RESUMO
BACKGROUND: Heterozygous pathogenic variants in STUB1 are implicated in autosomal dominant spinocerebellar ataxia type 48 (SCA48), which is a rare familial ataxia disorder. We investigated the clinical, genetic and functional characteristics of STUB1 mutations identified from a Taiwanese ataxia cohort. METHODS: We performed whole genome sequencing in a genetically undiagnosed family with an autosomal dominant ataxia syndrome. Further Sanger sequencing of all exons and intron-exon boundary junctions of STUB1 in 249 unrelated patients with cerebellar ataxia was performed. The pathogenicity of the identified novel STUB1 variant was investigated. RESULTS: We identified a novel heterozygous frameshift variant, c.832del (p.Glu278fs), in STUB1 in two patients from the same family. This rare mutation is located in the U-box of the carboxyl terminus of the Hsc70-interacting protein (CHIP) protein, which is encoded by STUB1. Further in vitro experiments demonstrated that this novel heterozygous STUB1 frameshift variant impairs the CHIP protein's activity and its interaction with the E2 ubiquitin ligase, UbE2D1, leading to neuronal accumulation of tau and α-synuclein, caspase-3 activation, and promoting cellular apoptosis through a dominant-negative pathogenic effect. The in vivo study revealed the influence of the CHIP expression level on the differentiation and migration of cerebellar granule neuron progenitors during cerebellar development. CONCLUSIONS: Our findings provide clinical, genetic, and a mechanistic insight linking the novel heterozygous STUB1 frameshift mutation at the highly conserved U-box domain of CHIP as the cause of autosomal dominant SCA48. Our results further stress the importance of CHIP activity in neuronal protein homeostasis and cerebellar functions.
Assuntos
Mutação da Fase de Leitura , Ataxias Espinocerebelares/genética , Ubiquitina-Proteína Ligases/genética , Adulto , Idoso , Estudos de Coortes , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Taiwan , Ubiquitina-Proteína Ligases/metabolismoRESUMO
Alzheimer's disease (AD), with a typical pathological hallmark of amyloidbeta (Aß)containing plaques and neurofibrillary tangles, is one of the most common types of chronic neurodegenerative diseases. Aß oligomers serve a crucial role in the pathogenesis of AD, and lead to neuronal loss. However, the precise mechanism of Aß oligomers in AD remains to be elucidated. The present study demonstrated that 10 µM Aß42 activated the caspase signaling pathway, and induced significant apoptosis in primary cultured mouse cerebral cortical neurons. The results of reverse transcriptionquantitative polymerase chain reaction and western blotting demonstrated that Aß42 (10 µM) also significantly upregulated the transcription and expression of the mitochondrial fission protein dynaminrelated protein 1 (Drp1), and downregulated the transcription and expression of mitochondrial fusion proteins, including mitofusin 1/2 (Mfn1/2) and mitochondrial dynamin like GTPase (OPA1). Neurons were transfected with pDsRed2Mito for mitochondrial imaging, which revealed that 10 µM Aß42 induced mitochondrial fission in cortical neurons. In addition, 2',7'dichlorodihydrofluorescein diacetate and tetramethylrhodamine ethyl ester staining indicated that Aß42 increased the reactive oxygen species (ROS) level and reduced mitochondrial membrane potential in neurons. Inhibition of Drp1 activity by Mdivi1 efficiently prevented Aß42induced ROS production and disruption of mitochondrial membrane potential. Loss of mitochondrial membrane potential may activate PTENinduced putative kinase 1 (Pink1), the prominent sensor for mitochondrial damage, and trigger the process of mitophagy to remove the damaged mitochondria. In the present study, western blotting revealed that the levels of autophagy marker microtubuleassociated proteins 1A/1B light chain 3B (LC3B) and Pink1 were upregulated after Aß42 stimulation. In conclusion, these data indicated that Aß42 induces neuronal apoptosis by targeting mitochondria, including promotion of mitochondrial fission, disruption of mitochondrial membrane potential, increasing intracellular ROS level and activation of the process of mitophagy. Therefore, mitochondria may represent a potential therapeutic target for AD in the future.