Scalp fibroblasts have a shared expression profile in monogenic craniosynostosis

BACKGROUND: Craniosynostosis can be caused by both genetic and environmental factors, the relative contributions of which vary between patients. Genetic testing identifies a pathogenic mutation or chromosomal abnormality in ∼ 21% of cases, but it is likely that further causative mutations remain to be discovered. OBJECTIVE: To identify a shared signature of genetically determined craniosynostosis by comparing the expression patterns in three monogenic syndromes with a control group of patients with non-syndromic sagittal synostosis. METHODS: Fibroblasts from 10 individuals each with Apert syndrome (FGFR2 substitution S252W), Muenke syndrome (FGFR3 substitution P250R), Saethre-Chotzen syndrome (various mutations in TWIST1) and non-syndromic sagittal synostosis (no mutation detected) were cultured. The relative expression of ∼ 47,000 transcripts was quantified on Affymetrix arrays. RESULTS: 435, 45 and 46 transcripts were identified in the Apert, Muenke and Saethre-Chotzen groups, respectively, that differed significantly from the controls. Forty-six of these transcripts were shared between two or more syndromes and, in all but one instance, showed the same direction of altered expression level compared with controls. Pathway analysis showed over-representation of the shared transcripts in core modules involving cell-to-cell communication and signal transduction. Individual samples from the Apert syndrome cases could be reliably distinguished from non-syndromic samples based on the gene expression profile, but this was not possible for samples from patients with Muenke and Saethre-Chotzen syndromes. CONCLUSIONS: Common modules of altered gene expression shared by genetically distinct forms of craniosynostosis were identified. Although the expression profiles cannot currently be used to classify individual patients, this may be overcome by using more sensitive assays and sampling additional tissues

The phenotype of MEGF8-related Carpenter syndrome (CRPT2) is refined through the identification of eight new patients

Carpenter syndrome (CRPTS) is a rare autosomal recessive condition caused by biallelic variants in genes that encode negative regulators of hedgehog signalling (RAB23 [CRPT1] or, more rarely, MEGF8 [CRPT2]), and is characterised by craniosynostosis, polysyndactyly, and other congenital abnormalities. We describe a further six families comprising eight individuals with MEGF8-associated CRPT2, increasing the total number of reported cases to fifteen, and refine the phenotype of CRPT2 compared to CRPT1. The core features of craniosynostosis, polysyndactyly and (in males) cryptorchidism are almost universal in both CRPT1 and CRPT2. However, laterality defects are present in nearly half of those with MEGF8-associated CRPT2, but are rare in RAB23-associated CRPT1. Craniosynostosis in CRPT2 commonly involves a single midline suture in comparison to the multi-suture craniosynostosis characteristic of CRPT1. No patient to date has carried two MEGF8 gene alterations that are both predicted to lead to complete loss-of-function, suggesting that a variable degree of residual MEGF8 activity may be essential for viability and potentially contributing to variable phenotypic severity. These data refine the phenotypic spectrum of CRPT2 in comparison to CRPT1 and more than double the number of likely pathogenic MEGF8 variants in this rare disorder

Reassessing the association: Evaluation of a polyalanine deletion variant of RUNX2 in non‐syndromic sagittal and metopic craniosynostosis

The RUNT‐related transcription factor RUNX2 plays a critical role in osteoblast differentiation, and alterations to gene dosage cause distinct craniofacial anomalies. Uniquely amongst the RUNT‐related family, vertebrate RUNX2 encodes a polyglutamine/polyalanine repeat (Gln23‐Glu‐Ala17 in humans), with the length of the polyalanine component completely conserved in great apes. Surprisingly, a frequent 6‐amino acid deletion polymorphism, p.(Ala84_Ala89)del, occurs in humans (termed 11A allele), and a previous association study (Cuellar et al. Bone 137:115395;2020) reported that the 11A variant was significantly more frequent in non‐syndromic sagittal craniosynostosis (nsSag; allele frequency [AF] = 0.156; 95% confidence interval [CI] 0.126–0.189) compared to non‐syndromic metopic craniosynostosis (nsMet; AF = 0.068; 95% CI 0.045–0.098). However, the gnomAD v.2.1.1 control population used by Cuellar et al. did not display Hardy–Weinberg equilibrium, hampering interpretation. To re‐examine this association, we genotyped the RUNX2 11A polymorphism in 225 individuals with sporadic nsSag as parent–child trios and 164 singletons with sporadic nsMet, restricting our analysis to individuals of European ancestry. We compared observed allele frequencies to the non‐transmitted alleles in the parent–child trios, and to the genome sequencing data from gnomAD v.4, which display Hardy–Weinberg equilibrium. Observed AFs (and 95% CI) were 0.076 (0.053–0.104) in nsSag and 0.082 (0.055–0.118) in nsMet, compared with 0.062 (0.042–0.089) in non‐transmitted parental alleles and 0.065 (0.063–0.067) in gnomAD v.4.0.0 non‐Finnish European control genomes. In summary, we observed a non‐significant excess, compared to gnomAD data, of 11A alleles in both nsSag (relative risk 1.18, 95% CI 0.83–1.67) and nsMet (relative risk 1.29, 95% CI 0.87–1.92), but we did not replicate the much higher excess of RUNX2 11A alleles in nsSag previously reported (p = 0.0001)

Disruption of TWIST1 translation by 5' UTR variants in Saethre-Chotzen syndrome

Saethre-Chotzen syndrome (SCS), one of the most common forms of syndromic craniosynostosis (premature fusion of the cranial sutures), results from haploinsufficiency of TWIST1, caused by deletions of the entire gene or loss-of-function variants within the coding region. To determine whether non-coding variants also contribute to SCS, we screened 14 genetically undiagnosed SCS patients using targeted capture sequencing, and identified novel single nucleotide variants (SNVs) in the 5' untranslated region (UTR) of TWIST1 in two unrelated SCS cases. We show experimentally that these variants, which create translation start sites in the TWIST1 leader sequence, reduce translation from the main open reading frame (mORF). This is the first demonstration that non-coding SNVs of TWIST1 can cause SCS, and highlights the importance of screening the 5' UTR in clinically diagnosed SCS patients without a coding mutation. Similar 5' UTR variants, particularly of haploinsufficient genes, may represent an under-ascertained cause of monogenic disease. This article is protected by copyright. All rights reserved

Identification of the first ATRIP-deficient patient and novel mutations in ATR define a clinical spectrum for ATR-ATRIP Seckel Syndrome

A homozygous mutational change in the Ataxia-Telangiectasia and RAD3 related (ATR) gene was previously reported in two related families displaying Seckel Syndrome (SS). Here, we provide the first identification of a Seckel Syndrome patient with mutations in ATRIP, the gene encoding ATR-Interacting Protein (ATRIP), the partner protein of ATR required for ATR stability and recruitment to the site of DNA damage. The patient has compound heterozygous mutations in ATRIP resulting in reduced ATRIP and ATR expression. A nonsense mutational change in one ATRIP allele results in a C-terminal truncated protein, which impairs ATR-ATRIP interaction; the other allele is abnormally spliced. We additionally describe two further unrelated patients native to the UK with the same novel, heterozygous mutations in ATR, which cause dramatically reduced ATR expression. All patient-derived cells showed defective DNA damage responses that can be attributed to impaired ATR-ATRIP function. Seckel Syndrome is characterised by microcephaly and growth delay, features also displayed by several related disorders including Majewski (microcephalic) osteodysplastic primordial dwarfism (MOPD) type II and Meier-Gorlin Syndrome (MGS). The identification of an ATRIP-deficient patient provides a novel genetic defect for Seckel Syndrome. Coupled with the identification of further ATR-deficient patients, our findings allow a spectrum of clinical features that can be ascribed to the ATR-ATRIP deficient sub-class of Seckel Syndrome. ATR-ATRIP patients are characterised by extremely severe microcephaly and growth delay, microtia (small ears), micrognathia (small and receding chin), and dental crowding. While aberrant bone development was mild in the original ATR-SS patient, some of the patients described here display skeletal abnormalities including, in one patient, small patellae, a feature characteristically observed in Meier-Gorlin Syndrome. Collectively, our analysis exposes an overlapping clinical manifestation between the disorders but allows an expanded spectrum of clinical features for ATR-ATRIP Seckel Syndrome to be define

Quantitative Control of Organ Shape by Combinatorial Gene Activity

A novel combination of molecular genetics, shape analysis, and computational modelling shows how the complex three-dimensional shape of the Snapdragon flower can arise through local gene activity

The Human Phenotype Ontology project:linking molecular biology and disease through phenotype data

The Human Phenotype Ontology (HPO) project, available at http://www.human-phenotype-ontology.org, provides a structured, comprehensive and well-defined set of 10,088 classes (terms) describing human phenotypic abnormalities and 13,326 subclass relations between the HPO classes. In addition we have developed logical definitions for 46% of all HPO classes using terms from ontologies for anatomy, cell types, function, embryology, pathology and other domains. This allows interoperability with several resources, especially those containing phenotype information on model organisms such as mouse and zebrafish. Here we describe the updated HPO database, which provides annotations of 7,278 human hereditary syndromes listed in OMIM, Orphanet and DECIPHER to classes of the HPO. Various meta-attributes such as frequency, references and negations are associated with each annotation. Several large-scale projects worldwide utilize the HPO for describing phenotype information in their datasets. We have therefore generated equivalence mappings to other phenotype vocabularies such as LDDB, Orphanet, MedDRA, UMLS and phenoDB, allowing integration of existing datasets and interoperability with multiple biomedical resources. We have created various ways to access the HPO database content using flat files, a MySQL database, and Web-based tools. All data and documentation on the HPO project can be found online

Mutations in fam20b and xylt1 Reveal That Cartilage Matrix Controls Timing of Endochondral Ossification by Inhibiting Chondrocyte Maturation

Differentiating cells interact with their extracellular environment over time. Chondrocytes embed themselves in a proteoglycan (PG)-rich matrix, then undergo a developmental transition, termed “maturation,” when they express ihh to induce bone in the overlying tissue, the perichondrium. Here, we ask whether PGs regulate interactions between chondrocytes and perichondrium, using zebrafish mutants to reveal that cartilage PGs inhibit chondrocyte maturation, which ultimately dictates the timing of perichondral bone development. In a mutagenesis screen, we isolated a class of mutants with decreased cartilage matrix and increased perichondral bone. Positional cloning identified lesions in two genes, fam20b and xylosyltransferase1 (xylt1), both of which encode PG synthesis enzymes. Mutants failed to produce wild-type levels of chondroitin sulfate PGs, which are normally abundant in cartilage matrix, and initiated perichondral bone formation earlier than their wild-type siblings. Primary chondrocyte defects might induce the bone phenotype secondarily, because mutant chondrocytes precociously initiated maturation, showing increased and early expression of such markers as runx2b, collagen type 10a1, and ihh co-orthologs, and ihha mutation suppressed early perichondral bone in PG mutants. Ultrastructural analyses demonstrated aberrant matrix organization and also early cellular features of chondrocyte hypertrophy in mutants. Refining previous in vitro reports, which demonstrated that fam20b and xylt1 were involved in PG synthesis, our in vivo analyses reveal that these genes function in cartilage matrix production and ultimately regulate the timing of skeletal development

De novo and rare inherited mutations implicate the transcriptional coregulator TCF20/SPBP in autism spectrum disorder

BACKGROUND: Autism spectrum disorders (ASDs) are common and have a strong genetic basis, yet the cause of ∼70-80% ASDs remains unknown. By clinical cytogenetic testing, we identified a family in which two brothers had ASD, mild intellectual disability and a chromosome 22 pericentric inversion, not detected in either parent, indicating de novo mutation with parental germinal mosaicism. We hypothesised that the rearrangement was causative of their ASD and localised the chromosome 22 breakpoints. METHODS: The rearrangement was characterised using fluorescence in situ hybridisation, Southern blotting, inverse PCR and dideoxy-sequencing. Open reading frames and intron/exon boundaries of the two physically disrupted genes identified, TCF20 and TNRC6B, were sequenced in 342 families (260 multiplex and 82 simplex) ascertained by the International Molecular Genetic Study of Autism Consortium (IMGSAC). RESULTS: IMGSAC family screening identified a de novo missense mutation of TCF20 in a single case and significant association of a different missense mutation of TCF20 with ASD in three further families. Through exome sequencing in another project, we independently identified a de novo frameshifting mutation of TCF20 in a woman with ASD and moderate intellectual disability. We did not identify a significant association of TNRC6B mutations with ASD. CONCLUSIONS: TCF20 encodes a transcriptional coregulator (also termed SPBP) that is structurally and functionally related to RAI1, the critical dosage-sensitive protein implicated in the behavioural phenotypes of the Smith-Magenis and Potocki-Lupski 17p11.2 deletion/duplication syndromes, in which ASD is frequently diagnosed. This study provides the first evidence that mutations in TCF20 are also associated with ASD