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HGNC Approved Gene Symbol: CLP1
SNOMEDCT: 782720005;
Cytogenetic location: 11q12.1 Genomic coordinates (GRCh38): 11:57,657,762-57,661,865 (from NCBI)
| Location | Phenotype |
Phenotype MIM number |
Inheritance |
Phenotype mapping key |
|---|---|---|---|---|
| 11q12.1 | Pontocerebellar hypoplasia, type 10 | 615803 | Autosomal recessive | 3 |
The CLP1 gene encodes a component of the tRNA endonuclease complex (TSEN), which removes introns from pre-tRNA transcripts. CLP1 is a multifunctional kinase implicated in tRNA, mRNA, and siRNA maturation (summary by Karaca et al., 2014 and Schaffer et al., 2014).
By cloning and sequencing a complex rearrangement involving chromosomes 10 and 11, Tanabe et al. (1996) obtained a partial genomic clone of HEAB. By 5-prime and 3-prime RACE of several cDNA libraries, they obtained a full-length cDNA. The 3-prime UTR contains 2 putative polyadenylation sites. The deduced 425-amino acid protein is valine- and leucine-rich and contains a domain sharing similarity with the ATP/GTP-binding domain of ABC transporters. It also has a possible tyrosine kinase phosphorylation site. Northern blot analysis detected a major 2.0-kb transcript that was expressed ubiquitously, with highest expression in testis and skeletal muscle. A 3.2-kb transcript was less abundant in many tissues, with highest expression in peripheral blood leukocytes and thymus.
De Vries et al. (2000) purified pre-mRNA cleavage factor IIm (CFIIm) of the 3-prime end processing complex (see 604978) from HeLa cell nuclear extracts and determined that it contains PCF11 (608876) and CLP1. By searching databases and PCR of a HeLa cell cDNA library, they cloned CLP1. The deduced protein contains highly conserved Walker A and B motifs, which have been implicated in ATP/GTP binding.
De Vries et al. (2000) found that immunodepletion of CLP1 reduced pre-mRNA cleavage activity, but not polyadenylation activity, in purified HeLa cell CFIIm. CLP1 interacted with CFIm and cleavage-polyadenylation specificity factor (CPSF; see 606027), suggesting that it bridges these two 3-prime end processing factors within the cleavage complex.
Paushkin et al. (2004) identified and characterized the human tRNA splicing endonuclease (see SEN2; 608753). They found that human endonuclease complexes are associated with pre-mRNA 3-prime end processing factors, including CLP1. Small interfering RNA-mediated depletion of SEN2 led to defects in maturation of both pre-tRNA and pre-mRNA. These findings demonstrated a link between pre-tRNA splicing and pre-mRNA 3-prime end formation, suggesting that the endonuclease subunits function in multiple RNA processing events.
Weitzer and Martinez (2007) applied a chromatographic approach that resulted in the identification of the human protein CLP1, a component of both tRNA splicing and mRNA 3-prime-end formation machineries, as the RNA kinase responsible for phosphorylation of the 5-prime end of siRNAs necessary for their subsequent incorporation into the RISC complex (RNA-induced silencing complex). Weitzer and Martinez (2007) reported that the kinase CLP1 phosphorylates and licenses synthetic siRNAs to become assembled into RISC for subsequent target RNA cleavage. More importantly, Weitzer and Martinez (2007) demonstrated that the physiologic role of CLP1 as the RNA kinase that phosphorylates the 5-prime end of the 3-prime exon during human tRNA splicing, allowing the subsequent ligation of both exon halves by an unknown tRNA ligase.
Using in vitro RNA kinase and small interfering RNA (siRNA) efficiency assays, Fujinami et al. (2020) demonstrated that Clp1, rather than Nol9 (620304), was the main RNA kinase in mice.
By somatic cell hybrid analysis and FISH, Tanabe et al. (1996) mapped the CLP1 gene to chromosome 11q12.
Tanabe et al. (1996) identified an invins(10;11)(p12;q23q12) and other complex chromosomal rearrangements in a 2-year old boy with acute monoblastic leukemia (AML-M5). Cloning of the proximal 10p breakpoint showed that the MLL gene (159555) at chromosome 11q23 was fused to the 3-prime portion of AF10 (MLLT10; 602409) at chromosome 10p12. Cloning of the telomeric 10p junction revealed that the 5-prime portion of AF10 was fused with the HEAB gene. The 5-prime AF10/HEAB fusion transcript was out of frame.
Simultaneously and independently, Karaca et al. (2014) and Schaffer et al. (2014) identified the same homozygous missense mutation in the CLP1 gene (R140H; 608757.0001) in affected members of 9 consanguineous Turkish families with a neurodevelopmental and neurodegenerative disorder called pontocerebellar hypoplasia type 10 (PCH10; 615803). Haplotype analysis indicated a founder effect. The patients were severely affected with delayed or absent psychomotor development, microcephaly, spasticity, seizures, and inability to walk. Brain imaging showed variable abnormalities, including cortical dysgenesis, pontocerebellar atrophy or hypoplasia, and thin corpus callosum. The patients reported by Karaca et al. (2014) had an axonal sensorimotor peripheral neuropathy. In vitro functional expression studies by both groups showed that the mutation caused a loss of kinase and cleavage activity and impaired function of the TSEN complex, resulting in an abnormal cellular accumulation of unspliced pre-tRNAs.
In 2 distantly related girls from a Turkish family with PCH10, Wafik et al. (2018) identified homozygosity for the founder R140H mutation in the CLP1 gene. The mutation was found by trio-based whole-exome sequencing and confirmed by Sanger sequencing. Both sets of parents were heterozygous for the mutation.
Hanada et al. (2013) generated kinase-dead Clp1 mice that showed a progressive loss of spinal motor neurons associated with axonal degeneration in the peripheral nerves and denervation of neuromuscular junctions, resulting in impaired motor function, muscle weakness, paralysis, and fatal respiratory failure. Transgenic rescue experiments showed that Clp1 functions in motor neurons. Mechanistically, loss of Clp1 activity results in accumulation of a novel set of small RNA fragments, derived from aberrant processing of tyrosine pre-transfer RNA. These tRNA fragments sensitize cells to oxidative stress-induced p53 (191170) activation and p53-dependent cell death. Genetic inactivation of p53 rescues Clp1 kinase-dead mice from the motor neuron loss, muscle denervation, and respiratory failure. Hanada et al. (2013) concluded that their experiments uncovered a mechanistic link between tRNA processing, formation of a new RNA species, and progressive loss of lower motor neurons regulated by p53.
Karaca et al. (2014) observed that kinase-dead Clp1 mice had reduced brain weight and volume, as well as reduced cortical thickness and reduced numbers of neurons compared to wildtype, consistent with microcephaly. There was an increase in cell death among neuronal progenitor cells, which resulted in reduced numbers of cortical neurons. Cortical layering was normal, and cerebellar volume was not affected.
Schaffer et al. (2014) found that zebrafish homozygous for a null clp1 mutation did not survive beyond 5 days postfertilization and showed abnormal swimming behavior, abnormal head shape, and curved tail, suggesting a neuromotor defect. Mutant zebrafish larvae showed progressive neuronal loss, particularly in the forebrain and hindbrain, as well as altered motor neuron morphology. The loss was shown to be p53-dependent.
In 11 affected children from 5 consanguineous Turkish families with pontocerebellar hypoplasia (PCH10; 615803), Karaca et al. (2014) identified a homozygous c.419G-A transition in the CLP1 gene, resulting in an arg140-to-his (R140H) substitution at a highly conserved residue. The mutation, which was found in the first 2 families by whole-exome sequencing, was not present in the 1000 Genomes Project or Exome Variant Server databases or in 2,500 in-house control exomes. The mutation was identified by Sanger sequencing and segregated with the disorder in 3 additional families with a similar phenotype. Haplotype analysis indicated a founder effect for the 5 families. In vitro functional expression assays in E. coli showed that the R140H mutant protein retained some RNA kinase activity, but did not interact with other members of the tRNA splicing endonuclease (TSEN) complex, resulting in decreased pre-tRNA cleavage activity and abnormal cellular accumulation of linear tRNA introns, although pre- and mature tRNA levels were largely unaffected. Patient cultured fibroblasts showed decreased RNA kinase activity and only minor detectable pre-tRNA cleavage activity.
Simultaneously and independently, Schaffer et al. (2014) identified a homozygous R140H mutation in affected members of 4 consanguineous Turkish families with PCH10. The mutation, which was found by exome sequencing and confirmed by Sanger sequencing, segregated with the disorder in the families and was not present in public databases. It was observed twice in heterozygosity among more than 2,000 in-house exomes, yielding a carrier frequency of 1:1,100. Haplotype analysis indicated a founder effect. Recombinant R140H CLP1 showed defective kinase activity that was reduced by more than half of wildtype levels. Induced neurons derived from patient fibroblasts showed reduced nuclear localization of mutant CLP1, also suggesting impaired function. Northern blot analysis identified accumulation of pre-tRNAs and depletion of mature tRNAs specifically in patient-induced neurons. These abnormalities were associated with loss of CLP1 affinity for the TSEN complex and with impaired pre-tRNA cleavage activity. Patient cells showed increased sensitivity to oxidative stress-induced death exacerbated by the addition of unphosphorylated 3-prime-tRNA exon halves.
In 2 distantly related girls (proband and her first cousin once removed) from a Turkish family with PCH10, Wafik et al. (2018) identified homozygosity for the founder R140H mutation in the CLP1 gene. The mutation was found by trio-based whole-exome sequencing and confirmed by Sanger sequencing. Both sets of parents were heterozygous for the mutation.
de Vries, H., Ruegsegger, U., Hubner, W., Friedlein, A., Langen, H., Keller, W. Human pre-mRNA cleavage factor IIm contains homologs of yeast proteins and bridges two other cleavage factors. EMBO J. 19: 5895-5904, 2000. [PubMed: 11060040] [Full Text: https://doi.org/10.1093/emboj/19.21.5895]
Fujinami, H., Shiraishi, H., Hada, K., Inoue, M., Morisaki, I., Higa, R., Shin, T., Kobayashi, T., Hanada, R., Penninger, J. M., Mimata, H., Hanada, T. CLP1 acts as the main RNA kinase in mice. Biochem. Biophys. Res. Commun. 525: 129-134, 2020. [PubMed: 32081435] [Full Text: https://doi.org/10.1016/j.bbrc.2020.02.066]
Hanada, T., Weitzer, S., Mair, B., Bernreuther, C., Wainger, B. J., Ichida, J., Hanada, R., Orthofer, M., Cronin, S. J., Komnenovic, V., Minis, A., Sato, F., and 12 others. CLP1 links tRNA metabolism to progressive motor-neuron loss. Nature 495: 474-480, 2013. [PubMed: 23474986] [Full Text: https://doi.org/10.1038/nature11923]
Karaca, E., Weitzer, S., Pehlivan, D., Shiraishi, H., Gogakos, T., Hanada, T., Jhangiani, S. N., Wiszniewski, W., Withers, M., Campbell, I. M., Erdin, S., Isikay, S., and 33 others. Human CLP1 mutations alter tRNA biogenesis, affecting both peripheral and central nervous system function. Cell 157: 636-650, 2014. [PubMed: 24766809] [Full Text: https://doi.org/10.1016/j.cell.2014.02.058]
Paushkin, S. V., Patel, M., Furia, B. S., Peltz, S. W., Trotta, C. R. Identification of a human endonuclease complex reveals a link between tRNA splicing and pre-mRNA 3-prime end formation. Cell 117: 311-321, 2004. [PubMed: 15109492] [Full Text: https://doi.org/10.1016/s0092-8674(04)00342-3]
Schaffer, A. E., Eggens, V. R. C., Caglayan, A. O., Reuter, M. S., Scott, E., Coufal, N. G., Silhavy, J. L., Xue, Y., Kayserili, H., Yasuno, K., Rosti, R. O., Abdellateef, M., and 21 others. CLP1 founder mutation links tRNA splicing and maturation to cerebellar development and neurodegeneration. Cell 157: 651-663, 2014. [PubMed: 24766810] [Full Text: https://doi.org/10.1016/j.cell.2014.03.049]
Tanabe, S., Bohlander, S. K., Vignon, C. V., Espinosa, R., III, Zhao, N., Strissel, P. L., Zeleznik-Le, N. J., Rowley, J. D. AF10 is split by MLL and HEAB, a human homolog to a putative Caenorhabditis elegans ATP/GTP-binding protein in an invins(10;11)(p12;q23q12). Blood 88: 3535-3545, 1996. [PubMed: 8896421]
Wafik, M., Taylor, J., Lester, T., Gibbons, R. J., Shears, D. J. 2 new cases of pontocerebellar hypoplasia type 10 identified by whole exome sequencing in a Turkish family. Europ. J. Med. Genet. 61: 273-279, 2018. [PubMed: 29307788] [Full Text: https://doi.org/10.1016/j.ejmg.2018.01.002]
Weitzer, S., Martinez, J. The human RNA kinase hClp1 is active on 3-prime transfer RNA exons and short interfering RNAs. Nature 447: 222-226, 2007. [PubMed: 17495927] [Full Text: https://doi.org/10.1038/nature05777]