The J Domain of Sacsin Disrupts Intermediate Filament Assembly

Abstract

Autosomal Recessive Spastic Ataxia of the Charlevoix Saguenay (ARSACS) is caused by mutation in the SACS gene resulting in loss of function of the protein sacsin. A key feature is the formation of abnormal bundles of neurofilaments (NF) in neurons and vimentin intermediate filaments (IF) in cultured fibroblasts, suggesting a role of sacsin in IF homeostasis. Sacsin contains a J domain (SacsJ) homologous to Hsp40, that can interact with Hsp70 chaperones. The SacsJ domain resolved NF bundles in cultured Sacs−/− neurons. Having studied the mechanism using NF assembled in vitro from purified NF proteins, we report that the SacsJ domain interacts with NF proteins to disassemble NFL filaments, and to inhibit their initial assembly. A cell-penetrating peptide derived from this domain, SacsJ-myc-TAT was efficient in disassembling NF bundles in cultured Sacs−/− motor neurons, restoring the NF network; however, there was some loss of vimentin IF and NF in cultured Sacs+/+ fibroblasts and motor neurons, respectively. These results suggest that sacsin through its SacsJ domain is a key regulator of NF and vimentin IF networks in cells.

Neurofilaments in health and Charcot-Marie-Tooth disease

“…While this supports the idea that the intrinsic nature of NF polypeptides can drive assembly, this does not exclude that cofactors can regulate their assembly in vivo ( Chernyatina et al, 2015 ; Herrmann and Aebi, 2016 ). As examples, Sacsin (mutated in the ataxia ARSACs presenting NF aggregation), NUDEL and the 14-3-3 protein have been shown to bind to NF-L and regulate the assembly/disassembly state of NFs ( Nguyen et al, 2004 ; Miao et al, 2013 ; Dabbaghizadeh et al, 2022 ). Investigation of the intrinsic capacities of single NF subunits to polymerise revealed that only NF-L, peripherin and α-internexin are able to form homopolymeric filaments in vitro ( Ching and Liem, 1993 ; Cui et al, 1995 ; Carter et al, 1998 ).…”

Interactors of sacsin’s DNAJ domain identify function in organellar transport and membrane composition relevant to ARSACS pathogenesis

“…Recently, using STRING network analysis, sacsin was shown to interact with multiple heat-shock protein (HSP) chaperones, and to be a key element for trafficking to the plasma membrane of alpha integrins, which were sequestered into IF bundles [6]. We recently demonstrated direct interaction between IF and the sacsin J domain [21]; however, DNAJ domains typically have multiple clients [22]. Hence, we aimed to identify the SacsJ interactome to better understand the functions of sacsin and key elements of ARSACS pathogenic cascade.…”

Subcellular, biochemical and biophysical alterations in two glial cell models of ARSACS

“…lipofuscin aggregates) in skin biopsies (Stevens et al, 2013), and ARSACS-derived fibroblasts show important cellular and molecular alteration (Bradshaw et al, 2016), suggesting that sacsin biological roles go beyond the central nervous system. Disruption of intermediate filament (IF) cytoskeleton and of organelle distribution are largely the most common hallmarks in all cellular and animal models of ARSACS (Dabbaghizadeh et al, 2022;Gentil et al, 2018;Murtinheira et al, 2022), including nerve and glial cells. IFs play an important role in maintaining the mechanical and viscoelastic properties of cells (Sivaramakrishnan et al, 2008), which are important for a wide range of cellular functions, such as cell migration, division, and differentiation.…”