Novel Insights into Pontocerebellar Hypoplasia Type 3: Discovery of a New Disease-causing PCLO Variant and Development of a CRISPR-generated Cell Model

Authors

  • Maryam Baneshi 1-Department of Medical Genetics, Shiraz University of Medical Sciences, Shiraz, Iran /2-Department of Medical Biotechnology, School of Advanced Medical Sciences and Technologies, Shiraz University of Medical Sciences, Shiraz, Iran
  • Sedigheh Mohammadi Department of Medical Genetics, Shiraz University of Medical Sciences, Shiraz, Iran
  • Hossein Jafari Khamirani 1-Department of Medical Genetics, Shiraz University of Medical Sciences, Shiraz, Iran /2-Student Research Committee, Shiraz University of Medical Sciences, Shiraz, Iran
  • Jafar Fallahi Department of Molecular Medicine, Shiraz University of Medical Sciences, Shiraz, Iran
  • Mahintaj Dara Stem Cells Technology Research Center, Shiraz University of Medical Sciences, Shiraz, Iran
  • Fatemeh Sadat Tabei Student Research Committee, Shiraz University of Medical Sciences, Shiraz, Iran
  • Maryam Ranjbar Department of Medical Biotechnology, School of Advanced Medical Sciences and Technologies, Shiraz University of Medical Sciences, Shiraz, Iran
  • Seyed Mohammad Bagher Tabei 1-Department of Medical Genetics, Shiraz University of Medical Sciences, Shiraz, Iran 2-Maternal-fetal Medicine Research Center, Shiraz University of Medical Sciences, Shiraz, Iran

DOI:

https://doi.org/10.31661/gmj.v14i.3727

Keywords:

PCLO; Piccolo; Pontocerebellar Hypoplasia Type 3; Mutation; CRISPR/Cas9

Abstract

Background: Pathogenic variations in the PCLO gene cause Pontocerebellar Hypoplasia type 3 (PCH3), an extremely rare autosomal recessive disease characterized by seizure, intellectual disability, developmental delay, and microcephaly. PCLO encodes the Piccolo protein, which plays a critical role in synaptic function and neurological disorders. To date, only one pathogenic PCLO variant associated with PCH3 has been reported in the literature. While research on PCH3 is ongoing, the rarity of the condition has limited the number of studies. Materials and Methods: A novel homozygous variant in PCLO (NM_033026: c.458T>C, p. Met153Thr) was identified through wholeexome sequencing and confirmed by Sanger sequencing. Functional studies were conducted to assess the pathogenicity of this variant using next-generation sequencing (NGS), in silico analysis, CRISPR-edited cells, and real-time PCR. Results: The proband presented with seizure, microcephaly, mild ataxia, and behavioral issues. Notably, in addition to previously reported symptoms, the patient also exhibited toe-walking, loss of tendon reflexes, and unilateral paralysis. The PCLO knockout cell model and molecular analysis confirmed the loss of function of the Piccolo protein in the homozygous variant. Our findings also demonstrated that Piccolo deficiency may affect the expression of other genes, including CtBp1 and BSN. Conclusion: We identified a novel PCLO variant responsible for PCH3 in a second known family worldwide. Additionally, a CRISPR-based cell model for PCH3 was developed, providing a valuable foundation for further research into the molecular mechanisms underlying Piccolo function and disease pathogenesis.

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Published

2025-08-10

How to Cite

Baneshi, M., Mohammadi, S., Jafari Khamirani, H., Fallahi, J., Dara, M., Tabei , F. S., … Tabei, S. M. B. (2025). Novel Insights into Pontocerebellar Hypoplasia Type 3: Discovery of a New Disease-causing PCLO Variant and Development of a CRISPR-generated Cell Model. Galen Medical Journal, 14, e3727. https://doi.org/10.31661/gmj.v14i.3727

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