Novel mutations in TPM2 and PIEZO2 are responsible for distal arthrogryposis (DA) 2B and mild DA in two Chinese families

Abstract

Background: Distal arthrogryposis (DA) is a group of clinically and genetically heterogeneous disorders that involve multiple congenital limb contractures and comprise at least 10 clinical subtypes. Here, we describe our findings in two Chinese families: Family 1 with DA2B (MIM 601680) and Family 2 with mild DA.

Methods: To map the disease locus, two-point linkage analysis was performed with microsatellite markers closed to TPM2, TNNI2/TNNT3 and TNNC2. In Family 1, a positive LOD (logarithm of odds) score was only obtained at the microsatellite marker close to TPM2 and mutation screening was performed using direct sequencing of TPM2 in the proband. In Family 2, for the LOD score that did not favor linkage to any markers, whole-exome sequencing (WES) was performed on the proband. PCR-restriction fragment length polymorphism (RFLP) and bioinformatics analysis were then applied to identify the pathogenic mutations in two families. In order to correlate genotype with phenotype in DA, retrospective analyses of phenotypic features according to the TPM2 and PIEZO2 mutation spectrums were carried out.

Results: A heterozygous missense mutation c.308A > G (p.Q103R) in TPM2 in Family 1, and a novel variation c.8153G > A (p.R2718Q) in PIEZO2 in Family 2 were identified. Each of the two novel variants was co-segregated with the DA manifestations in the corresponding family. Bioinformatics analysis from several tools supported the pathogenicity of the mutations. Furthermore, our study suggests that there is no relation between the types or locations of TPM2 mutations and the clinical characteristics, and that different inheritance modes and mutation types concerning PIEZO2 cause distinct clinical manifestations.

Conclusions: We report two novel mutations within TPM2 and PIEZO2 responsible for DA2B and mild DA in two Chinese families, respectively. Our study expands the spectrum of causal mutations in the TPM2 and PIEZO2 genes.

Keywords: Distal arthrogryposis; Genotype–phenotype; Novel mutation; PIEZO2; TPM2.

Fig. 1

Pedigrees of the two families with DA2B (Family 1) and mild DA1 (Family 2). Arrows indicate the probands in each family

Fig. 2

Clinical features of patients in Family 1 with DA2B (a-h) and Family 2 with mild DA (i-k). a Ulnar deviation and contractures in patient II5. b Camptodactyly and adducted thumbs in patient III1. c Flexed toes and talipes equinovarus in patient II5. d Dysplastic ear and attached ear lobes in patient II5. e Small mouth with limited opening in patient II5. f Downward-slanting palpebral fissures, a small chin, and deep folds in the nasolabial area and forehead in patient II5. g Flexion contracture yielding stiff elbows in patient II5. h Short stature of patient II5 (unaffected II6 [left], 178 cm; patient II5 [right], 156.5 cm). i Camptodactyly and ulnar deviation in patient II5. j Mild contractures of the fingers in patient II2. k X-ray findings of patient III1 in 5 years old indicate camptodactyly and ulnar deviation

Fig. 3

Identification of a missense mutation in TPM2 in Family 1. a Genetic linkage analysis was carried out with 3 microsatellites, and TPM2 was identified as the candidate gene. b Sequencing results indicate the heterozygous mutation c.308A > G in exon 3 of TPM2. c DNA fragments from the affected individuals (II5, III1, III3, IV1, and IV4). Three fragments were separated by electrophoresis (394 bp, 264 bp, and 130 bp). d Multiple-species sequence alignment shows the evolutionary conservation of position p.Q103 in TPM2. e The schematic of TPM2 mutation spectrum on functional module. Mutation-related diseases are exhibited in different literal colors

Fig. 4

Identification of a missense mutation in PIEZO2 in affected members of Family 2. a The novel mutation c.8153G > A (p.R2718Q) in PIEZO2 verified by Sanger sequencing. b PCR-RFLP findings with Taq I indicate two DNA fragments in affected patients (II2, III1, II5, and I1; 104 bp and 70 bp) but not in unaffected family members. c Multiple-species sequence alignment shows the evolutionary conservation of position p.R718 in PIEZO2. d The schematic of PIEZO2 mutation spectrum on functional module. Mutation-related diseases are exhibited in different literal colors

Fig. 5

SWISS-MODEL prediction of the p.Q103R mutation in tropomyosin. a The 3-dimensional structure of wild-type tropomyosin with position 103 indicated by a black arrow. Nearby acidic (red) and basic (blue) surface properties are illustrated with the neutral glutamine residue shown as a yellow line. b Acidic surface properties are destroyed with the substitution of basic arginine for glutamine. The substitution is depicted as a blue line

Fig. 6

Three substitutions in position 2718 of PIEZO2 are simulated by means of SWISS-MODEL and are represented with Ribbon. a The wild-type amino acid at position 2718 is depicted, with H-bonds shown by yellow dashed lines. b The p.R2718Q substitution maintains the most parts of H-bonding interactions. c Leucine tends to lose H-bond with D2713, and the nearby α-helical structure is disrupted. d Substitution with proline eliminates most of the H-bonds with neighboring amino acids