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Updated: Oct 28, 2025
Modeling Charcot-Marie-Tooth Disease In Vitro by Transfecting Mouse Primary Motoneurons
Published on: January 7, 2019
Modeling a human CLP1 mutation in mouse identifies an accumulation of tyrosine pre-tRNA fragments causing
Ikuko Morisaki1, Hiroshi Shiraishi1, Hiroyuki Fujinami1
1Department of Cell Biology, Oita University Faculty of Medicine, Yufu, Oita, 879-5593, Japan.
Abstract:
Cleavage factor polyribonucleotide kinase subunit 1 (CLP1), an RNA kinase, plays essential roles in protein complexes involved in the 3'-end formation and polyadenylation of mRNA and the tRNA splicing endonuclease complex, which is involved in precursor tRNA splicing. The mutation R140H in human CLP1 causes pontocerebellar hypoplasia type 10 (PCH10), which is characterized by microcephaly and axonal peripheral neuropathy. Previously, we reported that RNA fragments derived from isoleucine pre-tRNA introns (Ile-introns) accumulate in fibroblasts of patients with PCH10. Therefore, it has been suggested that this intronic RNA fragment accumulation may trigger PCH10 onset. However, the molecular mechanism underlying PCH10 pathogenesis remains elusive. Thus, we generated knock-in mutant mice that harbored a CLP1 mutation consistent with R140H. As expected, these mice showed progressive loss of the upper motor neurons, resulting in impaired locomotor activity, although the phenotype was milder than that of the human variant. Mechanistically, we found that the R140H mutation causes intracellular accumulation of Ile-introns derived from isoleucine pre-tRNAs and 5' tRNA fragments derived from tyrosine pre-tRNAs, suggesting that these two types of RNA fragments were cooperatively or independently involved in the onset and progression of the disease. Taken together, the CLP1-R140H mouse model provided new insights into the pathogenesis of neurodegenerative diseases, such as PCH10, caused by genetic mutations in tRNA metabolism-related molecules.
Insights
A mutation in CLP1 (Cleavage factor polyribonucleotide kinase subunit 1) causes pontocerebellar hypoplasia type 10 (PCH10). This study reveals that accumulated RNA fragments, specifically Ile-introns and 5' tRNA fragments, contribute to PCH10 pathogenesis.
Area of Science:
- Molecular Biology
- Neuroscience
- Genetics
Background:
- Cleavage factor polyribonucleotide kinase subunit 1 (CLP1) is crucial for mRNA processing and tRNA splicing.
- The R140H mutation in CLP1 is linked to pontocerebellar hypoplasia type 10 (PCH10), a neurodegenerative disorder.
- Previous studies indicated accumulation of isoleucine pre-tRNA introns (Ile-introns) in PCH10 patients.
Purpose of the Study:
- To investigate the molecular mechanisms underlying PCH10 pathogenesis caused by the CLP1 R140H mutation.
- To establish and characterize a CLP1 R140H knock-in mouse model for PCH10 research.
- To identify the specific RNA species accumulating due to the CLP1 mutation.
Main Methods:
- Generation of CLP1 R140H knock-in mutant mice.
- Phenotypic analysis of mutant mice, including motor function and neuronal integrity.
- Molecular analysis to detect and quantify accumulating RNA fragments in affected tissues.
Main Results:
- CLP1 R140H mutant mice exhibited progressive upper motor neuron loss and impaired locomotor activity.
- Intracellular accumulation of Ile-introns and 5' tRNA fragments from tyrosine pre-tRNAs was observed in mutant mice.
- The observed phenotypes, while milder than human PCH10, validate the mouse model for studying disease mechanisms.
Conclusions:
- The CLP1 R140H mutation leads to the accumulation of specific tRNA-derived RNA fragments, contributing to neurodegeneration.
- These accumulated RNA fragments, Ile-introns and 5' tRNA fragments, are implicated in the pathogenesis of PCH10.
- The CLP1 R140H mouse model offers valuable insights into neurodegenerative diseases linked to tRNA metabolism dysfunction.
