Comparative Analysis and Functional Mapping of SACS Mutations Reveal Novel Insights into Sacsin Repeated Architecture

Alessandro Romano,Alessandra Terracciano,Fabiana Fattori,Carlo Storelli,Maria Fulvia De Leva,Tiziano Verri,Amilcare Barca,Alessandra Tessa,Filippo Maria Santorelli

doi:10.1002/humu.22269

Abstract

Autosomal recessive spastic ataxia of Charlevoix–Saguenay (ARSACS) is a neurological disease with mutations in SACS, encoding sacsin, a multidomain protein of 4,579 amino acids. The large size of SACS and its translated protein has hindered biochemical analysis of ARSACS, and how mutant sacsins lead to disease remains largely unknown. Three repeated sequences, called sacsin repeating region (SRR) supradomains, have been recognized, which contribute to sacsin chaperone-like activity. We found that the three SRRs are much larger (≥1,100 residues) than previously described, and organized in discrete subrepeats. We named the large repeated regions Sacsin Internal RePeaTs (SIRPT1, SIRPT2, and SIRPT3) and the subrepeats sr1, sr2, sr3, and srX. Comparative analysis of vertebrate sacsins in combination with fine positional mapping of a set of human mutations revealed that sr1, sr2, sr3, and srX are functional. Notably, the position of the pathogenic mutations in sr1, sr2, sr3, and srX appeared to be related to the severity of the clinical phenotype, as assessed by defining a severity scoring system. Our results suggest that the relative position of mutations in subrepeats will variably influence sacsin dysfunction. The characterization of the specific role of each repeated region will help in developing a comprehensive and integrated pathophysiological model of function for sacsin.

Highlights

Autosomal recessive spastic ataxia of Charlevoix–Saguenay (ARSACS; MIM #270550) is an early-onset neurological disease presenting a founder effect in the Quebec regions of Charlevoix and Saguenay–Lac-St-Jean where the estimated carrier frequency is 1/22 [Bouchard et al, 1978; 1998]
Along with the original description of human SACS [Engert et al, 2000], it was suggested that repeating regions, two of which containing the putative ATP-binding domain of Hsp90, might have occurred in the sacsin protein
Similar results were obtained by using the HHRepID program [Biegert and Soding, 2008]. We named these three large homologous repeating regions Sacsin Internal RePeaTs. In spite of their low overall similarity (e.g., 16%– 18% in human sacsin, with SIRPT1 vs. SIRPT2: 17%, SIRPT1 vs. SIRPT3: 16%, and SIRPT2 vs. SIRPT3: 18%), each Sacsin internal repeat (SIRPT) displayed, based on the degree of local similarity, at least three subrepeats (Fig. 1B) that were distanced by regions of extremely low similarity

Summary

Introduction

Autosomal recessive spastic ataxia of Charlevoix–Saguenay (ARSACS; MIM #270550) is an early-onset neurological disease presenting a founder effect in the Quebec regions of Charlevoix and Saguenay–Lac-St-Jean where the estimated carrier frequency is 1/22 [Bouchard et al, 1978; 1998]. The major clinical features of ARSACS include early-onset ataxia, later occurrence of spastic paraparesis, and brisk tendon reflexes, and an axonal sensory-motor peripheral neuropathy, with some instances of mental retardation or cognitive decline. Hypermyelination of the retinal nerve fibers [Bouchard et al, 1978, 1998] has long been considered a cardinal feature in Quebecois French–Canadian patients, and is not so obvious in cases from elsewhere [Criscuolo et al, 2004; Hara et al, 2005] or even absent. Several aspects including early appearance of abnormal pontocerebellar and retinal fibers seen at brain neuroimaging speak for a neurodevelopmental anomaly in ARSACS [Gazulla et al, 2012]. The progressive clinical course with involvement of the corticospinal tract and peripheral nerves in patients as well as studies in model mice questioned this hypothesis and suggested the occurrence of a neurodegenerative process [Girard et al, 2012; Prodi et al, 2012]

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Results

Conclusion