Whole-exome sequencing identified a novel variant in an Iranian patient affected by pycnodysostosis

Abstract

Background: Whole-exome sequencing (WES) has emerged as a successful diagnostic tool in molecular genetics laboratories worldwide. In this study, we aimed to find the potential genetic cause of skeletal disease, a heterogeneous disease, revealing the obvious short stature phenotype. In an Iranian family, we used solo-WES in a suspected patient to decipher the potential genetic cause(s).

Methods: A comprehensive clinical and genotyping examination was applied to suspect the disease of the patient. The solo clinical WES was exploited, and the derived data were filtered according to the standard pipelines. In order to validate the WES finding, the region harboring the candidate variant in the CTSK gene was amplified from genomic DNA and sequenced directly by Sanger sequencing.

Results: Sequence analysis revealed a rare novel nonsense variant, p.(Trp320*); c.905G>A, in the CTSK gene (NM_000396.3). In silico analysis shed light on the contribution of the variant to the pathogenicity of pycnodysostosis. This variant was confirmed by Sanger sequencing and further clinical examinations of the patient confirmed the disease.

Conclusion: The present study shows a rare variant of the CTSK gene, which inherited as autosomal recessive, in an Iranian male patient with pycnodysostosis. Taken together, the novel nonsense CTSK variant meets the criteria of being likely pathogenic according to the American College of Medical Genetics and Genomics-the Association for Molecular Pathology (ACMG-AMP) variant interpretation guidelines.

Keywords: cathepsin K; nonsense variant; pycnodysostosis; rare disease; whole-exome sequencing.

Figure 1

(a) This pedigree is comprised of four generations. The arrow appoints the proband of the family. The genetic status is shown as heterozygote: W/c.905G>A or homozygote: c.905G>A/c.905G>A; in this pedigree, white symbols: unaffected; red symbol: affected; squares: men; circles: females; parallel lines: consanguineous marriage, W: wild‐type allele. (b) Schematic genetic and protein maps of the CTSK gene (NM_000396.3). c.905G>A variant is located in exon 8, which encodes the mature domain of CTSK protein. Similar to other most papain‐like cysteine proteases, CTSK contains 329 amino acids that can be categorized in three distinct sections: a 15‐amino acid preregion, a 99‐amino acid proregion, and a 215‐amino acid mature active enzyme (Toral‐López et al., 2011). The low‐PH environment changes inactive CTSK to the active form by the removal of the N‐terminal proregion. (c) UCSC database used to show the conservation of specific nucleotide (G; highlighted as red) including the variant site in vertebrates, particularly in primates. The amino acid sequence of CTSK colored based on conservation scores by the ConSurf database. Scores ranged from 1 to 9, where a score of 9 represented a highly conserved residue. ConSurf demonstrates evolutionary conservation profiles for proteins of known/unknown structure according to the phylogenetic relations between homologous sequences as well as amino acid's structural and functional importance. (d) The 3D structure of CTSK is shown. The picture was rendered with PyMOL (v.0.99rc6). The original site of Trp302 is emphasized by a highlighted zone and locally zoomed

Figure 2

(a) Schematic representation of filtering strategies exploited in this research. For more investigation, the filtering steps evaluated by regard to this fact that the disease could be engendered by autosomal dominant; however, we could detect no relevant variant according to this supposition. (b) Sequence chromatogram shows a homozygous state of the nucleotide sequence of c.905G>A. (c) Radiography shows the large head circumstance, obtuse mandibular angle, and scoliosis, which was evident in the patient. (d) Short fingers revealed by X‐ray and the dysplastic flat nails were evident in the patient (For detailed, please refer to Figure S1)