Biallelic COQ4 Variants in Hereditary Spastic Paraplegia: Clinical and Molecular Characterization.

BACKGROUND: Hereditary spastic paraplegias (HSP) are neurologic disorders characterized by progressive lower-extremity spasticity. Despite the identification of several HSP-related genes, many patients lack a genetic diagnosis. OBJECTIVES: The aims were to confirm the pathogenic role of biallelic COQ4 mutations in HSP and elucidate the clinical, genetic, and functional molecular features of COQ4-associated HSP. METHODS: Whole exome sequences of 310 index patients with HSP of unknown cause from three distinct populations were analyzed to identify potential HSP causal genes. Clinical data obtained from patients harboring candidate causal mutations were examined. Functional characterization of COQ4 variants was performed using bioinformatic tools, single-cell RNA sequencing, biochemical assays in cell lines, primary fibroblasts, induced pluripotent stem cell-derived pyramidal neurons, and zebrafish. RESULTS: Compound heterozygous variants in COQ4, which cosegregated with HSP in pedigrees, were identified in 7 patients from six unrelated families. Patients from four of the six families presented with pure HSP, whereas probands of the other two families exhibited complicated HSP with epilepsy or with cerebellar ataxia. In patient-derived fibroblasts and COQ4 knockout complementation lines, stable expression of these missense variants exerted loss-of-function effects, including mitochondrial reactive oxygen species accumulation, decreased mitochondrial membrane potential, and lower ubiquinone biosynthesis. Whereas differentiated pyramidal neurons expressed high COQ4 levels, coq4 knockdown zebrafish displayed severe motor dysfunction, reflecting motor neuron dysregulation. CONCLUSIONS: Our study confirms that loss-of-function, compound heterozygous, pathogenic COQ4 variants are causal for autosomal recessive pure and complicated HSP. Moreover, reduced COQ4 levels attributable to variants correspond with decreased ubiquinone biosynthesis, impaired mitochondrial function, and higher phenotypic disease severity. © 2023 International Parkinson and Movement Disorder Society.

Loss of function of CMPK2 causes mitochondria deficiency and brain calcification.

Brain calcification is a critical aging-associated pathology and can cause multifaceted neurological symptoms. Cerebral phosphate homeostasis dysregulation, blood-brain barrier defects, and immune dysregulation have been implicated as major pathological processes in familial brain calcification (FBC). Here, we analyzed two brain calcification families and identified calcification co-segregated biallelic variants in the CMPK2 gene that disrupt mitochondrial functions. Transcriptome analysis of peripheral blood mononuclear cells (PBMCs) isolated from these patients showed impaired mitochondria-associated metabolism pathways. In situ hybridization and single-cell RNA sequencing revealed robust Cmpk2 expression in neurons and vascular endothelial cells (vECs), two cell types with high energy expenditure in the brain. The neurons in Cmpk2-knockout (KO) mice have fewer mitochondrial DNA copies, down-regulated mitochondrial proteins, reduced ATP production, and elevated intracellular inorganic phosphate (Pi) level, recapitulating the mitochondrial dysfunction observed in the PBMCs isolated from the FBC patients. Morphologically, the cristae architecture of the Cmpk2-KO murine neurons was also impaired. Notably, calcification developed in a progressive manner in the homozygous Cmpk2-KO mice thalamus region as well as in the Cmpk2-knock-in mice bearing the patient mutation, thus phenocopying the calcification pathology observed in the patients. Together, our study identifies biallelic variants of CMPK2 as novel genetic factors for FBC; and demonstrates how CMPK2 deficiency alters mitochondrial structures and functions, thereby highlighting the mitochondria dysregulation as a critical pathogenic mechanism underlying brain calcification.

Advances in gene therapy for neurogenetic diseases: a brief review.

Neurogenetic diseases are neurological conditions with a genetic cause (s). There are thousands of neurogenetic diseases, and most of them are incurable. The development of bioinformatics and elucidation of the mechanism of pathogenesis have allowed the development of gene therapy approaches, which show great potential in treating neurogenetic diseases. Viral vectors delivery, antisense oligonucleotides, gene editing, RNA interference, and burgeoning viroid delivery technique are promising gene therapy strategies, and commendable therapeutic effects in the treatment of neurogenetic diseases have been achieved (Fig. 1). This review highlights a sampling of advances in gene therapies for neurogenetic disorders. Fig. 1 Examples of gene therapy strategies used in the treatment of neurogenetic diseases. The schematic diagram shows different gene therapy approaches used for treating a sampling of neurogenetic disorders, such as ASO therapy, gene editing, gene augmentation, and RNA interference.

Advances in detecting and reducing off-target effects generated by CRISPR-mediated genome editing.

CRISPR-mediated genome editing is a revolutionary technology for genome manipulation that uses the CRISPR-Cas systems and base editors. Currently, poor efficiency and off-target problems have impeded the application of CRISPR systems. The on-target efficiency has been improved in several advanced versions of CRISPR systems, whereas the off-target detection still remains a key challenge. Here, we outline the different versions of CRISPR systems and off-target detection strategies, discuss the merits and limitations of off-target detection methods, and provide potential implications for further gene editing research.