The deleted in brachydactyly B domain of ROR2 is required for receptor activation by recruitment of Src

The transmembrane receptor 'ROR2' resembles members of the receptor tyrosine kinase family of signalling receptors in sequence but its' signal transduction mechanisms remain enigmatic. This problem has particular importance because mutations in ROR2 are associated with two human skeletal dysmorphology syndromes, recessive Robinow Syndrome (RS) and dominant acting Brachydactyly type B (BDB). Here we show, using a constitutive dimerisation approach, that ROR2 exhibits dimerisation-induced tyrosine kinase activity and the ROR2 C-terminal domain, which is deleted in BDB, is required for recruitment and activation of the non-receptor tyrosine kinase Src. Native ROR2 phosphorylation is induced by the ligand Wnt5a and is blocked by pharmacological inhibition of Src kinase activity. Eight sites of Src-mediated ROR2 phosphorylation have been identified by mass spectrometry. Activation via tyrosine phosphorylation of ROR2 receptor leads to its internalisation into Rab5 positive endosomes. These findings show that BDB mutant receptors are defective in kinase activation as a result of failure to recruit Src

Isolated brachydactyly type E caused by a HOXD13 nonsense mutation: a case report

<p>Abstract</p> <p>Background</p> <p>Brachydactyly type E (BDE; MIM#113300) is characterized by shortening of the metacarpal, metatarsal, and often phalangeal bones, and predominantly affects postaxial ray(s) of the limb. BDE may occur as an isolated trait or as part of a syndrome. Isolated BDE is rare and in the majority of cases the molecular pathogenesis has so far not been resolved. Originally, the molecular cause of isolated BDE has been unravelled in 2 families and shown to result from heterozygous missense mutations in the homeodomain of the <it>HOXD13 </it>gene. Since the initial manuscript, one further <it>HOXD13 </it>mutation has been reported only in a single family manifesting isolated BDE.</p> <p>Case Presentation</p> <p>In this paper, we report on a Polish family exhibiting isolated BDE caused by a novel nonsense heterozygous <it>HOXD13 </it>mutation. We investigated a Polish female proband and her father, both affected by isolated BDE, in whom we identified a nonsense heterozygous mutation c.820C > T(p.R274X) in the <it>HOXD13 </it>gene. So far, only two missense <it>HOXD13 </it>substitutions (p.S308C and p.I314L), localized within the homeodomain of the HOXD13 transcription factor, as well as a single nonsense mutation (p.E181X) were associated with BDE. Both missense changes were supposed to alter DNA binding affinity of the protein.</p> <p>Conclusion</p> <p>The variant p.R274X identified in our proband is the fourth <it>HOXD13 </it>mutation, and the second truncating (nonsense) mutation, reported to result in typical isolated BDE. We refer our clinical and molecular findings to the previously described <it>HOXD13 </it>associated phenotypes and mutations.</p

Congenital syndactyly in cattle: four novel mutations in the low density lipoprotein receptor-related protein 4 gene (LRP4)

BACKGROUND: Isolated syndactyly in cattle, also known as mulefoot, is inherited as an autosomal recessive trait with variable penetrance in different cattle breeds. Recently, two independent mutations in the bovine LRP4 gene have been reported as the primary cause of syndactyly in the Holstein and Angus cattle breeds. RESULTS: We confirmed the previously described LRP4 exon 33 two nucleotide substitution in most of the affected Holstein calves and revealed additional evidence for allelic heterogeneity by the identification of four new LRP4 non-synonymous point mutations co-segregating in Holstein, German Simmental and Simmental-Charolais families. CONCLUSION: We confirmed a significant role of LRP4 mutations in the pathogenesis of congenital syndactyly in cattle. The newly detected missense mutations in the LRP4 gene represent independent mutations affecting different conserved protein domains. However, the four newly described LRP4 mutations do still not explain all analyzed cases of syndactyly

Functional Analysis of Alleged NOGGIN Mutation G92E Disproves Its Pathogenic Relevance

We identified an amino acid change (p.G92E) in the Bone Morphogenetic Protein antagonist NOGGIN in a 22-month-old boy who presented with a unilateral brachydactyly type B phenotype. Brachydactyly type B is a skeletal malformation that has been associated with increased Bone Morphogenetic Protein pathway activation in other patients. Previously, the amino acid change p.G92E in NOGGIN was described as causing fibrodysplasia ossificans progressiva, a rare genetic disorder characterized by limb malformations and progressive heterotopic bone formation in soft tissues that, like Brachydactyly type B, is caused by increased activation of Bone Morphogenetic Protein signaling. To determine whether G92E-NOGGIN shows impaired antagonism that could lead to increased Bone Morphogenetic Protein signaling, we performed functional assays to evaluate inhibition of BMP signaling. Interestingly, wt-NOGGIN shows different inhibition efficacies towards various Bone Morphogenetic Proteins that are known to be essential in limb development. However, comparing the biological activity of G92E-NOGGIN with wt-NOGGIN, we observed that G92E-NOGGIN inhibits activation of bone morphogenetic protein signaling with equal efficiency as wt-NOGGIN, supporting that G92E-NOGGIN does not cause pathological effects. Genetic testing of the child's parents revealed the same amino acid change in the healthy father, further supporting that p.G92E is a neutral amino acid substitution in NOGGIN. We conclude that p.G92E represents a rare polymorphism of the NOGGIN gene - causing neither brachydactyly nor fibrodysplasia ossificans progressiva. This study highlights that a given genetic variation should not be considered pathogenic unless supported by functional analyses

Brachydactyly

Brachydactyly ("short digits") is a general term that refers to disproportionately short fingers and toes, and forms part of the group of limb malformations characterized by bone dysostosis. The various types of isolated brachydactyly are rare, except for types A3 and D. Brachydactyly can occur either as an isolated malformation or as a part of a complex malformation syndrome. To date, many different forms of brachydactyly have been identified. Some forms also result in short stature. In isolated brachydactyly, subtle changes elsewhere may be present. Brachydactyly may also be accompanied by other hand malformations, such as syndactyly, polydactyly, reduction defects, or symphalangism

Ciliary Beating Recovery in Deficient Human Airway Epithelial Cells after Lentivirus Ex Vivo Gene Therapy

Primary Ciliary Dyskinesia is a heterogeneous genetic disease that is characterized by cilia dysfunction of the epithelial cells lining the respiratory tracts, resulting in recurrent respiratory tract infections. Despite lifelong physiological therapy and antibiotics, the lungs of affected patients are progressively destroyed, leading to respiratory insufficiency. Recessive mutations in Dynein Axonemal Intermediate chain type 1 (DNAI1) gene have been described in 10% of cases of Primary Ciliary Dyskinesia. Our goal was to restore normal ciliary beating in DNAI1–deficient human airway epithelial cells. A lentiviral vector based on Simian Immunodeficiency Virus pseudotyped with Vesicular Stomatitis Virus Glycoprotein was used to transduce cultured human airway epithelial cells with a cDNA of DNAI1 driven by the Elongation Factor 1 promoter. Transcription and translation of the transduced gene were tested by RT–PCR and western blot, respectively. Human airway epithelial cells that were DNAI1–deficient due to compound heterozygous mutations, and consequently had immotile cilia and no outer dynein arm, were transduced by the lentivirus. Cilia beating was recorded and electron microscopy of the cilia was performed. Transcription and translation of the transduced DNAI1 gene were detected in human cells treated with the lentivirus. In addition, immotile cilia recovered a normal beat and outer dynein arms reappeared. We demonstrated that it is possible to obtain a normalization of ciliary beat frequency of deficient human airway epithelial cells by using a lentivirus to transduce cells with the therapeutic gene. This preliminary step constitutes a conceptual proof that is indispensable in the perspective of Primary Ciliary Dyskinesia's in vivo gene therapy. This is the first time that recovery of cilia beating is demonstrated in this disease

Stat3 Activates the Receptor Tyrosine Kinase Like Orphan Receptor-1 Gene in Chronic Lymphocytic Leukemia Cells

BACKGROUND: The receptor tyrosine kinase like orphan receptor (ROR)-1 gene is overexpressed in chronic lymphocytic leukemia (CLL). Because Stat3 is constitutively activated in CLL and sequence analysis revealed that the ROR1 promoter harbors gamma-interferon activation sequence-like elements typically activated by Stat3, we hypothesized that Stat3 activates ROR1. METHODOLOGY/PRINCIPAL FINDINGS: Because IL-6 induced Stat3 phosphorylation and upregulated Ror1 protein levels in MM1 cells, we used these cells as a model. We transfected MM1 cells with truncated ROR1 promoter luciferase reporter constructs and found that IL-6 induced luciferase activity of ROR1-195 and upstream constructs. Co-transfection with Stat3 siRNA reduced the IL-6-induced luciferase activity, suggesting that IL-6 induced luciferase activity by activating Stat3. EMSA and the ChIP assay confirmed that Stat3 binds ROR1, and EMSA studies identified two Stat3 binding sites. In CLL cells, EMSA and ChIP studies determined that phosphorylated Stat3 bound to the ROR1 promoter at those two ROR1 promoter sites, and ChIP analysis showed that Stat3 co-immunoprecipitated DNA of STAT3, ROR1, and several Stat3-regulated genes. Finally, like STAT3-siRNA in MM1 cells, STAT3-shRNA downregulated STAT3, ROR1, and STAT3-regulated genes and Stat3 and Ror1 protein levels in CLL cells. CONCLUSION/SIGNIFICANCE: Our data suggest that constitutively activated Stat3 binds to the ROR1 promoter and activates ROR1 in CLL cells

Goal-directed and habitual control in the basal ganglia: implications for Parkinson's disease

Progressive loss of the ascending dopaminergic projection in the basal ganglia is a fundamental pathological feature of Parkinson's disease. Studies in animals and humans have identified spatially segregated functional territories in the basal ganglia for the control of goal-directed and habitual actions. In patients with Parkinson's disease the loss of dopamine is predominantly in the posterior putamen, a region of the basal ganglia associated with the control of habitual behaviour. These patients may therefore be forced into a progressive reliance on the goal-directed mode of action control that is mediated by comparatively preserved processing in the rostromedial striatum. Thus, many of their behavioural difficulties may reflect a loss of normal automatic control owing to distorting output signals from habitual control circuits, which impede the expression of goal-directed action. © 2010 Macmillan Publishers Limited. All rights reserved

The Epidemiology, Genetics and Future Management of Syndactyly

Syndactyly is a condition well documented in current literature due to it being the most common congenital hand defect, with a large aesthetic and functional significance