Machine learning discriminates a movement disorder in a zebrafish model of Parkinson's disease.

Animal models of human disease provide an in vivo system that can reveal molecular mechanisms by which mutations cause pathology, and, moreover, have the potential to provide a valuable tool for drug development. Here, we have developed a zebrafish model of Parkinson's disease (PD) together with a novel method to screen for movement disorders in adult fish, pioneering a more efficient drug-testing route. Mutation of the PARK7 gene (which encodes DJ-1) is known to cause monogenic autosomal recessive PD in humans, and, using CRISPR/Cas9 gene editing, we generated a Dj-1 loss-of-function zebrafish with molecular hallmarks of PD. To establish whether there is a human-relevant parkinsonian phenotype in our model, we adapted proven tools used to diagnose PD in clinics and developed a novel and unbiased computational method to classify movement disorders in adult zebrafish. Using high-resolution video capture and machine learning, we extracted novel features of movement from continuous data streams and used an evolutionary algorithm to classify parkinsonian fish. This method will be widely applicable for assessing zebrafish models of human motor diseases and provide a valuable asset for the therapeutics pipeline. In addition, interrogation of RNA-seq data indicate metabolic reprogramming of brains in the absence of Dj-1, adding to growing evidence that disruption of bioenergetics is a key feature of neurodegeneration.This article has an associated First Person interview with the first author of the paper.

Skeletal muscle differentiation drives a dramatic downregulation of RNA polymerase III activity and differential expression of Polr3g isoforms.

Gene regulatory networks underpinning skeletal muscle determination and differentiation have been extensively investigated, providing molecular insights into how cell lineages are established during development. These studies have exclusively focused on the transcriptome downstream of RNA polymerase II (Pol II). RNA polymerase III (Pol III) drives the production of tRNAs and other small RNAs essential for the flow of genetic information from gene to protein and we have found that a specific isoform of a subunit unique to Pol III is expressed early in the myogenic lineage. This points to the possibility that additional regulatory networks exist to control the production of Pol III transcripts during skeletal muscle differentiation. We describe the differential expression of Polr3g and its alternate isoform Polr3gL during embryonic development and using a custom tRNA microarray, we demonstrate their distinct activity on the synthesis of tRNA isoacceptors. We show that Pol III dependent transcripts are dramatically down-regulated during the differentiation of skeletal muscle, as are mRNAs coding for Pol III associated proteins Brf1 and Brf2, while Polr3gL is up-regulated alongside contractile protein genes. Forcing Polr3g expression in this context results in a partial reversal of myogenic differentiation.

Transcriptional upregulation of Bag3, a chaperone-assisted selective autophagy factor, in animal models of KY-deficient hereditary myopathy.

The importance of kyphoscoliosis peptidase (KY) in skeletal muscle physiology has recently been emphasised by the identification of novel human myopathies associated with KY deficiency. Neither the pathogenic mechanism of KY deficiency nor a specific role for KY in muscle function have been established. However, aberrant localisation of filamin C (FLNC) in muscle fibres has been shown in humans and mice with loss-of-function mutations in the KY gene. FLNC turnover has been proposed to be controlled by chaperone-assisted selective autophagy (CASA), a client-specific and tension-induced pathway that is required for muscle maintenance. Here, we have generated new C2C12 myoblast and zebrafish models of KY deficiency by CRISPR/Cas9 mutagenesis. To obtain insights into the pathogenic mechanism caused by KY deficiency, expression of the co-chaperone BAG3 and other CASA factors was analyzed in the cellular, zebrafish and ky/ky mouse models. Ky-deficient C2C12-derived clones show trends of higher transcription of CASA factors in differentiated myotubes. The ky-deficient zebrafish model (kyyo1/kyyo1 ) lacks overt signs of pathology, but shows significantly increased bag3 and flnca/b expression in embryos and adult muscle. Additionally, kyyo1/kyyo1 embryos challenged by swimming in viscous media show an inability to further increase expression of these factors in contrast with wild-type controls. The ky/ky mouse shows elevated expression of Bag3 in the non-pathological exterior digitorum longus (EDL) and evidence of impaired BAG3 turnover in the pathological soleus. Thus, upregulation of CASA factors appears to be an early and primary molecular hallmark of KY deficiency.