Comparison of the effects of mutaions modelling early-onset familial Alzheimer's disease and Sanfilippo syndrome childhood dementia in zebrafish larvae

Neural cells are under immense pressure to maintain proper degradation of molecules using their endo-lysosomal pathway (ELP). In Alzheimer's disease (AD), the gradual cognitive worsening of this age-related disorder is partially due to dysfunction of the ELP. However, AD is a complex disorder and how the genetic landscape eventually leads to AD is unclear. Sanfilippo syndrome childhood dementia (mucopolysaccharidosis type III, MPSIII) is an extreme consequence of ELP dysfunction, where inherited recessive mutations result in the inability to properly catabolise a particular polysaccharide, which ultimately accumulates and interferes with the function of the ELP. Unlike AD, Sanfilippo is genetically simple (all causative genes are identified). The primary biochemical consequences are understood and animal models of Sanfilippo are considered excellent models of the human disease. Since Sanfilippo and AD show similar pathological changes, they likely share disease-associated mechanisms. Here, we dissect the similarity between AD and Sanfilippo using RNA-seq. We have generated zebrafish knock-in models of both diseases: the early-onset familial AD-like (EOfAD-like) mutation psen1 Q96_K97del and the MPSIII-like mutation naglu A603fs. We generated a family of zebrafish containing wild type, heterozygous EOfAD-like and homozygous MPSIII-like genotypes. We raised the family until 7 days post fertilisation and performed mRNA-seq on n = 8 induvial larvae per genotype. A power calculation revealed that this experiment was only contained ~50% power to detect differentially expressed genes. Nevertheless, changes to the expression of genes encoding proteins involved in lysosomal function could be detected in the MPS-III mutant larvae, suggesting a homeostatic response as the cells which make up the larval RNA-seq samples respond to dysfunctional lysosomes. Overall design: We first paired fish a female zebrafish with genotype psen1 Q96_K97del/+ ; naglu A603fs/+ with a male zebrafish with genotype psen1 +/+ ; naglu A603fs/+. This gave a large family of > 100 sibling larvae with various psen1 and naglu genotypes. We raised the family of sibling larvae together in a large petri dish until 7 days post fertilisation. The, the entire family was euthanised using an ice slurry. Individual larvae were placed in individual microfuge tubes containing 100 µL of RNAlater solution. Samples were then incubated in the RNAlater solution overnight at 4°C. Then, the tail from each larvae was cut from the rest of the head at the end of the yolk sac extension. The tail piece was used for genomic DNA extraction and PCR genotyping. The head was placed back in RNAlater. Once each larvae was genotyped for psen1 and naglu mutations, a total of 24 individual larvae were used for RNA extraction and sequencing: n = 8 wild type larvae, n = 8 EOfAD-like/+ larvae and n = 8 MPS-IIIB homozygous larvae.