A lysyl oxidase-responsive collagen peptide illuminates collagen remodeling in wound healing.

Tissue repair and fibrosis involve the dynamic remodeling of collagen, and accurate detection of these sites is of utmost importance. Here, we use a collagen peptide sensor (1) to visualize collagen formation and remodeling during wound healing in mice and humans. We show that the probe binds selectively to sites of collagen formation and remodeling at different stages of healing. Compared to conventional methods, the peptide sensor localizes preferentially to areas of collagen synthesis and remodeling at the wound edge and not in matured fibrillar collagen. We also demonstrate its applicability for in vivo wound imaging and for discerning differential remodeling in wounds of transgenic mice with altered collagen dynamics. Our findings show the value of 1 as a diagnostic tool to rapidly identify the sites of matrix remodeling in tissue sections, which will aid in the conception of new therapeutic strategies for fibrotic disorders and defective tissue repair.

Regions of homozygosity and a novel variant in Steel syndrome: An added dilemma to diagnosis.

Steel syndrome is an autosomal recessive disorder that is caused by mutations in COL27A1 gene. The majority of reported cases have been of Puerto Rican origin, with few reports from India. The present case adds to the repertoire of homozygous recessive disorders from non-consanguineous Indian families. With the present case, a 4-year-old girl, we wish to signify that although mutations in several genes are known to cause skeletal abnormalities, identification of underlying mutations is important as it not only helps with the ascertainment of diagnosis but also aids in determining the role of surgical interventions which is particularly true for Steel syndrome, where the outcome of surgical intervention is usually dismal.

Molecular Basis of Pathogenic Variants in the Fibrillar Collagens.

The fibrillar collagen family is comprised of the quantitatively major types I, II and III collagens and the quantitatively minor types V and XI. These form heterotypic collagen fibrils (composed of more than a single collagen type) where the minor collagens have a regulatory role in controlling fibril formation and diameter. The structural pre-requisites for normal collagen biosynthesis and fibrillogenesis result in many places where this process can be disrupted, and consequently a wide variety of phenotypes result when pathogenic changes occur in these fibrillar collagen genes. Another contributing factor is alternative splicing, both naturally occurring and as the result of pathogenic DNA alterations. This article will discuss how these factors should be taken into account when assessing DNA sequencing results from a patient.

Mutations in COL1A1 and COL27A1 Associated with a Pectus Excavatum Phenotype in 2 Siblings with Osteogenesis Imperfecta.

BACKGROUND Osteogenesis imperfecta is a skeletal disease with a range of phenotypes, depending on the genetic mutation. Individuals with osteogenesis imperfecta type I often have mutations in COL1A genes. This disease can be associated with chest wall deformities such as pectus excavatum, but the number of patients with this presentation is limited, and genetic variants associated with this phenotype have not been reported. CASE REPORT We studied the Skeletal Disorders Genetic Panel of 2 siblings with osteogenesis imperfecta type I and severe pectus excavatum requiring surgical correction. Both had severe respiratory symptoms secondary to the chest wall deformity, and the male patient had evidence of mitral valve insufficiency on an echocardiogram. Results of the genetic panel were remarkable for a homozygous copy number gain in exons 2 to 51 in gene COL1A1. Additionally, both had a heterozygous pathogenic variant in exon 7 of gene COL27A1 (replacement of a glycine with arginine in codon 697 of the protein). CONCLUSIONS Gene COL27A1 plays a role during the calcification of cartilage to bone and is associated with Steel syndrome, a skeletal disorder mainly found in the Puerto Rican population. Heterozygous carriers of the p.Gly697Arg variant in COL27A1 have not been described to have a phenotype with chest wall deformities. Additionally, a genotype-phenotype relationship regarding pectus excavatum in patients with osteogenesis imperfecta has not been described, suggesting that having COL1A gene mutations and simultaneous haploinsufficiency of COL27A1 can result in a phenotype of osteogenesis imperfecta with pectus excavatum and predispose these patients to additional phenotypic features.

Steel syndrome: Report of three patients, including monozygotic twins and review of clinical and mutation profiles.

Steel syndrome (MIM# 615155) is an autosomal recessive skeletal disorder, characterized by dislocations of the hips and radial heads, carpal coalition, short stature, facial dysmorphism, and scoliosis. Until date 47 patients have been reported. However, disease causing variants have been identified only in twenty Puerto Rican and nine non-Puerto Rican families. Here we report two monozygotic twins and a boy from two families with novel missense variants, c.295G > A p.(Ala99 Thr), c.3056C > A p.(Pro1019His) and c.2521G > A p.(Gly841Arg) in COL27A1. We describe for the first time, cleft palate and delayed carpal bone ossification as features of Steel syndrome. We reviewed clinical features in all mutation-proven Steel syndrome patients. Short stature and dislocation/subluxation of hip joint are consistently observed. Other features include dislocated radial heads, scoliosis, lordosis, carpal coalition, facial dysmorphism, hearing loss, bilateral fifth finger clinodactyly, knee deformities and developmental delay. Seven missense variants and eight null variants are reported in COL27A1 until date. We also looked into the genotype-phenotype correlation in Puerto Rican and non-Puerto Rican patients.

Bioinformatics Analysis and Experimental Verification Identify Downregulation of COL27A1 in Poor Segmental Congenital Scoliosis.

BACKGROUND: Congenital scoliosis (CS) represents the congenital defect disease, and poor segmental congenital scoliosis (PSCS) represents one of its types. Delayed intervention can result in disability and paralysis. In this study, we would identify the core biomarkers for PSCS progression through bioinformatics analysis combined with experimental verification. METHODS: This work obtained the GSE11854 expression dataset associated with somite formation in the GEO database, which covers data of 13 samples. Thereafter, we utilized the edgeR of the R package to obtain DEGs in this dataset. Then, GO annotation, KEGG analyses, and DO annotation of DEGs were performed by "clusterProfiler" of the R package. This study performed LASSO regression for screening the optimal predicting factors for somite formation. Through RNA sequencing based on peripheral blood samples from healthy donors and PSCS cases, we obtained the RNA expression patterns and screen out DEGs using the R package DESeq2. The present work analyzed COL27A1 expression in PSCS patients by the RT-PCR assay. RESULTS: A total of 443 genes from the GSE11854 dataset were identified as DEGs, which were involved in BP associated with DNA replication, CC associated with chromosomal region, and MF associated with ATPase activity. These DEGs were primarily enriched in the TGF-ß signaling pathway and spinal deformity. Further, LASSO regression suggested that 9 DEGs acted as the signature markers for somite formation. We discovered altogether 162 DEGs in PSCS patients, which were involved in BP associated with cardiac myofibril assembly and MF associated with structural constituent of muscle. However, these 162 DEGs were not significantly correlated with any pathways. Finally, COL27A1 was identified as the only intersected gene between the best predictors for somite formation and PSCS-related DEGs, which was significantly downregulated in PSCS patients. CONCLUSION: This work sheds novel lights on DEGs related to the PSCS pathogenic mechanism, and COL27A1 is the possible therapeutic target for PSCS. Findings in this work may contribute to developing therapeutic strategies for PSCS.

Collagen fibers provide guidance cues for capillary regrowth during regenerative angiogenesis in zebrafish.

Although well investigated, the importance of collagen fibers in supporting angiogenesis is not well understood. In this study, we demonstrate that extracellular collagen fibers provide guidance cues for endothelial cell migration during regenerative angiogenesis in the caudal zebrafish fin. Inhibition of collagen cross-linking by ß-Aminopropionitrile results in a 70% shorter regeneration area with 50% reduced vessel growth and disintegrated collagen fibers. The disrupted collagen scaffold impedes endothelial cell migration and induces formation of abnormal angioma-like blood vessels. Treatment of the Fli//colRN zebrafish line with the prodrug Nifurpirinol, which selectively damages the active collagen-producing 1α2 cells, reduced the regeneration area and vascular growth by 50% with wider, but less inter-connected, capillary segments. The regenerated area contained larger vessels partially covered by endothelial cells embedded in atypical extracellular matrix containing cell debris and apoptotic bodies, macrophages and granulocytes. Similar experiments performed in early embryonic zebrafish suggested that collagens are important also during embryonic angiogenesis. In vitro assays revealed that collagen I allows for the most efficient endothelial cell migration, followed by collagen IV relative to the complete absence of exogenous matrix support. Our data demonstrates severe vascular defects and restricted fin regeneration when collagens are impaired. Collagen I therefore, provides support and guidance for endothelial cell migration while collagen IV is responsible for proper lumen formation and vascular integrity.

Ubiquitin ligase Wwp1 gene deletion attenuates diastolic dysfunction in pressure-overload hypertrophy.

Heart failure with a preserved left ventricular (LV) ejection fraction (HFpEF) often arises from a prolonged LV pressure overload (LVPO) and accompanied by abnormal extracellular matrix (ECM) accumulation. The E3 ubiquitin ligase WWP1 is a fundamental determinant ECM turnover. We tested the hypothesis that genetic ablation of Wwp1 would alter the progression of LVPO-induced HFpEF. LV echocardiography in mice with global Wwp1 deletion (n = 23; Wwp1-/-) was performed at 12 wk of age (baseline) and then at 2 and 4 wk following LVPO (transverse aortic banding) or surgery without LVPO induction. Age-matched wild-type mice (Wwp1+/+; n = 23) underwent identical protocols. LV EF remained constant and unchanged with LVPO and LV mass increased in both groups but was lower in the Wwp1-/- mice. With LVPO, the E/A ratio, an index of LV filling, was 3.97 ± 0.46 in Wwp1+/+ but was 1.73 ± 0.19 in the Wwp1-/- group (P < 0.05). At the transcriptional level, mRNA for fibrillar collagens (types I and III) decreased by approximately 50% in Wwp1-/- compared with the Wwp1+/+ group at 4 wk post-LVPO (P < 0.05) and was paralleled by a similar difference in LV fibrillar collagen content as measured by histochemistry. Moreover, mRNA levels for determinants favoring ECM accumulation, such as transforming growth factor (TGF), increased with LVPO, but were lower in the Wwp1-/- group. The absence of Wwp1 reduced the development of left ventricular hypertrophy and subsequent progression to HFpEF. Modulating the WWP1 pathway could be a therapeutic target to alter the natural history of HFpEF.NEW & NOTEWORTHY Heart failure with a preserved left ventricular (LV) ejection fraction (HFpEF) often arises from a prolonged LV pressure overload (LVPO) and is accompanied by abnormal extracellular matrix (ECM) accumulation. It is now recognized that the ECM is a dynamic entity that is regulated at multiple post-transcriptional levels, including the E3 ubiquitin ligases, such as WWP1. In the present study, WWP1 deletion in the context of an LVPO stimulus reduced functional indices of HFpEF progression and determinants of ECM remodeling.

Tendon and motor phenotypes in the Crtap-/- mouse model of recessive osteogenesis imperfecta.

Osteogenesis imperfecta (OI) is characterized by short stature, skeletal deformities, low bone mass, and motor deficits. A subset of OI patients also present with joint hypermobility; however, the role of tendon dysfunction in OI pathogenesis is largely unknown. Using the Crtap-/- mouse model of severe, recessive OI, we found that mutant Achilles and patellar tendons were thinner and weaker with increased collagen cross-links and reduced collagen fibril size at 1- and 4-months compared to wildtype. Patellar tendons from Crtap-/- mice also had altered numbers of CD146+CD200+ and CD146-CD200+ progenitor-like cells at skeletal maturity. RNA-seq analysis of Achilles and patellar tendons from 1-month Crtap-/- mice revealed dysregulation in matrix and tendon marker gene expression concomitant with predicted alterations in TGF-ß, inflammatory, and metabolic signaling. At 4-months, Crtap-/- mice showed increased αSMA, MMP2, and phospho-NFκB staining in the patellar tendon consistent with excess matrix remodeling and tissue inflammation. Finally, a series of behavioral tests showed severe motor impairments and reduced grip strength in 4-month Crtap-/- mice - a phenotype that correlates with the tendon pathology.

Suppression of pancreatic ductal adenocarcinoma growth and metastasis by fibrillar collagens produced selectively by tumor cells.

Pancreatic ductal adenocarcinoma (PDAC) has a collagen-rich dense extracellular matrix (ECM) that promotes malignancy of cancer cells and presents a barrier for drug delivery. Data analysis of our published mass spectrometry (MS)-based studies on enriched ECM from samples of progressive PDAC stages reveal that the C-terminal prodomains of fibrillar collagens are partially uncleaved in PDAC ECM, suggesting reduced procollagen C-proteinase activity. We further show that the enzyme responsible for procollagen C-proteinase activity, bone morphogenetic protein1 (BMP1), selectively suppresses tumor growth and metastasis in cells expressing high levels of COL1A1. Although BMP1, as a secreted proteinase, promotes fibrillar collagen deposition from both cancer cells and stromal cells, only cancer-cell-derived procollagen cleavage and deposition suppresses tumor malignancy. These studies reveal a role for cancer-cell-derived fibrillar collagen in selectively restraining tumor growth and suggest stratification of patients based on their tumor epithelial collagen I expression when considering treatments related to perturbation of fibrillar collagens.

Histopathology of recurrent Steel syndrome in fetuses caused by novel variants of COL27A1 gene.

Steel syndrome (STLS) encompasses characteristic facies, dwarfness, irreducible bilateral hip and radial head dislocation, and carpal bone coalition due to COL27A1 mutations. Two consecutive pregnancies in a non-consanguineous couple were terminated because of severe fetal anomalies. Complete autopsies with microscopic exam were performed on both fetuses. Next-generation-based clinical exome sequencing was applied to the first fetus. Exome sequencing results, parental segregation, and affection of the second fetus were confirmed by Sanger sequencing. Both fetuses had signs consistent with STLS. Bilateral capitulum humeri absence explained radial head dislocation in STLS. Metaphyseal cartilage showed severe disorganization. Resting cartilage was hypercellular, organized in irregular nests limited by acellular matrix. Two variants in COL27A1 (c.2548G>A -p.Gly850Arg- and c.3249+1G> T) were found in both fetuses in compound heterozygosity with parental Mendelian segregation. This is the first report to include histology of STLS. The COL27A1 variants here described increase the number of mutations associated with STLS.

Brothers with novel compound heterozygous mutations in COL27A1 causing dental and genital abnormalities.

COL27A1 encodes a collagen type XXVII alpha 1 chain. It is the product of this gene that provides the structural support of connective tissue and is reported to be the causative gene of Steel syndrome (OMIM #615155). The primary symptoms of patients with this defect are consistent with systemic bone disease; however, recent reports note findings of intellectual disability and hearing loss. In this study, we identified novel COL27A1 compound heterozygous variants in two brothers with rhizomelia and congenital hip dislocation as well as dental and genital abnormalities that have not yet been reported in Steel syndrome. This variant, of maternal origin, caused an amino acid substitution of arginine for glycine, c.2026G>C or p.G676R, in the collagen helix domain, which is assumed to damage the structure of the helix. The paternally transmitted variant, c.2367G>A, is located at the 3' end of exon 12, and cDNA analysis revealed a splicing alteration. These novel, compound heterozygous COL27A1 variants might indicate an association of the gene with tooth and genital abnormalities.

Genome-wide association of phenotypes based on clustering patterns of hand osteoarthritis identify WNT9A as novel osteoarthritis gene.

BACKGROUND: Despite recent advances in the understanding of the genetic architecture of osteoarthritis (OA), only two genetic loci have been identified for OA of the hand, in part explained by the complexity of the different hand joints and heterogeneity of OA pathology. METHODS: We used data from the Rotterdam Study (RSI, RSII and RSIII) to create three hand OA phenotypes based on clustering patterns of radiographic OA severity to increase power in our modest discovery genome-wide association studies in the RS (n=8700), and sought replication in an independent cohort, the Framingham Heart Study (n=1203). We used multiple approaches that leverage different levels of information and functional data to further investigate the underlying biological mechanisms and candidate genes for replicated loci. We also attempted to replicate known OA loci at other joint sites, including the hips and knees. RESULTS: We found two novel genome-wide significant loci for OA in the thumb joints. We identified WNT9A as a possible novel causal gene involved in OA pathogenesis. Furthermore, several previously identified genetic loci for OA seem to confer risk for OA across multiple joints: TGFa, RUNX2, COL27A1, ASTN2, IL11 and GDF5 loci. CONCLUSIONS: We identified a robust novel genetic locus for hand OA on chromosome 1, of which WNT9A is the most likely causal gene. In addition, multiple genetic loci were identified to be associated with OA across multiple joints. Our study confirms the potential for novel insight into the genetic architecture of OA by using biologically meaningful stratified phenotypes.

Collagen XII mediated cellular and extracellular mechanisms regulate establishment of tendon structure and function.

Tendons have a uniaxially aligned structure with a hierarchical organization of collagen fibrils crucial for tendon function. Collagen XII is expressed in tendons and has been implicated in the regulation of fibrillogenesis. It is a non-fibrillar collagen belonging to the Fibril-Associated Collagens with Interrupted Triple Helices (FACIT) family. Mutations in COL12A1 cause myopathic Ehlers Danlos Syndrome with a clinical phenotype involving both joints and tendons supporting critical role(s) for collagen XII in tendon development and function. Here we demonstrate the molecular function of collagen XII during tendon development using a Col12a1 null mouse model. Col12a1 deficiency altered tenocyte shape, formation of interacting cell processes, and organization resulting in impaired cell-cell communication and disruption of hierarchal structure as well as decreased tissue stiffness. Immuno-localization revealed that collagen XII accumulated on the tenocyte surface and connected adjacent tenocytes by building matrix bridges between the cells, suggesting that collagen XII regulates intercellular communication. In addition, there was a decrease in fibrillar collagen I in collagen XII deficient tenocyte cultures compared with controls suggesting collagen XII signaling specifically alters tenocyte biosynthesis. This suggests that collagen XII provides feedback to tenocytes regulating extracellular collagen I. Together, the data indicate dual roles for collagen XII in determination of tendon structure and function. Through association with fibrils it functions in fibril packing, fiber assembly and stability. In addition, collagen XII influences tenocyte organization required for assembly of higher order structure; intercellular communication necessary to coordinate long range order and feedback on tenocytes influencing collagen synthesis. Integration of both regulatory roles is required for the acquisition of hierarchal structure and mechanical properties.

The tricuspid valve also maladapts as shown in sheep with biventricular heart failure.

Over 1.6 million Americans suffer from significant tricuspid valve leakage. In most cases this leakage is designated as secondary. Thus, valve dysfunction is assumed to be due to valve-extrinsic factors. We challenge this paradigm and hypothesize that the tricuspid valve maladapts in those patients rendering the valve at least partially culpable for its dysfunction. As a first step in testing this hypothesis, we set out to demonstrate that the tricuspid valve maladapts in disease. To this end, we induced biventricular heart failure in sheep that developed tricuspid valve leakage. In the anterior leaflets of those animals, we investigated maladaptation on multiple scales. We demonstrated alterations on the protein and cell-level, leading to tissue growth, thickening, and stiffening. These data provide a new perspective on a poorly understood, yet highly prevalent disease. Our findings may motivate novel therapy options for many currently untreated patients with leaky tricuspid valves.

Collagen XI regulates the acquisition of collagen fibril structure, organization and functional properties in tendon.

Collagen XI is a fibril-forming collagen that regulates collagen fibrillogenesis. Collagen XI is normally associated with collagen II-containing tissues such as cartilage, but it also is expressed broadly during development in collagen I-containing tissues, including tendons. The goals of this study are to define the roles of collagen XI in regulation of tendon fibrillar structure and the relationship to function. A conditional Col11a1-null mouse model was created to permit the spatial and temporal manipulation of Col11a1 expression. We hypothesize that collagen XI functions to regulate fibril assembly, organization and, therefore, tendon function. Previous work using cho mice with ablated Col11a1 alleles supported roles for collagen XI in tendon fibril assembly. Homozygous cho/cho mice have a perinatal lethal phenotype that limited the studies. To circumvent this, a conditional Col11a1flox/flox mouse model was created where exon 3 was flanked with loxP sites. Breeding with Scleraxis-Cre (Scx-Cre) mice yielded a tendon-specific Col11a1-null mouse line, Col11a1Δten/Δten. Col11a1flox/flox mice had no phenotype compared to wild type C57BL/6 mice and other control mice, e.g., Col11a1flox/flox and Scx-Cre. Col11a1flox/flox mice expressed Col11a1 mRNA at levels comparable to wild type and Scx-Cre mice. In contrast, in Col11a1Δten/Δten mice, Col11a1 mRNA expression decreased to baseline in flexor digitorum longus tendons (FDL). Collagen XI protein expression was absent in Col11a1Δten/Δten FDLs, and at ~50% in Col11a1+/Δten compared to controls. Phenotypically, Col11a1Δten/Δten mice had significantly decreased body weights (p < 0.001), grip strengths (p < 0.001), and with age developed gait impairment becoming hypomobile. In the absence of Col11a1, the tendon collagen fibrillar matrix was abnormal when analyzed using transmission electron microscopy. Reducing Col11a1 and, therefore collagen XI content, resulted in abnormal fibril structure, loss of normal fibril diameter control with a significant shift to small diameters and disrupted parallel alignment of fibrils. These alterations in matrix structure were observed in developing (day 4), maturing (day 30) and mature (day 60) mice. Altering the time of knockdown using inducible I-Col11a1-/- mice indicated that the primary regulatory foci for collagen XI was in development. In mature Col11a1Δten/Δten FDLs a significant decrease in the biomechanical properties was observed. The decrease in maximum stress and modulus suggest that fundamental differences in the material properties in the absence of Col11a1 expression underlie the mechanical deficiencies. These data demonstrate an essential role for collagen XI in regulation of tendon fibril assembly and organization occurring primarily during development.

Prognostic significance of abnormal matrix collagen remodeling in colorectal cancer based on histologic and bioinformatics analysis.

As the major component of the tumor matrix, collagen greatly influences tumor invasion and prognosis. The present study compared the remodeling of collagen and collagenase in 56 patients with colorectal cancer (CRC) using Sirius red stain and immunohistochemistry, exploring the relationship between collagen remodeling and the prognosis of CRC. Weak or strong changes in collagen fiber arrangement in birefringence were observed. With the exception of a higher density, weak changes equated to a similar arrangement in normal collagen, while strong changes facilitated cross­linking into bundles. Compared with normal tissues, collagen I (COL I) and III (COL III) deposition was significantly increased in CRC tissues, and was positively correlated with the metastasis status. In tissues without distant metastasis, collagen IV (COL IV) levels were higher than that in normal tissues, while in tissues with distant metastasis, collagen IV expression was significantly lower. Furthermore, the expression of matrix metalloproteinase (MMP)­1, MMP­2, MMP­7, MMP­9 and lysyl oxidase­like 2 (LOXL2) was found to be elevated in the cancer stroma, which contributed to the hyperactive remodeling of collagen. The association between collagen­related genes and the occurrence and prognosis of CRC were analyzed using biometric databases. The results indicated that patients with upregulated expression of a combination of coding genes for collagen and collagenase exhibited poorer overall survival times. The coding genes COL1A1­2, COL3A1, COL4A3, COL4A6 and MMP2 may therefore be used as biomarkers to predict the prognosis of patients with CRC. Furthermore, the results of Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis suggest that collagen may promote tumor development by activating platelets. Collectively, the abnormal collagen remodeling, including associated protein and coding genes is associated with the tumorigenesis and metastasis, affecting the prognosis of patients with CRC.

Comparative Genomics Reveals Evolution of a Beak Morphology Locus in a High-Altitude Songbird.

The Ground Tit (Pseudopodoces humilis) has lived on the Qinghai-Tibet Plateau for ∼5.7 My and has the highest altitudinal distribution among all parids. This species has evolved an elongated beak in response to long-term selection imposed by ground-foraging and cavity-nesting habits, yet the genetic basis for beak elongation remains unknown. Here, we perform genome-wide analyses across 14 parid species and identify 25 highly divergent genomic regions that are significantly associated with beak length, finding seven candidate genes involved in bone morphogenesis and remolding. Neutrality tests indicate that a model allowing for a selective sweep in the highly conserved COL27A1 gene best explains variation in beak length. We also identify two nonsynonymous fixed mutations in the collagen domain that are predicted to be functionally deleterious yet may have facilitated beak elongation. Our study provides evidence of adaptive alleles in COL27A1 with major effects on beak elongation of Ps. humilis.

First reported case of Steel syndrome in the European population: A novel homozygous mutation in COL27A1 and review of the literature.

Steel syndrome is an autosomal recessive disorder that primarily affects the skeletal system causing a variety of manifestations. Sixteen individuals with Steel syndrome, mainly Puerto Ricans (11/16), were previously reported to carry bi-allelic mutations in the COL27A1 gene. Here, we present the first patient with Steel syndrome in Europe and the sixth non-Puerto Rican carrying a novel homozygous mutation in COL27A1. The patient is a 4-year-old boy born to non-consanguineous healthy parents, with dysmorphic facial features, absent hip ossification centres, external rotation of both feet, relatively short stature, mild skin syndactyly, short mid phalanges and bilateral sensorineural hearing loss. Whole exome sequencing (WES) revealed a novel homozygous missense variant p.(Gly802Glu) in COL27A1. The homozygous mutation was confirmed by Sanger sequencing in the proband and carrier status was confirmed in both parents and his unaffected sibling. According to online and in-house minor allele frequency (MAF) databases, this is the first COL27A1 mutation reported in the European population. Additional screening of healthy Greek-Cypriot individuals was thus performed, which did not reveal any additional carriers in the population for the variant in question.

Functional biology of the Steel syndrome founder allele and evidence for clan genomics derivation of COL27A1 pathogenic alleles worldwide.

Previously we reported the identification of a homozygous COL27A1 (c.2089G>C; p.Gly697Arg) missense variant and proposed it as a founder allele in Puerto Rico segregating with Steel syndrome (STLS, MIM #615155); a rare osteochondrodysplasia characterized by short stature, congenital bilateral hip dysplasia, carpal coalitions, and scoliosis. We now report segregation of this variant in five probands from the initial clinical report defining the syndrome and an additional family of Puerto Rican descent with multiple affected adult individuals. We modeled the orthologous variant in murine Col27a1 and found it recapitulates some of the major Steel syndrome associated skeletal features including reduced body length, scoliosis, and a more rounded skull shape. Characterization of the in vivo murine model shows abnormal collagen deposition in the extracellular matrix and disorganization of the proliferative zone of the growth plate. We report additional COL27A1 pathogenic variant alleles identified in unrelated consanguineous Turkish kindreds suggesting Clan Genomics and identity-by-descent homozygosity contributing to disease in this population. The hypothesis that carrier states for this autosomal recessive osteochondrodysplasia may contribute to common complex traits is further explored in a large clinical population cohort. Our findings augment our understanding of COL27A1 biology and its role in skeletal development; and expand the functional allelic architecture in this gene underlying both rare and common disease phenotypes.