Abstract
Background
Carbonic anhydrase II deficiency syndrome is an autosomal recessive osteopetrosis with renal tubular acidosis and cerebral calcifications. We tried to detect the causative mutation for carbonic anhydrase II deficiency syndrome in a five-generation Chinese family.
Materials and Methods
Genomic DNA was extracted from whole blood of the proband, his grandmother, parents, aunt, uncle and sister. The exomes were sequenced by whole exon sequencing followed by genetic analysis and Sanger sequencing validation. Then, physical and chemical properties studies and structure analysis were performed on mutated protein. Finally, Minigene model of vector plasmids for wild type and mutant type was constructed and transfected into human embryonic kidney 293T cells to further explore the expression change of CA2 transcript and protein after mutation.
Results
Sequencing and genetic analysis have revealed the homozygous nonsense mutation of CA2 gene (c.368G > A, p.W123X) in the exon 4 of chromosome 8 of the proband, while it was not found in his grandmother, parents, aunt, uncle and sister. Furthermore, Sanger sequencing in the proband and his parents validated the mutation. Properties and structure of mutated CA2 proteins changed after mutation, especially in change of protein modification and hindrance of zinc ions binding, which may lead to decreased protein expression level of CA2.
Conclusions
We found a new homozygous nonsense mutation in CA2 gene (c.368G > A, p.W123X), which may be valuable in the early diagnosis and therapy of carbonic anhydrase II deficiency syndrome.
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Conception and design: Yi Yang; Administrative support: Pengqiu Li; Supply of materials and samples: Nie Tang; Data collection and collation: Yan Yang and Yi Yang; Data analysis and interpretation: Yan Yang, Yi Yang, Ying Zhu, Lei Zhang, Xu Cao, Limei Liu and Wei Xia. All authors approve and revise the manuscript.
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This study was approved by the ethics committee of Sichuan Academy of Medical Sciences & Sichuan Provincial People’s Hospital (2017 − 175).
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Supplementary Information
Supplementary Figure 1
Agarose gel electrophoresis of PCR product of wild type and mutant type CA2 mRNA. The sequence of PCR product electrophoresis was marker, normal control (1), wild type (2) and mutant type (3). DNA Marker is DM2000. From bottom to top is 100, 250, 500, 750, 1000 and 2000bp successively. Among which, 750bp is the bright band. (PNG 889 kb)
Supplementary Figure 2
Protein expression detection of CA2 in wild type and mutant type HEK 293T cells. Photomicrographs show the distribution of CA2 protein (red) and DAPI-stained nuclei (blue). (PNG 385 kb)
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Yang, Y., Tang, N., Zhu, Y. et al. A novel homozygous nonsense mutation in the CA2 gene (c.368G>A, p.W123X) linked to carbonic anhydrase II deficiency syndrome in a Chinese family. Metab Brain Dis 36, 589–599 (2021). https://doi.org/10.1007/s11011-021-00677-9
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DOI: https://doi.org/10.1007/s11011-021-00677-9