Triphalangeal Thumb-polysyndactyly Syndrome Research Articles

The pZRS non-coding regulatory mutation resulting in triphalangeal thumb-polysyndactyly syndrome changes the pattern of local interactions.

Herein, we report on a large Polish family presenting with a classical triphalangeal thumb-polysyndactyly syndrome (TPT-PS). This rare congenital limb anomaly is generally caused by microduplications encompassing the Sonic Hedgehog (SHH) limb enhancer, termed the zone of polarizing activity (ZPA) regulatory sequence (ZRS). Recently, a pathogenic variant in the pre-ZRS (pZRS), a conserved sequence located near the ZRS, has been described in a TPT-PS Dutch family. We performed targeted ZRS sequencing, array comparative genomic hybridization, and whole-exome sequencing. Next, we sequenced the recently described pZRS region. Finally, we performed a circular chromatin conformation capture-sequencing (4C-seq) assay on skin fibroblasts of one affected family member and control samples to examine potential alterations in the SHH regulatory domain and functionally characterize the identified variant. We found that all affected individuals shared a recently identified pathogenic point mutation in the pZRS region: NC_000007.14:g.156792782C>G (GRCh38/hg38), which is the same as in the Dutch family. The results of 4C-seq experiments revealed increased interactions within the whole SHH regulatory domain (SHH-LMBR1 TAD) in the patient compared to controls. Our study expands the number of TPT-PS families carrying a pathogenic alteration of the pZRS and underlines the importance of routine pZRS sequencing in the genetic diagnostics of patients with TPT-PS or similar phenotypes. The pathogenic mutation causative for TPT-PS in our patient gave rise to increased interactions within the SHH regulatory domain in yet unknown mechanism.

The ZPA Regulatory Sequence (ZRS) Hand2 and Twist1 Binding Sites are Not Necessary for ZRS Activity

Anterior-posterior patterning in the developing limb is regulated by Sonic Hedgehog (Shh), which is expressed in the Zone of Polarizing Activity (ZPA). Shh is regulated by a limb-specific enhancer known as the ZPA Regulatory Sequence (ZRS). Mutations in the highly conserved ZRS have been linked to limb malformations in humans. Single nucleotide variants (SNVs) cause digit duplications, such as preaxial polydactyly (PPD) and triphalangeal thumb-polysyndactyly syndrome (TPTPS). The ZRS mechanism is not well understood. Conditional knockout of Hand2 results in a delta-Shh phenotype, indicating the importance of Hand2 in Shh regulation. Hand2 and Twist1 are basic helix-loop-helix (bHLH) transcription factors, capable of homo- and hetero-dimerization, that bind E-boxes (CANNTG) with some promiscuity. The objective of the present study was to determine whether the three ZRS E-boxes are necessary for activity. We hypothesize that altering E-boxes will disrupt ZRS function. Enhancer activity was evaluated using a ZRS-GFP reporter plasmid. We generated a construct with all three E-boxes altered using site-directed mutagenesis. Constructs were electroporated into presumptive forelimb buds in Hamburger-Hamilton stage 14 chicken embryos. Enhancer activity was recorded using fluorescent microscopy. Surprisingly, modification of the Hand2 and Twist1 binding sites did not affect ZRS activity. Future work is needed to identify what transcription factors are indispensable for ZRS regulation of Shh.

A 300-kb microduplication of 7q36.3 in a patient with triphalangeal thumb-polysyndactyly syndrome combined with congenital heart disease and optic disc coloboma: a case report

BackgroundTriphalangeal thumb-polysyndactyly syndrome (TPT-PS) is a rare well-defined autosomal dominant disorder characterized by long thumbs with three phalanges combined with pre- and postaxial polydactyly/syndactyly of limbs.By now, the syndrome has been reported in several large families from different ethnic backgrounds, with a high degree of inter- and intrafamilial variability. The genome locus responsible for TPT-PS has been mapped to the 7q36.3 region harboring a long-range sonic hedgehog (SHH) regulatory sequence (ZRS). Both single-nucleotide variants and complete duplications of ZRS were shown to cause TPT-PS and similar limb phenotypes. TPT-PS usually forms as isolated limb pathology not associated with additional malformations, in particular, with cardiovascular abnormalities.Case presentationHere we report on a rare Russian neonatal case of TPT-PS combined with severe congenital heart disease, namely double outlet right ventricle, and microphthalmia with optic disc coloboma. Pedigree analysis revealed TPT-PS of various expressivity in 10 family members throughout five generations, while the cardiac defect and the eye pathology were detected only in the proband. To extend the knowledge on genotype–phenotype spectrum of TPT-PS, the careful clinical and genomic analysis of the family was performed. High-resolution array-based comparative genomic hybridization (array-CGH) revealed a ~ 300 kb microduplication of 7q36.3 locus (arr[GRCh37] 7q36.3(156385810_156684811) × 3) that co-segregated with TPT-PS in the proband and her mother. The duplication encompassed three genes including LMBR1, the intron 5 of which is known to harbor ZRS. Based on whole-exome sequencing data, no additional pathogenic mutations or variants of uncertain clinical significance were found in morbid cardiac genes or genes associated with a microphthalmia/anophthalmia/coloboma spectrum of ocular malformations.ConclusionsThe results support the previous data, indicating that complete ZRS duplication underlies TPT-PS, and suggest a broader phenotypic impact of the 7q36.3 microduplication. Potential involvement of the 7q36.3 microduplication in the patient’s cardiac and eye malformations is discussed. However, the contribution of some additional genetic/epigenetic factors to the complex patient`s phenotype cannot be excluded entirely. Further comprehensive functional studies are needed to prove the possible involvement of the 7q36.3 locus in congenital heart disease and eye pathology.

Large duplication in LMBR1 gene in a large Chinese pedigree with triphalangeal thumb polysyndactyly syndrome.

Polydactyly and syndactyly are digital abnormalities in limb-associated birth defects usually caused by genetic disorders. In this study, a five-generation Chinese pedigree was found with triphalangeal thumb polysyndactyly syndrome (TPTPS), showing an autosomal dominant pattern of inheritance. We utilized linkage analysis and whole genome sequencing (WGS) for the genetic diagnosis of this pedigree. Linkage analysis was performed using a genome-wide single nucleotide polymorphism (SNP) chip and three genomic regions were identified in chromosomes 2, 6, and 7 with significant linkage signals. WGS discovered a copy number variation (CNV) mutation caused by a large duplication region at the tail of chromosome 7 located in exons 1-5 of the LMBR1 gene, including the zone of polarizing activity regulatory sequence (ZRS), with a length of approximately 180 kb. A real-time polymerase chain reaction (PCR) assay confirmed the duplication. The findings of our study supported the notion that large duplications including the ZRS caused TPTPS. Our study showed that linkage analysis in combination with WGS could successfully identify the disease locus and causative mutation in TPTPS, which could help elucidate the molecular mechanisms and genotype-phenotype correlations in polydactyly.

A point mutation in the pre-ZRS disrupts sonic hedgehog expression in the limb bud and results in triphalangeal thumb–polysyndactyly syndrome

PurposeThe zone of polarizing activity regulatory sequence (ZRS) is an enhancer that regulates sonic hedgehog during embryonic limb development. Recently, mutations in a noncoding evolutionary conserved sequence 500 bp upstream of the ZRS, termed the pre-ZRS (pZRS), have been associated with polydactyly in dogs and humans. Here, we report the first case of triphalangeal thumb–polysyndactyly syndrome (TPT-PS) to be associated with mutations in this region and show via mouse enhancer assays how this mutation leads to ectopic expression throughout the developing limb bud. MethodsWe used linkage analysis, whole-exome sequencing, Sanger sequencing, fluorescence in situ hybridization, multiplex ligation-dependent probe amplification, single-nucleotide polymorphism array, and a mouse transgenic enhancer assay. ResultsTen members of a TPT-PS family were included in this study. The mutation was linked to chromosome 7q36 (LOD score 3.0). No aberrations in the ZRS could be identified. A point mutation in the pZRS (chr7:156585476G>C; GRCh37/hg19) was detected in all affected family members. Functional characterization using a mouse transgenic enhancer essay showed extended ectopic expression dispersed throughout the entire limb bud (E11.5). ConclusionOur work describes the first mutation in the pZRS to be associated with TPT-PS and provides functional evidence that this mutation leads to ectopic expression of this enhancer within the developing limb.

An increased duplication of ZRS region that caused more than one supernumerary digits preaxial polydactyly in a large Chinese family.

Preaxial polydactyly (PPD) is inherited in an autosomal dominant fashion and characterized by the presence of one or more supernumerary digits on the thumb side. It had been identified that point mutation or genomic duplications of the long-range limb-specific cis-regulator - zone of polarizing activity regulatory sequence (ZRS) cause PPD or other limb deformities such as syndactyly type IV (SD4) and Triphalangeal thumb-polysyndactyly syndrome (TPTPS). Most previously reported cases involved with no more than one extra finger; however, the role of the point mutation or genomic duplications of ZRS in the case of more than one redundant finger polydactyly remains unclear. In this article, we reported a family case of more than one redundant finger polydactyly on the thumb side for bilateral hands with a pedigree chart of the family. Results of quantitative PCR (qPCR) and sequence analysis suggested that the relative copy number (RCN) of ZRS but not point mutation (including insertion and deletion) was involved in all affected individuals.

Mutation analysis of a Chinese pedigree with triphalangeal thumb-polysyndactyly syndrome

Triphalangeal thumb-polysyndactyly syndrome (TPTPS) is an autosomal dominant limb disorder with triphalangeal thumbs, polysyndactyly, and syndactyly. In this study, we describe a four-generation Han Chinese family with eight affected members. Haplotype analysis, Affymetrix SNP 6.0 arrays, qPCR, and gap-PCR were performed. Haplotyping results linked the disease-causing region to the 7q36 region that includes the zone of polarizing activity-regulatory sequence. A 442-kb duplication was found on chromosome 7 that co-segregated with the disease phenotype. The extent of the duplication was determined by qPCR, and the breakpoints were identified by gap-PCR and direct sequencing. This mutation was not detected in normal members in the same family. Our data therefore suggest that this novel microduplication, between 155,913,768 and 156,355,553 bp on chromosome 7, could be considered the cause of TPTPS in this kindred.

Confirmation of genetic homogeneity of syndactyly type IV and triphalangeal thumb–polysyndactyly syndrome in a Chinese family and review of the literature

Syndactyly type IV (SD4) is inherited in an autosomal dominant fashion and characterized by complete cutaneous syndactyly of all fingers accompanied with polydactyly. Triphalangeal thumb-polysyndactyly syndrome (TPTPS) consists of a triphalangeal thumb, polydactyly, and syndactyly and is transmitted in an autosomal dominant manner with variable expression. Genomic duplications of the long-range limb-specific cis-regulator (ZRS) cause SD4 and TPTPS. Here, we report two individuals from a Chinese family with syndactyly. One individual had overlapping clinical symptoms of TPTPS and SD4, while the other had a typical SD4 with postaxial polydactyly of the toe. Results of quantitative PCR suggested that the duplication of ZRS involved all affected individuals, and array comparative genomic hybridization detected its size as 115.3 kb. This work confirms the genetic homogeneity of SD4 and TPTPS. Our result expands the spectrum of ZRS duplications. TPTPS and SD4 should be considered as a continuum of phenotypes.

A specific mutation in the distant sonic hedgehog (SHH)cis-regulator (ZRS) causes Werner mesomelic syndrome (WMS) while complete ZRS duplications underlie Haas type polysyndactyly and preaxial polydactyly (PPD) with or without triphalangeal thumb

Werner mesomelic syndrome (WMS) is an autosomal dominant disorder with unknown molecular etiology characterized by hypo- or aplasia of the tibiae in addition to the preaxial polydactyly (PPD) of the hands and feet and/or five-fingered hand with absence of thumbs. We show that point mutations of a specific nucleotide within the sonic hedgehog (SHH) regulatory region (ZRS) cause WMS. In a previously unpublished WMS family, we identified the causative G>A transition at position 404 of the ZRS, and in six affected family members of a second WMS family we found a 404G>C mutation of the ZRS. The 404G>A ZRS mutation is known as the "Cuban mutation" of PPD type II (PPD2). Interestingly, the index patient of that family had tibial hypoplasia as well. These data provide the first evidence that WMS is caused by a specific ZRS mutation, which leads to strong ectopic SHH expression. In contrast, we show that complete duplications of the ZRS region lead to type Haas polysyndactyly or triphalangeal thumb-polysyndactyly syndrome, but do not affect lower limb development. We suggest the term "ZRS-associated syndromes" and a clinical subclassification for the continuum of limb malformations caused by different molecular alterations of the ZRS.

Triphalangeal thumb–polysyndactyly syndrome and syndactyly type IV are caused by genomic duplications involving the long range, limb-specific SHH enhancer

Background:The Sonic hedgehog (SHH) protein produced in the zone of polarising activity (ZPA) is a major determinant of the identity and numbers of digits in early limb development. Preaxial polydactyly...

A microduplication of the long range SHH limb regulator (ZRS) is associated with triphalangeal thumb-polysyndactyly syndrome

Background:Sonic hedgehog (SHH) plays an important role in defining the anterior–posterior axis in the developing limbs. A highly conserved non-coding sequence about ∼ 1 Mb upstream from the sonic hedgehog...

A single C to T transition in intron 5 of LMBR1 gene is associated with triphalangeal thumb-polysyndactyly syndrome in a Chinese family

Triphalangeal thumb-polysyndactyly syndrome (TPT-PS) is a type of human hand-foot malformation. In this study, we collected data from a Chinese family with TPT-PS and mapped the disease region to chromosome 7q36. By using a fine mapping study and a haplotype analysis, we narrowed the affected region to 1.7 cM between markers D7S2465 and D7S2423, which contains four candidate genes: HLXB9, LMBR1, NOM1, and RNF32. By sequence analysis, we found no sequence alterations, which are specific to the patients in the transcribed regions and in the intron–exon boundaries among the four genes. After closely examining intron 5 of the LMBR1 gene, we discovered a single C to T transition in the affected TPT-PS individuals of the Chinese subject family. The position of this C to T transition is located close to other sequence alterations involved in several preaxial polydactyly (PPD) families, supporting the notion that intron 5 of LMBR1 contains a cis-acting regulator of limb-specific Sonic Hedgehog ( SHH). We postulate that the disruption of this cis-regulator via a single C to T transition results in the dysregulation of SHH, which leads to the TPT-PS found in this case.

Werner mesomelic dysplasia with Hirschsprung disease

Mesomelic dysplasia type Werner is defined by absence of tibiae and preaxial polysyndactyly of hands and feet. Occasional findings are triphalangeal thumbs, absence of patella(e), and dislocated fibula(e). The molecular basis is unknown and autosomal dominant inheritance with variable expressivity is currently postulated. Hirschsprung disease was reported previously in one case. We report here on a new case of mesomelic dysplasia Werner type associated with Hirschsprung disease and bilateral cryptorchidism. We discuss the overlap with the triphalangeal thumb polysyndactyly syndrome located in chromosome 7q36.

Laurin-Sandrow syndrome with additional associated manifestations.

A Thai man with Laurin-Sandrow syndrome (LSS, MIM 135750), the ninth reported case, is described. He had an underdeveloped nasal bone, scar-like seams under the nose, large heads of mandibular condyles, and brachymesophalangy of toes as newly observed findings of the syndrome. He also had mental retardation. The patient had duplication of ulna, with triphalangeal thumbs, and polydactyly of one finger. The triphalangeal thumbs were non-opposable. Carpal bones were malformed. Mirror image polydactyly of the toes was present. There were nine toes on the right and eight on the left. Joint abnormalities were observed at his elbows, wrists, knees, ankles, fingers, and toes. Synostosis of severely malformed tarsal bones was noted. This appears to be the first case of LSS with anomalies not limited to the nose and limbs. The relationship between LSS, tibial hemimelia-polysyndactyly-triphalangeal thumbs syndrome, triphalangeal thumb-polysyndactyly syndrome, preaxial polydactyly types 2 and 3, and Haas-type syndactyly is discussed.

Phenotypic variability of triphalangeal thumb-polysyndactyly syndrome linked to chromosome 7q36

Triphalangeal thumb-polysyndactyly (TPT-PS) is an isolated limb malformation consisting of pre- and postaxial polysyndactyly of hands and feet. The only family reported so far is of Dutch origin, and the genetic mapping study localized the TPT-PS locus at chromosome region 7q36 where the isolated triphalangeal thumb (TPT) anomaly has also been mapped. It was suggested that TPT-PS is a phenotypic variation of isolated TPT, and the same ancestral mutation may produce both phenotypes. Here we report on the second family with this malformation from the Turkish population. The characteristic findings in this family are triphalangeal thumb, webbing between 3rd, 4th, and 5th fingers associated with bony synostosis in the distal phalanges of the same fingers, and pre- and postaxial polysyndactyly of feet. Some individuals show a more severe phenotype with a complete syndactyly of all fingers giving a "cup-like" appearance to the hands. Genetic linkage study with DNA markers D7S1823, D7S550, D7S559, and D7S2423 demonstrated that this family is also linked to chromosome band 7q36. Identification of a second family from a distinct ethnic background suggests that TPT-PS and isolated TPT are not caused by the same ancestral mutation as it was originally anticipated.