The Role of HSP70 and Its Co-chaperones in Protein Misfolding, Aggregation and Disease - The Networking of Chaperones by Co-chaperones - page 260

protein were identified as causing ARSACS in a Canadian population, ARSACS
patients have now been identified worldwide with cases in Europe, North Africa,
Turkey, Japan and Brazil (Bouhlal et al. 2011 ; Pyle et al. 2012 ). Patients outside of
Canada display considerable phenotypic heterogeneity and a later disease onset is
more common (Pyle et al. 2012 ; Synofzik et al. 2013 ). The SACS gene was initially
thought to be encoded by a single gigantic exon (Engert et al. 2000 ), but a further
eight coding exons and a tenth non-coding exon have been identified upstream of
this, forming a 13,737 bp open reading frame (Ouyang et al. 2006 ; Vermeer et al.
2008 ). SACS encodes the protein sacsin (SACS), a multimodular protein of 4579
amino acids (520 kDa), one of the largest known proteins in the human genome.
SACS is expressed in many tissues, with high expression in large neurons, particu-
larly cerebellar Purkinje cells (Parfitt et al. 2009 ). Subcellular localisation shows
a predominantly cytoplasmic localisation with a significant mitochondrial compo-
nent (Vermeer et al. 2008 ; Girard et al. 2012 ).
SACS is a putative co-chaperone of the HSP40 family based upon the presence
of a J domain at the C-terminus of its predicted amino acid sequence (~ 60 % iden-
tity over 30 residues compared to hdj-1 (DNAJB1)). Although the SACS J domain
is divergent from that of HSP40 it does contain the highly conserved His-Pro-Asp
(HPD) motif essential for the stimulation of HSP70 ATPase activity. The SACS J-
domain was found to function with bacterial HSP70 (DnaK) by an in vivo comple-
mentation assay (Parfitt et al. 2009 ), and a recombinant version of the J-domain
from mouse SACS increased the ATPase activity of HSP70 (Anderson et al. 2010 ).
This confirmed that SACS is a type III HSP40 protein. Type III HSP40 proteins
recruit HSP70s to specialised roles. Interestingly, mutations in the HSP40 family
DNAJB2 gene have recently been shown to cause distal hereditary motor neuropa-
thy (dHMN), characterised by motor neuron degeneration in the anterior horn spinal
cord, muscle weakness and atrophy (Blumen et al. 2012 ).
SACS consists of three repeated regions, known as the SACS-repeating region
(SRR), which cover ~84% of the protein sequence (Romano et al. 2013 ). Each
repeated region contains discrete sub-repeats, the first of which is homologous to
the HATPase (Histidine kinase-like ATPases; SMART acc. no. SM00387) domain
of HSP90 (Anderson et al. 2010 ; Romano et al. 2013 ). Biochemical characterisation
identified the repeating regions to be ATPase active. Furthermore, a disease caus-
ing mutation (D168Y) within the first sub-repeat region, homologous to HSP90,
abolished the ability to hydrolyse ATP. From this, it was suggested that ATPase
activity is a requirement for SACS function, as this mutation leads to essentially
the same clinical phenotype as nonsense mutations close to the N-terminus of the
protein that result in truncations of the protein (Anderson et al. 2010 ). In HSP90,
the middle domain contains an arginine residue that accepts phosphate after ATP
hydrolysis (Pearl and Prodromou 2006 ). This arginine is conserved in each of the
three SRR domains and a mutation occurring in one of these conserved arginines,
namely c.1420C > T (p.R474C), leads to a severe clinical phenotype (Romano et al.
2013 ). Previously, HSP90 and the hdj-2 (DNAJA1) protein have been implicated
to function together in folding pathways, for example in the maturation of the glu-
cocorticoid receptor.