Hybrid Structure of the RagA/C-Ragulator mTORC1 Activation Complex

Su, M.; Morris, Kyle K.; Kim, Dojin; Fu, Yangxue; Lawrence, Rosalie; Stjepanović, Goran; Zoncu, Roberto; Hurley, James H.

doi:10.1016/j.bpj.2017.11.193

Cited by 5 publications

(8 citation statements)

References 45 publications

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“…Our results confirm the essential role of HBXIP-C7orf59 dimer as an integral part of pentameric Ragulator and support a sequential model of Ragulator assembly in which HBXIP-C7orf59 stabilizes p18 and allows subsequent binding of MP1-p14. These findings are in agreement with the recently reported structural studies of pentameric Ragulator [20][21][22][23][24].…”

supporting

confidence: 93%

“…The p18 regions assigned here as responsible for stable binding to HBXIP-C7orf59 (residues: 130-149) and MP1-p14 (residues: 80-108) by truncation and pulldown analysis are found associated with HBXIP and MP1, respectively, in the structure of the pentamer [20][21][22][23][24]. The two p18-derived peptides which copurified with HBXIP-C7orf59 dimer obtained from dissociation of the trimer are part of the interface with C7orf59 in the pentamer structure.…”

Section: Discussionmentioning

confidence: 74%

“…Comparison of the apo HBXIP-C7orf59 structure solved in this study with the HBXIP-C7orf59 dimer present in the pentameric Ragulator [20][21][22][23][24] demonstrates that the unfolded N terminus of C7orf59 (residues: 1-15) undergoes a structural rearrangement upon p18 binding, adopting a helical structure similar to the N terminus of other Roadblock subunits. Although the lack of electron density in the N-terminal region of C7orf59 in other structures of unbound C7orf59-HBXIP dimer [21,23] and HDX analysis [20] suggested that this region is unfolded, our structure is the first to capture the full N terminus of C7orf59 in an extended conformation due to its unique crystal packing. The strict requirement for the N terminus of C7orf59 for binding to p18, demonstrated by deletion of this region, highlights the importance of its flexible nature.…”

Section: Discussionmentioning

confidence: 80%

“…The overall structure and dimer interface of HBXIP-C7orf59 are similar to previously reported structures [21,23] except for the presence of defined electron density corresponding to the unfolded N-terminal region (residues: 1-15) in C7orf59 ( Figs 1A,B and S1). Comparison with the pentameric Ragulator complex [20][21][22][23][24] shows that the N terminus of C7orf59 undergoes a transition to alpha-helical conformation upon binding of p18. Strikingly, deletion of residues 1-15 of C7orf59 severely impairs the dimer's interaction with p18, highlighting its importance in stabilizing the complex ( Figs 1C and S2D).…”

Section: Resultsmentioning

confidence: 99%

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C7orf59/LAMTOR4 phosphorylation and structural flexibility modulate Ragulator assembly

et al. 2019

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Ragulator is a pentamer composed of p18, MP1, p14, C7orf59, and hepatitis B virus X‐interacting protein (HBXIP; LAMTOR 1—5) which acts as a lysosomal scaffold of the Rag GTPases in the amino acid sensitive branch of TORC1 signaling. Here, we present the crystal structure of human HBXIP‐C7orf59 dimer (LAMTOR 4/5) at 2.9 Å and identify a phosphorylation site on C7orf59 which modulates its interaction with p18. Additionally, we demonstrate the requirement of HBXIP‐C7orf59 to stabilize p18 and allow further binding of MP1‐p14. The structure of the dimer revealed an unfolded N terminus in C7orf59 (residues 1–15) which was shown to be essential for p18 binding. Full‐length p18 does not interact stably with MP1‐p14 in the absence of HBXIP‐C7orf59, but deletion of p18 residues 108–161 rescues MP1‐p14 binding. C7orf59 was phosphorylated by protein kinase A (PKA) in vitro and mutation of the conserved Ser67 residue to aspartate prevented phosphorylation and negatively affected the C7orf59 interaction with p18 both in cell culture and in vitro . C7orf59 Ser67 was phosphorylated in human embryonic kidney 293T cells. PKA activation with forskolin induced dissociation of p18 from C7orf59, which was prevented by the PKA inhibitor H‐89. Our results highlight the essential role of HBXIP‐C7orf59 dimer as a nucleator of pentameric Ragulator and support a sequential model of Ragulator assembly in which HBXIP‐C7orf59 binds and stabilizes p18 which allows subsequent binding of MP1‐p14.

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supporting

confidence: 93%

Section: Discussionmentioning

confidence: 74%

Section: Discussionmentioning

confidence: 80%

Section: Resultsmentioning

confidence: 99%

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C7orf59/LAMTOR4 phosphorylation and structural flexibility modulate Ragulator assembly

et al. 2019

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“…The lysosomal platform upon which mTORC1 activation occurs is known as the Ragulator, a protein complex composed of the proteins MP1, p14, HBXIP1, C7ORF59 and p18 (encoded by MAPKSP1, ROBLD3, HBXIP, c7orf59 and c11orf59, respectively), and is anchored to the lysosome by p18. [46][47][48] The Ragulator, which is orthologous to the yeast EGO complex, 49,50 tethers the Rag GTPases to the lysosomal surface and has guanine nucleotide exchange factor (GEF) activity for two of the Rag proteins, RagA and RagB, 47,51 with the Gdomains of the Rag proteins projecting away from the Ragulator surface. [51][52][53][54] Control of Ragulator activity is one mechanism by which amino acids regulate mTORC1.…”

Section: Regulators Of the Rag Gtpasesmentioning

confidence: 99%

Lysosome: The metabolic signaling hub

Lamming

Bar-Peled

2018

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127

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For the past five decades the lysosome has been characterized as an unglamorous cellular recycling center. This notion has undergone a radical shift in the last ten years, with new research revealing that this organelle serves as a major hub for metabolic signaling pathways. The discovery that master growth regulators, including the protein kinase mTOR (mechanistic Target Of Rapamycin) make their home at the lysosomal surface has generated intense interest in the lysosome’s key role in nutrient sensing and cellular homeostasis. The transcriptional networks required for lysosomal maintenance and function are just being unraveled and their connection to lysosome-based signaling pathways revealed. The catabolic and anabolic pathways that converge on the lysosome connect this organelle with multiple facets of cellular function; when these pathways are deregulated they underlie multiple human diseases, and promote cellular and organismal aging. Thus, understanding how lysosome-based signaling pathways function will not only illuminate the fascinating biology of this organelle but will also be critical in unlocking its therapeutic potentials.

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A conformational change in the N terminus of SLC38A9 signals mTORC1 activation

Lei

Hattne

et al. 2021

Structure

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mTORC1 is a central hub that integrates environmental cues, such as cellular stresses and nutrient availability to modulate metabolism and cellular responses. Recently, SLC38A9, a lysosomal amino acid transporter, emerged as a sensor for luminal arginine and as an activator of mTORC1. The amino acid-mediated activation of mTORC1 is regulated by the N-terminal domain of SLC38A9. Here, we determined the crystal structure of zebrafish SLC38A9 (drSLC38A9) and found the N-terminal fragment inserted deep within the transporter, bound in the substrate-binding pocket where normally arginine would bind. This represents a significant conformational change of the N-terminal domain (N-plug) when compared with our recent arginine-bound structure of drSLC38A9. We propose a ball-and-chain model for mTORC1 activation, where N-plug insertion and Rag GTPase binding with SLC38A9 is regulated by luminal arginine levels. This work provides important insights into nutrient sensing by SLC38A9 to activate the mTORC1 pathways in response to dietary amino acids.

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Hybrid Structure of the RagA/C-Ragulator mTORC1 Activation Complex

Cited by 5 publications

References 45 publications

C7orf59/LAMTOR4 phosphorylation and structural flexibility modulate Ragulator assembly

C7orf59/LAMTOR4 phosphorylation and structural flexibility modulate Ragulator assembly

Lysosome: The metabolic signaling hub

A conformational change in the N terminus of SLC38A9 signals mTORC1 activation

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