Structural basis for FLCN RagC GAP activation in MiT-TFE substrate-selective mTORC1 regulation

Abstract

The mechanistic target of rapamycin complex 1 (mTORC1) regulates cell growth and catabolism in response to nutrients through phosphorylation of key substrates. The tumor suppressor folliculin (FLCN) is a RagC/D guanosine triphosphatase (GTPase)–activating protein (GAP) that regulates mTORC1 phosphorylation of MiT-TFE transcription factors, controlling lysosome biogenesis and autophagy. We determined the cryo–electron microscopy structure of the active FLCN complex (AFC) containing FLCN, FNIP2, the N-terminal tail of SLC38A9, the RagA GDP :RagC GDP.BeFx- GTPase dimer, and the Ragulator scaffold. Relative to the inactive lysosomal FLCN complex structure, FLCN reorients by 90°, breaks contact with RagA, and makes previously unseen contacts with RagC that position its Arg 164 finger for catalysis. Disruption of the AFC-specific interfaces of FLCN and FNIP2 with RagC eliminated GAP activity and led to nuclear retention of TFE3, with no effect on mTORC1 substrates S6K or 4E-BP1. The structure provides a basis for regulation of an mTORC1 substrate-specific pathway and a roadmap to discover MiT-TFE family selective mTORC1 antagonists.

Sex differences in response to obesity and caloric restriction on cognition and hippocampal markers of autophagic-lysosomal pathway function

“…Atg12 was signi cantly upregulated in Obese males, regardless of CR treatment, but also ubiquitously decreased by CR. A similar effect of obesity was detected in Slc38a9, an amino acid (arginine) sensor on lysosomes which aides in mTORC1 regulation [50], and disassembly of the lysosomal-folliculin complex allowing folliculin GAP activation of RagC [57]. CR also had some malespeci c effects.…”

“…Substantial evidence links folliculin, FNIP2 and TFE3; however, these effects vary greatly based on tissue type, and neither have, currently, been characterized in the brain. Manipulation of folliculin and FNIP2 active sites prevents GAP activity leading to nuclear retention of TFE3 [50]. In cancer tissues, loss of folliculin leads to greater TFE3 activation and induction of related pathways [51].…”

TFEB-mediated lysosomal exocytosis alleviates high-fat diet–induced lipotoxicity in the kidney

“…Indeed, PA-induced TFEB nuclear translocation was inhibited by the overexpression of a constitutively active form of RagC (pRK5-HA GST RagC 75 L), while Torin 1 translocated TFEB into the nucleus even in the presence of active RagC ( Figure 2C ). To further explore the possible mechanism of PA-induced TFEB nuclear translocation, we focused on folliculin (FLCN), which is an amino acid–dependent GTPase-activating protein (GAP) for RagC/D ( 27 ). As expected, amino acid stimulation after starvation increased TFEB phosphorylation in BSA-treated PTECs but not in PA-treated PTECs, whereas S6RP and 4E-BP1 were similarly phosphorylated by amino acid stimulation in both BSA- and PA-treated PTECs ( Figure 2D ).…”

Structure of the Commander endosomal trafficking complex linked to X-linked intellectual disability/Ritscher-Schinzel syndrome

“…It is localised to late endosomes and its knockdown perturbs aspects of endosomal morphology and function (21, 75), although it appears dispensable for assembly of other Commander subunits (21). To date only one crystal structure of a DENN domain in complex with a RAB GEF substrate has been determined, DENDD1B with RAB35 (74), while the only other structures known are Longin and DENN domain-containing dimers of C9orf72/SMCR8 and Folliculin/FNIP2 that actually act as RAB GAP complexes (100–104). DENND10 is distinct from all of these, as it lacks key residues found in DENND1B required for GEF activity, and the interface with CCDC22-CCDC93 would preclude RAB binding in the same region.…”