Identification of variants in ACAN and PAPSS2 leading to spondyloepi(meta)physeal dysplasias in four Chinese families

Abstract

Background: Spondyloepi(meta)physeal dysplasias (SE[M]D) are a group of inherited skeletal disorders that mainly affect bone and cartilage, and next-generation sequencing has aided the detection of genetic defects of such diseases. In this study, we aimed to identify causative variants in four Chinese families associated with SE(M)D.

Methods: We recruited four unrelated Chinese families all displaying short stature and growth retardation. Clinical manifestations and X-ray imaging were recorded for all patients. Candidate variants were identified by whole-exome sequencing (WES) and verified by Sanger sequencing. Pathogenicity was assessed by conservation analysis, 3D protein modeling and in silico prediction, and was confirmed according to American College of Medical Genetics and Genomics.

Results: Three novel SE(M)D-related variants c.1090dupG, c.7168 T > G, and c.2947G > C in ACAN, and one reported variant c.712C > T in PAPSS2 were identified. Among them, c.1090dupG in ACAN and c.712C > T in PAPSS2 caused truncated protein and the other two variants led to amino acid alterations. Conservation analysis revealed sites of the two missense variants were highly conserved, and bioinformatic findings confirmed their pathogenicity. 3D modeling of mutant protein encoded by c.7168 T > G(p.Trp2390Gly) in ACAN proved the structural alteration in protein level.

Conclusion: Our data suggested ACAN is a common pathogenic gene of SE(M)D. This study enriched the genetic background of skeletal dysplasias, and expanded the mutation spectra of ACAN and PAPSS2.

Keywords: bioinformatic analysis; short stature; spondyloepimetaphyseal dysplasia; spondyloepiphyseal dysplasia; variation; whole-exome sequencing.

FIGURE 1

Pedigrees of four Chinese families with SE(M)D. Squares represent males and circles represent females. Affected members are in black symbols, and unaffected individuals are in open symbols. Symbols with half black indicate the carriers, and the arrows are pointed to the probands. Height (cm), genotypes, and whether WES or Sanger sequencing were performed were labeled next to the members with available data

FIGURE 2

Clinical characteristics of the four probands. (a and b) Metatarsal dysplasia, phalangeal dysplasia, and metacarpal dysplasia in proband 1; (c and d) Decreased bone density in epiphyses and underdevelopment of first distal phalanx in hand in proband 2. (e–g) Short and disproportionate upper limbs (e), and blurring growth plate region in proband 3. (h–j) Short stature (h), abnormal trochlea phalanges (i), and short middle/distal phalanx (j) in proband 4

FIGURE 3

Sequencing analysis of four variants associated with SE(M)D. Sequencing analysis of all available members in families 1–4. (a) In family 1, II‐3, III‐1, and IV‐1 were heterozygous of c.1090dupG(p.Val364Glyfs*4) in ACAN. (b) In family 2, III‐1, III‐3, IV‐1, IV‐2, and V‐1 were heterozygous of c.7168 T > G(p.Trp2390Gly) in ACAN. (c) In family 3, III‐1 was homozygous of c.2947G > C(p.Glu983Gln) in ACAN, while parents II‐1, II‐2, and grandfathers I‐1, I‐3 were heterozygous of the variant. (d) In family 4, II‐1 and II‐2 were homozygous of c.712C > T(p.Arg238*) in PAPSS2, and the parents (I‐1, I‐2) were heterozygous of the variant. Red arrows point to the mutation sites

FIGURE 4

Bioinformatic prediction by conservation analysis and 3D protein structure. (a) Structure of aggrecan containing the main domains that are drawn to scale is shown above. Structure of the ACAN is shown below, and the numbers indicate the corresponding exons. Variants reported in this study are labeled in red. G, globular domain; IGD, interglobular domain; KS, keratan sulfate attachment region; CS, chondroitin sulfate attachment region; EGF, epidermal growth factor‐like domain; CLD, C‐type lectin domain; CRP, complement regulatory‐like domain. (b) Sequence alignment of two missense variants in ACAN was analyzed by comparing the amino acid sequences in human, mouse, cow, pig, rabbit, and monkey. (c–e) Overall structure of the ACAN protein (c), tertiary structure of the wild‐type (d), and p.Trp2390Gly mutant protein (e)