012).”

“The active Rag heterodimer physically interacts with RAPTOR, causing mTORC1 to cluster onto the surface of late endosomes and lysosomes, where the Rag GTPases resid”

“pool (mainly leucine) present within the lumen of the lysosome by the vacuolar H-ATPase (v-ATPase) and a molecular complex called Ragulator (Figure 1).(24,25) This mechanism could underlie the negative feedback acting on autophagy”