Structural basis for the ARF GAP activity and specificity of the C9orf72 complex

Abstract

Mutation of C9ORF72 is the most common genetic cause of amyotrophic lateral sclerosis (ALS) and frontal temporal degeneration (FTD), which is attributed to both a gain and loss of function. C9orf72 forms a complex with SMCR8 and WDR41, which was reported to have GTPase activating protein activity toward ARF proteins, RAB8A, and RAB11A. We determined the cryo-EM structure of ARF1-GDP-BeF3- bound to C9orf72:SMCR8:WDR41. The SMCR8longin and C9orf72longin domains form the binding pocket for ARF1. One face of the C9orf72longin domain holds ARF1 in place, while the SMCR8longin positions the catalytic finger Arg147 in the ARF1 active site. Mutations in interfacial residues of ARF1 and C9orf72 reduced or eliminated GAP activity. RAB8A GAP required ~10-fold higher concentrations of the C9orf72 complex than for ARF1. These data support a specific function for the C9orf72 complex as an ARF GAP.

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

“…A recent cryo-EM structure of the GAP-competent interaction between Rag-Ragulator and GATOR1 suggested GATOR1 is stabilized by both Rag GTPases in its GAP mode (34). The overall geometry and the role of the Arg finger in the active GATOR1 complex resembles that of FLCN F118D :FNIP2 seen here, as well as that of another longin-domain based GAP, the C9orf72-SMCR8 complex with ARF1 (31). In comparison to GATOR1, the structure of the AFC revealed that FLCN solely interacts with RagC to stimulate GTP hydrolysis.…”

“…S6B). We also compared the RagC structure to that of Arf1, which is a substrate of another longin domain GAP, SMCR8 ( 31, 32 ). In the Arf1 model, FLCN F118D (Leu 13 and His 14 ) clash with Arf1 residues (Fig.…”

“…Residues in FLCN at interface that clash are indicated and labelled in red. (B) Model of RagC (pink) and SMCR8 (orange) interface generated by aligning SMCR8 from Arf1:SMCR8 structure (PDB: 7MGE) with FLCN in AFC structure ( 27 ). Residues in SMCR8 at interface that clash are indicated and labelled in red.…”

“…Structural investigation of RagC:SMCR8 and Arf1:FLCN interface. (A) Model of Arf1 (pink) and FLCN (orange) interface generated by aligning Arf1 from Arf1:SMCR8 structure (PDB: 7MGE) with RagC in AFC structure(27). Residues in FLCN at interface that clash are indicated and labelled in red.…”

Implicações de variantes bialélicas de perda de função no gene TBCK, no transporte de vesículas de células progenitoras neurais derivadas de células-tronco pluripotentes induzidas

“…Nesse contexto, foi demonstrado que C9ORF72 se liga a RAB1A-GTP para regular o transporte do complexo ULK1, central para a indução da autofagia, à membrana onde se dará a formação dos autofagossomos (Webster et al, 2016). Já a relevância da interação com ARF1 para a autofagia não é clara, uma vez que C9ORF72 nesse caso, parece atuar em conjunto com SMCR8, para inativar ARF1 (Su et al, 2021).…”