Acromesomelic Dysplasia With Homozygosity for a Likely Pathogenic BMPR1B Variant: Postaxial Polydactyly as a Novel Clinical Finding.

BACKGROUND: Acromesomelic chondrodysplasias are a rare subgroup of the clinically and genetically heterogeneous osteochondrodysplasias that are characterised by abnormalities in the limb development and short stature. Here, we report a 2-year-old boy, offspring of consanguineous parents, with acromesomelic dysplasia and postaxial polydactyly in which exome sequencing identified a novel homozygous missense variant in BMPR1B. The patient showed skeletal malformation of both hands and feet that included complex brachydactyly with the thumbs most severely affected, postaxial polydactyly of both hands, shortened toes as well as a bilateral hypoplasia of the fibula. METHODS: Whole trio exome sequencing was conducted to identify potential genetic variants in the patient. RESULTS: The analysis identified the biallelic variant NM_001203.3:c.821A > G;p.(Gln274Arg) in BMPR1B, a gene encoding bone morphogenetic protein receptor 1B. CONCLUSION: The skeletal phenotype can be brought in line with the phenotypes of previously reported cases of BMPR1B-associated chondrodysplasias. However, the postaxial polydactyly described here is a novel clinical finding in a BMPR1B-related case; notably, it has previously been reported in other acromesomelic dysplasia cases caused by homozygous pathogenic variants in GDF5-a gene which encodes for growth differentiation factor 5, a high-affinity ligand to BMPR1B.

The First Patient with Tibial Hemimelia-Polysyndactyly-Triphalangeal Thumb Syndrome Caused by De Novo c.423+4916 T>C ZRS Variant: A Case Report.

Genetic variants in the zone of polarizing activity regulatory sequence (ZRS) that induce ectopic expression of the SHH gene have been associated with different ZRS-related phenotypes. We report the first patient with a de novo variant, c.423+4916 T>C, in ZRS (previously classified as a variant of uncertain significance) that causes tibial hemimelia-polysyndactyly-triphalangeal thumb syndrome (THPTTS). A two-month-old male patient presented with bilateral preaxial polydactyly, triphalangeal thumb, and tibial agenesis and was heterozygous for the variant c.423+4916T>C (neither of his parents was a carrier). The findings obtained from the family study were sufficient to reclassify the variant from "uncertain significance" to "likely pathogenic" according to three criteria from the American College of Medical Genetics and Genomics guidelines, as follows: (1) absence of gnomAD, (2) confirmation of paternity and maternity, and (3) strong phenotype-genotype association. In ZRS-associated syndromes, a wide clinical spectrum has been observed, ranging from polydactyly to THPTTS; our patient has the most severe and rare phenotype. We did not perform functional assays. However, the c.423+4916T>C variant is flanked by three variants, which have been proven not only to cause the phenotype but also to increase the expression of SHH. Through all this data gathering, we consider the c.423+4916T>C variant to be causative of THPTTS.

Cant1 Affects Cartilage Proteoglycan Properties: Aggrecan and Decorin Characterization in a Mouse Model of Desbuquois Dysplasia Type 1.

Desbuquois dysplasia type 1 (DBQD1) is a recessive chondrodysplasia caused by mutations in the CANT1 gene, encoding for the Golgi Calcium-Activated Nucleotidase 1 (CANT1). The enzyme hydrolyzes UDP, the by-product of glycosyltransferase reactions, but it might play other roles in different cell types. Using a Cant1 knock-out mouse, we demonstrated that CANT1 is crucial for glycosaminoglycan (GAG) synthesis; however, its impact on the biochemical properties of cartilage proteoglycans remains unknown. Thus, in this work, we characterized decorin and aggrecan from primary chondrocyte cultures and cartilage biopsies of mutant mice at post-natal day 4 by Western blots and further investigated their distribution in the cartilage extracellular matrix (ECM) by immunohistochemistry. We demonstrated that the GAG synthesis defect caused by CANT1 impairment led to the synthesis and secretion of proteoglycans with shorter GAG chains compared with wild-type animals. However, this alteration did not result in the synthesis and secretion of decorin and aggrecan in the unglycanated form. Interestingly, the defect was not cartilage-specific since also skin decorin showed a reduced hydrodynamic size. Finally, immunohistochemical studies in epiphyseal sections of mutant mice demonstrated that the proteoglycan structural defect moderately affected decorin distribution in the ECM.

Identification of truncated variants in GLI family zinc finger 3 (GLI3) associated with polydactyly.

BACKGROUND: Polydactyly is a prevalent congenital anomaly with an incidence of 2.14 per 1000 live births in China. GLI family zinc finger 3 (GLI3) is a classical causative gene of polydactyly, and serves as a pivotal transcription factor in the hedgehog signaling pathway, regulating the development of the anterior-posterior axis in limbs. METHODS: Three pedigrees of polydactyly patients were enrolled from Hunan Province, China. Pathogenic variants were identified by whole-exome sequencing (WES) and Sanger sequencing. RESULTS: Three variants in GLI3 were identified in three unrelated families, including a novel deletion variant (c.1372del, p.Thr458GlnfsTer44), a novel insertion-deletion (indel) variant (c.1967_1968delinsAA, p.Ser656Ter), and a nonsense variant (c.2374 C > T, p.Arg792Ter). These variants were present exclusively in patients but not in healthy individuals. CONCLUSIONS: We identified three pathogenic GLI3 variants in polydactyly patients, broadening the genetic spectrum of GLI3 and contributing significantly to genetic counseling and diagnosis for polydactyly.

A novel missense variant located within the zinc finger domain of the GLI3 gene was identified in a Vietnamese pedigree with index finger polydactyly.

BACKGROUND: Polydactyly, particularly of the index finger, remains an intriguing anomaly for which no specific gene or locus has been definitively linked to this phenotype. In this study, we conducted an investigation of a three-generation family displaying index finger polydactyly. METHODS: Exome sequencing was conducted on the patient, with a filtration to identify potential causal variation. Validation of the obtained variant was conducted by Sanger sequencing, encompassing all family members. RESULTS: Exome analysis uncovered a novel heterozygous missense variant (c.1482A>T; p.Gln494His) at the zinc finger DNA-binding domain of the GLI3 protein within the proband and all affected family members. Remarkably, the variant was absent in unaffected individuals within the pedigree, underscoring its association with the polydactyly phenotype. Computational analyses revealed that GLI3 p.Gln494His impacts a residue that is highly conserved across species. CONCLUSION: The GLI3 zinc finger DNA-binding region is an essential part of the Sonic hedgehog signaling pathway, orchestrating crucial aspects of embryonic development through the regulation of target gene expression. This novel finding not only contributes valuable insights into the molecular pathways governing polydactyly during embryonic development but also has the potential to enhance diagnostic and screening capabilities for this condition in clinical settings.

Phenotypic and genotypic analysis of 11 fetal cases with Bardet-Biedl syndrome.

OBJECTIVE: To present the prenatal sonographic features and genomic spectrum of pregnancies with fetal Bardet-Biedl syndrome (BBS). METHODS: This was a retrospective study of 11 cases with BBS diagnosed by prenatal ultrasound and confirmed by genetic testing. Clinical and laboratory data were collected and reviewed for these cases, including maternal demographics, prenatal sonographic findings, molecular testing sequencing results, and pregnancy outcomes. RESULTS: All cases had unremarkable first-trimester ultrasound scans without reporting limb malformations. All had second-trimester abnormal ultrasounds: postaxial polydactyly in nine cases (9/11), renal abnormalities in seven (7/11), reduced amniotic fluid volume in two (2/11), central nervous system anomalies in two (2/11), and ascites in three (3/11). Ten fetuses presented with at least two-system anomalies, and one (Case 11) presented with only postaxial polydactyly. Variants were detected in five genes, including BBS2, ARL6/BBS3, BBS7, CEP290/BBS14 and IFT74/BBS22. Ten pregnancies were terminated in the second trimester, while one continued to term. CONCLUSION: Enlarged hyperechogenic kidneys and postaxial polydactyly are the two most common sonographic features of fetal BBS. Prenatal diagnosis of BBS can be done with ultrasound and genetic testing although the diagnosis may be made in the second trimester.

A novel homozygous FAM92A gene (CIBAR1) variant further confirms its association with non-syndromic postaxial polydactyly type A9 (PAPA9).

Polydactyly is a very common digit anomaly, having extra digits in hands and/or toes. Non-syndromic polydactyly in both autosomal dominant and autosomal recessive forms are caused by disease-causing variants in several genes, including GLI1, GLI3, ZNF141, FAM92A, IQCE, KIAA0825, MIPOL1, STKLD1, PITX1, and DACH1. Whole exome sequencing (WES) followed by bi-directional Sanger sequencing was performed for the single affected individual (II-1) of the family to reveal the disease causative variant/gene. 3D protein modeling and structural molecular docking was performed to determine the effect of the identified mutation on the overall protein structure. WES revealed a novel biallelic missense variant (c.472G>C; p.Ala158Pro) in exon 6 of the FAM92A gene. The identified variant segregated perfectly with the disease phenotype using Sanger sequencing. Furthermore, Insilco analysis revealed that the variant significantly changes the protein secondary structure, and substantially impact the stability of FAM92A. We report the second FAM92A disease-causing mutation associated with recessive non-syndromic postaxial polydactyly. The data further confirms the contribution of FAM92A in limb development and patterning.

A de novo Mutation (p.Gln277X) of Cyclin D2 is Responsible for a Child with Megalencephaly-Polymicrogyria-Polydactyly-Hydrocephalus Syndrome.

Megalencephaly-polymicrogyria-polydactyly-hydrocephalus syndrome (MPPH), a type of overgrowth syndrome, is characterized by progressive megalencephaly, cortical brain malformations, and distal limb anomalies. Previous studies have revealed that the overactivity of the phosphatidylinositol 3-kinase-Protein kinase B pathway and the increased cyclin D2 (CCND2) expression were the main factors contributing to this disease. Here, we present the case of a patient who exhibited megalencephaly, polymicrogyria, abnormal neuronal migration, and developmental delay. Serum tandem mass spectrometry and chromosome examination did not detect any metabolic abnormalities or copy number variants. However, whole-exome sequencing and Sanger sequencing revealed a de novo nonsense mutation (NM_001759.3: c.829C>T; p.Gln277X) in the CCND2 gene of the patient. Bioinformatics analysis predicted that this mutation may disrupt the structure and surface charge of the CCND2 protein. This disruption could potentially prevent polyubiquitination of CCND2, leading to its resistance against degradation. Consequently, this could drive cell division and growth by altering the activity of key cell cycle regulatory nodes, ultimately contributing to the development of MPPH. This study not only presents a new case of MPPH and expands the mutation spectrum of CCND2 but also enhances our understanding of the mechanisms connecting CCND2 with overgrowth syndromes.

Single-cell transcriptome analysis reveals characteristic transcription factors in polydactyly.

OBJECTIVE: The study aims to provide guidance on the identification of multiple-digit malformations as potential biomarkers and therapeutic targets. MATERIALS AND METHODS: Single-cell RNA sequencing (scRNA-seq) data of four multiple-finger malformation samples were downloaded from the GEO public database. Fibroblasts and keratinocytes were divided into cellular subpopulations and the transcription factors of different subpopulations were analyzed. The regulatory network of transcription factors and their target genes were constructed to analyze the functionality of regulons. RESULTS: Examination of the transcriptional profile data from 11,806 single cells uncovered significant associations between regulons and cell function in polydactyly. Specifically, the analysis highlighted the involvement of HOX family members and GLI2 transcription factors, including HOXD13, MSX2, LHX2, EMX2, LEF1, CREB3L2, and LHX2, in the polydactyly process within fibroblast cells. Furthermore, it sheds light on the roles of HES2 and GLIS1 in the formation and development of keratinocytes. CONCLUSIONS: Significant presence of transcription factors, especially HOXD13, MSX2, and LHX2, may be strongly related to the development of polydactyly.

Retinoblastoma and polydactyly in a child with 46, XY, 15pstk+ karyotype-A case report and literature review.

BACKGROUND: Retinoblastoma (Rb) is the most common intraocular malignancy in childhood, originating from primitive retinal stem cells or cone precursor cells. It can be triggered by mutations of the RB1 gene or amplification of the MYCN gene. Rb may rarely present with polydactyly. METHODS: We conducted karyotype analysis, copy number variation sequencing, and whole-genome sequencing on the infant proband and his family. The clinical course and laboratory results of the proband's infant were documented and collected. We also reviewed the relevant literature. RESULTS: A 68-day-old boy presented with preaxial polydactyly and corneal edema. His intraocular pressure (IOP) was 40/19 mmHg, and color Doppler imaging revealed vitreous solid mass-occupying lesions with calcification in the right eye. Ocular CT showed flaky high-density and calcification in the right eye. This was classified as an International Retinoblastoma Staging System group E retinoblastoma with an indication for enucleation. Enucleation and orbital implantation were performed on the child's right eye. Karyotype analysis revealed an abnormal 46, XY, 15pstk+ karyotype, and the mother exhibited diploidy of the short arm of chromosome 15. The Alx-4 development factor, 13q deletion syndrome, and the PAPA2 gene have been reported as potential mechanisms for Rb combined with polydactyly. CONCLUSION: We report the case of a baby boy with Rb and polydactyly exhibiting a 46, XY, 15pstk+ Karyotype. We discuss potential genetic factors related to both Rb and polydactyly. Furthermore, there is a need for further exploration into the impact of chromosomal polymorphisms in Rb with polydactyly.

PRKACA-related, atrial defects-polydactyly-multiple congenital malformation syndrome in an Indian patient.

PRKACA-related, atrial defects-polydactyly-multiple congenital malformation syndrome is a recently described skeletal ciliopathy, which is caused by disease-causing variants in PRKACA. The primary phenotypic description includes atrial septal defects, and limb anomalies including polydactyly and short limbs. To date, only four molecularly proven patients have been reported in the literature with a recurrent variant, c.409G>A p.Gly137Arg in PRKACA. In this study, we report the fifth affected individual with the same variant and review the clinical features and radiographic findings of this rare syndrome.

Increasing Sufu gene dosage reveals its unorthodox role in promoting polydactyly and medulloblastoma tumorigenesis.

Suppressor of fused (SUFU) is widely regarded as a key negative regulator of the sonic hedgehog (SHH) morphogenic pathway and a known tumor suppressor of medulloblastoma (MB). However, we report here that SUFU expression was markedly increased in 75% of specimens compiled in a tissue array comprising 49 unstratified MBs. The SUFU and GLI1 expression levels in this MB array showed strong positive correlation, which was also identified in a large public data set containing 736 MBs. We further report that increasing Sufu gene dosage in mice caused preaxial polydactyly, which was associated with the expansion of the Gli3 domain in the anterior limb bud and heightened Shh signaling responses during embryonic development. Increasing Sufu gene dosage also led to accelerated cerebellar development and, when combined with ablation of the Shh receptor encoded by Patched1 (Ptch1), promoted MB tumorigenesis. These data reveal multifaceted roles of SUFU in promoting MB tumorigenesis by enhancing SHH signaling. This revelation clarifies potentially counterintuitive clinical observation of high SUFU expression in MBs and may pave way for novel strategies to reduce or reverse MB progression.

Whole genome resequencing reveals candidate genes for postaxial polydactyly in Large White pigs.

Polydactyly is a genetic abnormality that affects both pig welfare and industry profits. Despite efforts to explore the genetic basis of pig polydactyly, progress remains limited. In this study, we analyzed a group of Large White pigs with postaxial polydactyly, including 29 cases and 79 controls from 24 families. High-depth sequencing was performed on 20 pigs, while low-depth sequencing was improved through imputation for the remaining pigs. A genome-wide association study (GWAS) and genetic differentiation were conducted using the resequencing dataset, resulting in the identification of 48 significantly associated SNPs and 27 candidate regions. The genetic differentiation regions on chromosomes 5 and 18, which harbored GWAS-identified SNPs, were delineated as confidence regions. The confidence region at Chr18: 1.850-1.925 Mb covers the fifth intron of LMBR1, a gene that contains an important regulatory element for SHH, known as ZRS. Mutations in this ZRS have been found to cause polydactyly in animals and humans. Therefore, we propose LMBR1 as a prospective candidate gene for postaxial polydactyly. These findings emphasize the importance of exploring the role of ZRS within LMBR1 in the pathogenesis of polydactyly in pigs.

Affinity-optimizing enhancer variants disrupt development.

Enhancers control the location and timing of gene expression and contain the majority of variants associated with disease1-3. The ZRS is arguably the most well-studied vertebrate enhancer and mediates the expression of Shh in the developing limb4. Thirty-one human single-nucleotide variants (SNVs) within the ZRS are associated with polydactyly4-6. However, how this enhancer encodes tissue-specific activity, and the mechanisms by which SNVs alter the number of digits, are poorly understood. Here we show that the ETS sites within the ZRS are low affinity, and identify a functional ETS site, ETS-A, with extremely low affinity. Two human SNVs and a synthetic variant optimize the binding affinity of ETS-A subtly from 15% to around 25% relative to the strongest ETS binding sequence, and cause polydactyly with the same penetrance and severity. A greater increase in affinity results in phenotypes that are more penetrant and more severe. Affinity-optimizing SNVs in other ETS sites in the ZRS, as well as in ETS, interferon regulatory factor (IRF), HOX and activator protein 1 (AP-1) sites within a wide variety of enhancers, cause gain-of-function gene expression. The prevalence of binding sites with suboptimal affinity in enhancers creates a vulnerability in genomes whereby SNVs that optimize affinity, even slightly, can be pathogenic. Searching for affinity-optimizing SNVs in genomes could provide a mechanistic approach to identify causal variants that underlie enhanceropathies.

Prenatal Ultrasound Diagnosis of Megalencephaly-Polymicrogyria-Polydactyly-Hydrocephalus Syndrome with Persistent Hyperplastic Primary Vitreous: A Case Report.

INTRODUCTION: Megalencephaly-polymicrogyria-polydactyly-hydrocephalus (MPPH) syndrome is a rare autosomal dominant disorder characterized by megalencephaly (i.e., overgrowth of the brain), polymicrogyria, focal hypoplasia of the cerebral cortex, and polydactyly. Persistent hyperplastic primary vitreous (PHPV) involves a spectrum of congenital ocular abnormalities that are characterized by the presence of a vascular membrane behind the lens. CASE PRESENTATION: Here, we present a case of foetal MPPH with PHPV that was diagnosed using prenatal ultrasound. Ultrasound revealed the presence of megalencephaly, multiple cerebellar gyri, and hydrocephalus. Whole-exome sequencing confirmed the mutation of the AKT3 gene, which led to the consideration of MPPH syndrome. Moreover, an echogenic band with an irregular surface was observed between the lens and the posterior wall of the left eye; therefore, MPPH with PHPV was suspected. CONCLUSION: MPPH syndrome with PHPV can be diagnosed prenatally.

Compound heterozygous variants in RAB34 in a rare skeletal ciliopathy syndrome.

Skeletal ciliopathies are a heterogenous group of congenital disorders characterized by multiple internal abnormalities, and distinct radiographic presentation. Pathogenic variants in at least 30 cilia genes are known to cause skeletal ciliopathies. Here we report a fetus with an atypical skeletal ciliopathy phenotype and compound heterozygous variants in the RAB34 gene. The affected fetus had multiple malformations, including posterior neck edema, micrognathia, low-set and small ears, auricular hypoplasia, cleft lip and palate, short extremities, and a combination of rarely occurring pre- and postaxial polydactyly. Genome sequencing identified compound heterozygous variants in the RAB34 gene: maternal c.254T>C, p.(Ile85Thr), and paternal c.691C>T, p.(Arg231*) variants. Only the paternal variant was present in the unaffected sibling. Evidence in the literature indicated that Rab34-/- mice displayed a ciliopathy phenotype with cleft palate and polydactyly. These features were consistent with malformations detected in our patient supporting the pathogenicity of the identified RAB34 variants. Overall, this case report further expands genetic landscape of human ciliopathy syndromes and suggests RAB34 as a candidate gene for skeletal ciliopathies.

Variants in EFCAB7 underlie nonsyndromic postaxial polydactyly.

Polydactyly is the most common limb malformation that occurs in 1.6-10.6 per one thousand live births, with incidence varying with ancestry. The underlying gene has been identified for many of the ~100 syndromes that include polydactyly. While for the more common form, nonsydromic polydactyly, eleven candidate genes have been reported. We investigated the underlying genetic cause of autosomal recessive nonsyndromic postaxial polydactyly in four consanguineous Pakistani families. Some family members with postaxial polydactyly also present with syndactyly, camptodactyly, or clinodactyly. Analysis of the exome sequence data revealed two novel homozygous frameshift deletions in EFCAB7: [c.830delG;p.(Gly277Valfs*5)]; in three families and [c.1350_1351delGA;p.(Asn451Phefs*2)] in one family. Sanger sequencing confirmed that these variants segregated with postaxial polydactyly, i.e., family members with postaxial polydactyly were found to be homozygous while unaffected members were heterozygous or wild type. EFCAB7 displays expressions in the skeletal muscle and on the cellular level in cilia. IQCE-EFCAB7 and EVC-EVC2 are part of the heterotetramer EvC complex, which is a positive regulator of the Hedgehog (Hh) pathway, that plays a key role in limb formation. Depletion of either EFCAB7 or IQCE inhibits induction of Gli1, a direct Hh target gene. Variants in IQCE and GLI1 have been shown to cause nonsyndromic postaxial polydactyly, while variants in EVC and EVC2 underlie Ellis van Creveld and Weyers syndromes, which include postaxial polydactyly as a phenotype. This is the first report of the involvement of EFCAB7 in human disease etiology.

Gain-of-function MYCN causes a megalencephaly-polydactyly syndrome manifesting mirror phenotypes of Feingold syndrome.

MYCN, a member of the MYC proto-oncogene family, regulates cell growth and proliferation. Somatic mutations of MYCN are identified in various tumors, and germline loss-of-function variants are responsible for Feingold syndrome, characterized by microcephaly. In contrast, one megalencephalic patient with a gain-of-function variant in MYCN, p.Thr58Met, has been reported, and additional patients and pathophysiological analysis are required to establish the disease entity. Herein, we report two unrelated megalencephalic patients with polydactyly harboring MYCN variants of p.Pro60Leu and Thr58Met, along with the analysis of gain-of-function and loss-of-function Mycn mouse models. Functional analyses for MYCN-Pro60Leu and MYCN-Thr58Met revealed decreased phosphorylation at Thr58, which reduced protein degradation mediated by FBXW7 ubiquitin ligase. The gain-of-function mouse model recapitulated the human phenotypes of megalencephaly and polydactyly, while brain analyses revealed excess proliferation of intermediate neural precursors during neurogenesis, which we determined to be the pathomechanism underlying megalencephaly. Interestingly, the kidney and female reproductive tract exhibited overt morphological anomalies, possibly as a result of excess proliferation during organogenesis. In conclusion, we confirm an MYCN gain-of-function-induced megalencephaly-polydactyly syndrome, which shows a mirror phenotype of Feingold syndrome, and reveal that MYCN plays a crucial proliferative role, not only in the context of tumorigenesis, but also organogenesis.