PMC7262964_01
(A) Pedigree of the HSAN-VI family with segregation analysis. The black symbols represent the affected members, and the arrow indicates the proband. Red text represents the variants. (B-G) Aspects of the phenotype of the proband. The proband has an ankle ulcer (B), necrosis of the astragalus (C), moderate scoliosis (D), phalangeal joint contracture (E,F), and cataract (G).
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F1
We identified a Chinese family with HSAN (Figure 1A). He consistently could not perceive noxious thermal/gelid stimuli or innocuous warm stimuli, which had resulted in the ankle ulcer and necrosis of the astragalus (Figures 1B,C). The proband had moderate scoliosis (with a Cobb angle of 33 ) without centrum malformation (Figure 1D). The brothers presented with arthrogryposes of the fingers, foot extroversion, knee recurvation deformities, and myasthenia; hence, they achieved independent ambulation at 3 years of age (Figures 1E,F). The proband had thin blue scleras and corneal ulcers and was subsequently diagnosed with a cataract (Figure 1G). HSAN-VI patients, suggesting changes in cell adhesion and the cytoskeleton, which is in accordance with the expectation that DST variants can alter cytoskeletal filament networks (Figure 2E and Supplementary Figure 1). However, we did not observe any change in DST expression by RNAseq or qPCR (Figure 2E).
(A,B) Sequencing results of the DST variants. Sequence chromatograms indicate compound heterozygous variants (c.3304G>A, p.V1102I and c.13796G>A, p.R4599H) in the family with HSAN-VI. (C,D) DST peptide sequences surrounding the mutated residues (p.V1102I and p.R4599H) with multiple interspecies alignments generated by ClustalX. The mutation position is indicated with a red arrow. (E) The results of qPCR. "***", statistical significance; "ns", no statistical significance. (F) The structures of three major tissue-specific dystonin isoforms. "ABD", an actin-binding domain; "plakin", a plakin domain; "rod", a rod domain; "PRD2", the plectin repeats; "spectrin repeats", spectrin repeats; and "TMBD", a combination of an EF hand domain and a growth arrest-specific 2 protein-related region. The red (V1102I) and gray (R4599H) arrows indicate the mutated AA site. (G) Schematic diagram of DST and the variants identified in HSAN-VI patients. Four putative promoters are indicated by horizontal arrows. Blue boxes indicate the unique exons of isoform-2, the red box indicates the unique exon of isoform-1, the green box indicates the unique exon of isoform-3, and the purple box indicates the unique exon of isoform-e. The blue line represents the transcript of isoform-2, the red line represents the transcript of isoform-1, the green line represents the transcript of isoform-3, and the purple line represents the transcript of isoform-e. Full vertical arrows show the positions of the variants of DST in the gene. The red text represents the present variants.
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F2
Sanger sequencing showed that the DST variant (c.3304G>A, p.V1102I) in the identical twins was inherited from their mother and the other (c.13796G>A, p.R4599H) was inherited from their father (Figures 2A,B). The amino acid sequence alignment analysis suggested that these two variants were both located in a highly evolutionarily conserved site (Figures 2C,D). HSAN-VI patients, suggesting changes in cell adhesion and the cytoskeleton, which is in accordance with the expectation that DST variants can alter cytoskeletal filament networks (Figure 2E and Supplementary Figure 1). However, we did not observe any change in DST expression by RNAseq or qPCR (Figure 2E).