Project description:Spinocerebellar ataxias (SCAs) are a group of cerebellar diseases characterized by loss and dysfunction of Purkinje cells and Spinocerebellar ataxia type 14 (SCA14) is caused by missense mutations or deletions in the Protein kinase C γ (PKCγ) gene. Until now, more than 40 different mutations or deletions in the PKCγ gene have been found in SCA14 patients. Many of these mutations have been shown to have an increased enzymatic activity in cell-based assays, but there is also evidence that the mutations may result in inefficient activation of downstream signalling pathways compatible with a loss of function. Therefore, it is still unclear how mutant PKCγ may cause the disease. We have previously generated a transgenic SCA14 mouse model with a human SCA14 mutation in the kinase domain. This transgenic mouse shows mild ataxia and abnormal Purkinje cell dendritic development with a morphology indistinguishable from that of PKC activator treated Purkinje cells, indicating that the PKCγ with this kinase domain mutation has indeed increased biological activity. In order to confirm that increased PKC activity in vivo perturbs Purkinje cell maturation and induces ataxia we have now created a new knock-in mouse model with a missense mutation in the PKCγ pseudosubstrate domain keeping the PKCγ protein in the open active conformation. This knock-in mouse shows indeed abnormal Purkinje cell maturation and ataxia, even in a heterozygous state corresponding to the human disease situation. Our findings confirm that constitutive activation of PKCγ is one way to induce a phenotype corresponding to human spinocerebellar ataxia.
Project description:Spinocerebellar Ataxia Autosomal Recessive 20 (SCAR20) is a rare disorder caused by mutation in the SNX14 gene. The symptoms of the disorder including coarse face, poor or lack of speech, delayed or absent of motor function, and most markedly, cerebella atrophy with Purkinje cell degeneration. SNX14 has been localized to ER and shown to play a vital role in facilitating the formation of lipid droplet and maintaining the lipid saturation balance in cell membrane. Meanwhile, it has also been localized to other subcellular organelles, including lysosome and endo-lysosome, and has been suggested to participate in the autophagy responses. However, the previous researches were performed in vitro or ex vivo, and how disrupted SNX14 led to Purkinje cell degeneration in the SCAR20 cerebella in vivo remained unexplored. Here, we report the first animal model that manifested the SCAR20 symptoms, by introducing a 1bp deletion in Snx14 in mice via CRISPR-Cas9. We performed RNAseq to analyse the pathway changes caused by Snx14 KO.
Project description:Spinocerebellar Ataxia Autosomal Recessive 20 (SCAR20) is a rare disorder caused by mutation in the SNX14 gene. The symptoms of the disorder including coarse face, poor or lack of speech, delayed or absent of motor function, and most markedly, cerebella atrophy with Purkinje cell degeneration. SNX14 has been localized to ER and shown to play a vital role in facilitating the formation of lipid droplet and maintaining the lipid saturation balance in cell membrane. Meanwhile, it has also been localized to other subcellular organelles, including lysosome and endo-lysosome, and has been suggested to participate in the autophagy responses. However, the previous researches were performed in vitro or ex vivo, and how disrupted SNX14 led to Purkinje cell degeneration in the SCAR20 cerebella in vivo remained unexplored. Here, we report the first animal model that manifested the SCAR20 symptoms, by introducing a 1bp deletion in Snx14 in mice via CRISPR-Cas9. We performed RNAseq to analyse the pathway changes caused by Snx14 KO.
Project description:Autosomal recessive spastic ataxia of Charlevoix-Saguenay (ARSACS) is a childhood-onset cerebellar ataxia caused by mutations in SACS, which encodes the protein sacsin. Cellular ARSACS phenotypes include mitochondrial dysfunction, intermediate filament disorganization, and the progressive death of cerebellar Purkinje neurons. It is unclear how the loss of sacsin function causes these deficits, or why they manifest as cerebellar ataxia. To investigate this, we performed multi-omic profiling of sacsin knockout cells and compared them to wild-type controls
Project description:Spinocerebellar ataxia type 22 (SCA22) caused by KCND3 mutations is an autosomal dominant disorder. We establish a mouse model carrying the Kcnd3 F227del mutation to study the molecular pathogenesis. Four findings are pinpointed. Firstly, the heterozygous mice exhibit an early onset of defects in motor coordination and balance, which mirror the SCA22 patients. The degeneration and a minor loss of Purkinje cells together with the concurrent presence of neuroinflammation, as well as the previous finding on electrophysiological changes, may all contribute to the development of the SCA22 ataxia phenotype in mice carrying the Kcnd3 F227del mutant protein. Secondly, the mutant protein is retained by the endoplasmic reticulum and Golgi leading to activation of the unfolded protein response and a severe trafficking defect that affects its membrane destination. Intriguingly, profound damage to Golgi is the earliest manifestation. Thirdly, transcriptomic analysis revealed that the Kcnd3 F227del mutation down-regulates a panel of genes involved in the functioning of synapse and neurogenesis which are tightly linked to the functioning of Purkinje cells. Finally, no ataxia phenotypes are detectable in knockout mice carrying a loss-of-function Kcnd3 mutation. Thus, the Kcnd3 F227del is a dominant-negative mutation. Additionally, this mouse model can serve as a pre-clinical model for exploring therapeutic strategies to treat patients.
Project description:GEMIN5 is a critical component of snRNP assembly complex. Patients carrying novel autosomal recessive variants in the GEMIN5 gene showed symptoms of developmental delay, central hypotonia, and cerebellar ataxia which are distinct than classical spinal muscular atrophy. We performed RNA-seq analysis in iPSC-derived differentiated neurons with biallelic GEMIN5-H913R mutation to identify global alterations in various genes and pathways mediated by GEMIN5 mutations in patients.
Project description:Uniparental disomy (UPD) refers to as both homologous chromosomes inherited from only one parent without identical copies from the other parent. Current knowledge of the phenotypes is associated with imprinting disorders or autosomal recessive mutations. Recent studies had reported that UPD for chromosomes 6, 7, 11, 14, 15, and 20 accompanied with definite imprinted disorders. However, the phenotypes and outcomes of UPD with other chromosomes have not been completely identified. Herein, we combined three remarkable UPD patients involving different chromosomes and presented their phenotypes. The chromosomal microarray (CMA) demonstrated a whole homozygous region on chromosome 2 in patient A, on chromosome 9 in patient B, accompanied by a gain mosaic region (ratio:22%) of 19.18Mb in 9p22.2p13.2, and a LOH region on 14q23.2q32.12 and several genes with compound heterozygous mutation were detected in patient C
Project description:Ataxia with oculomotor apraxia type 1 (AOA1) is an early onset progressive spinocerebellar ataxia caused by mutation in aprataxin (APTX). Here we use RNA-seq to identify genes that are affected by APTX-KO, APTX overexpression, and APTX mutant, thus contributes to understadning the mechanisms underlying AOA1 pathlogy. Examination the chages of gene expressions in APTX proficient and APTX deficienc cells.
Project description:To explore the mechanism associated with retinal degeneration and adeno-associated virus (AAV)-mediated gene therapy in rd10 mouse, a model of autosomal recessive retinitis pigmentosa (arRP) containing mutation of β subunit of the rod cGMP phosphodiesterase 6 (PDE6).
Project description:Spinocerebellar ataxia with axonal neuropathy (SCAN1) is a rare recessive neurodegenerative syndrome associated with cerebellar atrophy and peripheral neuropathy. It is caused by a homozygous missense mutation in the tyrosyl-DNA phosphodiesterase-1 (TDP1) gene (A1478G). resulting in a substitution of histidine for arginine-493 (H493R) in the TDP1 catalytic site, leading to reduced TDP1 activity. How TDP1 H493R mutation promotes the SCAN1 phenotype, which is associated with the death of post-mitotic neurons, is unclear. We have generated models of osteosarcoma U2OS cells homozygous for TDP1 H493R employing the CRISPR-Cas9 technique (2 clones, named “1P” and “3.3”). Here, we have generated transcriptional genome wide profile in order to characterize differences in gene expression that are specific of TDP1-mutated clones.