Project description:Biallelic loss of expression/function variants in MTMR5/SBF1 cause the inherited peripheral neuropathy Charcot-Marie-Tooth (CMT) Type 4B3. There is an incomplete understanding of the disease pathomechanism(s) underlying CMT4B3, and despite its severe clinical presentation, currently no disease modifying therapies. A key barrier to the study of CMT4B3 is the lack of pre-clinical models that recapitulate the clinical and pathologic features of the disease. To address this barrier, we generated a zebrafish CRISPR/Cas9 mutant line with a full gene deletion of mtmr5. Resulting homozygous deletion zebrafish are born at normal Mendelian ratios and have preserved motor function. However, starting by 10 days-post-fertilization, mutant zebrafish develop obvious morphometric changes in head size and brain volume. These changes are accompanied at the pathological level by abnormal axon outgrowths and by the presence of dysmyelination, changes reminiscent of the nerve pathology in human CMT4B3. Importantly, RNA sequencing from brain-enriched samples identifies novel disease pathways including transcriptional changes in genes responsible for neurogenesis, chromatin remodeling/organization, and synaptic membrane homeostasis. Overall, our mtmr5 knockout zebrafish mirror genetic, clinical, and pathologic features of human CMT4B3. As such, it represents a first pre-clinical model to phenocopy the disease, and an ideal tool for future studies on disease pathomechanism(s) and therapy development.
Project description:Aminoacyl-tRNA synthetases are indispensable enzymes in all cells, ensuring the correct pairing of amino acids to their cognate tRNAs to maintain translational fidelity. Autosomal dominant mutations V133F and Y330C in histidyl-tRNA synthetase (HARS) cause the genetic disorder Charcot-Marie-Tooth type 2W (CMT2W). HARS V133F and Y330C cause mistranslation as validated by mass spectrometry and growth defects that persist with histidine supplementation. The growth defects and translation fidelity for both V133F and Y330C mutants were rescued by supplementation with human tRNAHis in a humanized yeast model.
Project description:Charcot-Marie-Tooth disease (CMT) is a length-dependent peripheral neuropathy. The aminoacyl-tRNA synthetases constitute the largest protein family implicated in CMT. Aminoacyl-tRNA synthetases are predominantly cytoplasmic, but are also present in the nucleus. Here we show that a nuclear function of tyrosyl-tRNA synthetase (TyrRS) is implicated in a Drosophila model of CMT. CMT-causing mutations in TyrRS induce unique conformational changes, which confer capacity for aberrant interactions with transcriptional regulators in the nucleus, leading to transcription factor E2F1 hyperactivation. Using neuronal tissues, we reveal a broad transcriptional regulation network associated with wild-type TyrRS expression, which is disturbed when a CMT-mutant is expressed. Pharmacological inhibition of TyrRS nuclear entry with embelin reduces, whereas genetic nuclear exclusion of mutant TyrRS prevents hallmark phenotypes of CMT in the Drosophila model. These data highlight that this translation factor may contribute to transcriptional regulation in neurons, and suggest a therapeutic target for CMT.