Abstract
Mucopolysaccharidosis IIIA (MPS IIIA, Sanfilippo syndrome type A), a paediatric neurological lysosomal storage disease, is caused by impaired function of the enzyme N-sulfoglucosamine sulfohydrolase (SGSH) resulting in impaired catabolism of heparan sulfate glycosaminoglycan (HS GAG) and its accumulation in tissues. MPS IIIA represents a significant proportion of childhood dementias. This condition generally leads to patient death in the teenage years, yet no effective therapy exists for MPS IIIA and a complete understanding of the mechanisms of MPS IIIA pathogenesis is lacking. Here, we employ targeted CRISPR/Cas9 mutagenesis to generate a model of MPS IIIA in the zebrafish, a model organism with strong genetic tractability and amenity for high-throughput screening. The sgshΔex5−6 zebrafish mutant exhibits a complete absence of Sgsh enzymatic activity, leading to progressive accumulation of HS degradation products with age. sgshΔex5−6 zebrafish faithfully recapitulate diverse CNS-specific features of MPS IIIA, including neuronal lysosomal overabundance, complex behavioural phenotypes, and profound, lifelong neuroinflammation. We further demonstrate that neuroinflammation in sgshΔex5−6 zebrafish is largely dependent on interleukin-1β and can be attenuated via the pharmacological inhibition of Caspase-1, which partially rescues behavioural abnormalities in sgshΔex5−6 mutant larvae in a context-dependent manner. We expect the sgshΔex5−6 zebrafish mutant to be a valuable resource in gaining a better understanding of MPS IIIA pathobiology towards the development of timely and effective therapeutic interventions.
Highlights
IntroductionMPS IIIA initially presents in patients during early developmental stages as a failure to meet developmental milestones, coinciding with the onset of behavioural abnormalities including hyperactivity and impaired sleep patterns [5]
A dual-gRNA CRISPR/Cas9-mediated genome editing approach was employed to delete a conserved part of the sgsh gene and generate a stable sgsh loss-of-function zebrafish mutant
As exons 4 and 7 are not phase-compatible, we did not expect to preserve translation through to the original termination codon, as a clean splicing event between exons 4 and 7 should result in a premature termination codon (PTC) early in exon
Summary
MPS IIIA initially presents in patients during early developmental stages as a failure to meet developmental milestones, coinciding with the onset of behavioural abnormalities including hyperactivity and impaired sleep patterns [5] These clinical features precede a progressive decline in intellectual and cognitive functions [6,7,8], often leading to onset of dementia during early childhood. While much has been studied relating to the course of MPS IIIA progression and its clinical features, relatively little is known about the cellular mechanisms through which the disease manifests in patients. This represents a severe limitation for the development of disease-modifying therapeutic interventions. SGSH removes the sulfate group from N-Sulfoglucosamine by hydrolysis to produce D-Glucosamine [15]; when SGSH activity is absent, subsequent steps in the HS degradation pathway (e.g., NAGLU-mediated acetylglucosaminidase activity impaired in MPS IIIB, HGSNAT-mediated acetyltransferase activity which is impaired in MPS IIIC, or GNS-mediated sulfatase activity in MPS IIID)
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