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1.  Centriolar satellites assemble centrosomal microcephaly proteins to recruit CDK2 and promote centriole duplication 
eLife  null;4:e07519.
Primary microcephaly (MCPH) associated proteins CDK5RAP2, CEP152, WDR62 and CEP63 colocalize at the centrosome. We found that they interact to promote centriole duplication and form a hierarchy in which each is required to localize another to the centrosome, with CDK5RAP2 at the apex, and CEP152, WDR62 and CEP63 at sequentially lower positions. MCPH proteins interact with distinct centriolar satellite proteins; CDK5RAP2 interacts with SPAG5 and CEP72, CEP152 with CEP131, WDR62 with MOONRAKER, and CEP63 with CEP90 and CCDC14. These satellite proteins localize their cognate MCPH interactors to centrosomes and also promote centriole duplication. Consistent with a role for satellites in microcephaly, homozygous mutations in one satellite gene, CEP90, may cause MCPH. The satellite proteins, with the exception of CCDC14, and MCPH proteins promote centriole duplication by recruiting CDK2 to the centrosome. Thus, centriolar satellites build a MCPH complex critical for human neurodevelopment that promotes CDK2 centrosomal localization and centriole duplication.
eLife digest
When a cell divides, the chromosomes that contain the genetic blueprint for the cell must be replicated and shared between the two new cells. A structure called the centrosome organizes the cellular machinery that separates the chromosome copies during cell division. At the center of each centrosome are two cylindrical microtubule-based structures called centrioles.
Mutations in certain proteins that interact with the centrosome cause a neurodevelopmental disorder called primary microcephaly. People born with microcephaly have unusually small heads and brains. As a result, they may have difficulties with mental tasks. Scientists do not know exactly how these ‘microcephaly-associated’ proteins normally interact with the centrosomes or what they do at the centrosomes, so it is difficult to work out what goes wrong in people with microcephaly. One idea is that the proteins help to duplicate the centrioles before a cell divides. If this duplication does not occur, a cell cannot divide properly; so, people with mutations that interfere with centriole duplication cannot grow enough brain cells.
Now, Kodani et al. have examined how these microcephaly-associated proteins work with ‘satellite’ proteins that congregate near the centrosome to duplicate centrioles. The satellite proteins help to recruit four microcephaly-associated proteins to the centrosome, where they are built into a ring. The microcephaly-associated proteins congregate at the centrosome in a particular order, with each protein recruiting the next one in the sequence. Once all four are in place near the centrosome, an enzyme that helps to duplicate the centrioles joins them.
Further experiments suggest that mutations that affect one of the satellite proteins—known as CEP90—may cause microcephaly. Future analysis of how microcephaly-associated genes work may reveal the cell biological mechanisms by which centrioles participate in brain development.
PMCID: PMC4574112  PMID: 26297806
centriole; microcephaly; centrosome; centriolar satellite; cell cycle; brain development; human
2.  METTL23, a transcriptional partner of GABPA, is essential for human cognition 
Human Molecular Genetics  2014;23(13):3456-3466.
Whereas many genes associated with intellectual disability (ID) encode synaptic proteins, transcriptional defects leading to ID are less well understood. We studied a large, consanguineous pedigree of Arab origin with seven members affected with ID and mild dysmorphic features. Homozygosity mapping and linkage analysis identified a candidate region on chromosome 17 with a maximum multipoint logarithm of odds score of 6.01. Targeted high-throughput sequencing of the exons in the candidate region identified a homozygous 4-bp deletion (c.169_172delCACT) in the METTL23 (methyltransferase like 23) gene, which is predicted to result in a frameshift and premature truncation (p.His57Valfs*11). Overexpressed METTL23 protein localized to both nucleus and cytoplasm, and physically interacted with GABPA (GA-binding protein transcription factor, alpha subunit). GABP, of which GABPA is a component, is known to regulate the expression of genes such as THPO (thrombopoietin) and ATP5B (ATP synthase, H+ transporting, mitochondrial F1 complex, beta polypeptide) and is implicated in a wide variety of important cellular functions. Overexpression of METTL23 resulted in increased transcriptional activity at the THPO promoter, whereas knockdown of METTL23 with siRNA resulted in decreased expression of ATP5B, thus revealing the importance of METTL23 as a regulator of GABPA function. The METTL23 mutation highlights a new transcriptional pathway underlying human intellectual function.
PMCID: PMC4049305  PMID: 24501276
3.  TCTEX1D2 mutations underlie Jeune asphyxiating thoracic dystrophy with impaired retrograde intraflagellar transport 
Nature Communications  2015;6:7074.
The analysis of individuals with ciliary chondrodysplasias can shed light on sensitive mechanisms controlling ciliogenesis and cell signalling that are essential to embryonic development and survival. Here we identify TCTEX1D2 mutations causing Jeune asphyxiating thoracic dystrophy with partially penetrant inheritance. Loss of TCTEX1D2 impairs retrograde intraflagellar transport (IFT) in humans and the protist Chlamydomonas, accompanied by destabilization of the retrograde IFT dynein motor. We thus define TCTEX1D2 as an integral component of the evolutionarily conserved retrograde IFT machinery. In complex with several IFT dynein light chains, it is required for correct vertebrate skeletal formation but may be functionally redundant under certain conditions.
Severe congenital development defects such as Jeune syndrome can result from the malfunction of primary cilia and dynein. Here Schmidts et al. report unique biallelic null mutations in a gene encoding a dynein light chain, helping to explain the nature of ciliopathies in human patients.
PMCID: PMC4468853  PMID: 26044572
4.  A novel disorder reveals clathrin heavy chain-22 is essential for human pain and touch development 
Brain  2015;138(8):2147-2160.
Congenital inability to feel pain is rare, but the identification of causative genes is translating into the development of novel analgesics. Nahorski et al. describe insensitivity to pain caused by mutations affecting the second clathrin heavy chain (CHC22), and reveal a role for CHC22 in pain and touch development.
Congenital inability to feel pain is rare, but the identification of causative genes is translating into the development of novel analgesics. Nahorski et al. describe insensitivity to pain caused by mutations affecting the second clathrin heavy chain (CHC22), and reveal a role for CHC22 in pain and touch development.
Congenital inability to feel pain is very rare but the identification of causative genes has yielded significant insights into pain pathways and also novel targets for pain treatment. We report a novel recessive disorder characterized by congenital insensitivity to pain, inability to feel touch, and cognitive delay. Affected individuals harboured a homozygous missense mutation in CLTCL1 encoding the CHC22 clathrin heavy chain, p.E330K, which we demonstrate to have a functional effect on the protein. We found that CLTCL1 is significantly upregulated in the developing human brain, displaying an expression pattern suggestive of an early neurodevelopmental role. Guided by the disease phenotype, we investigated the role of CHC22 in two human neural crest differentiation systems; human induced pluripotent stem cell-derived nociceptors and TRKB-dependant SH-SY5Y cells. In both there was a significant downregulation of CHC22 upon the onset of neural differentiation. Furthermore, knockdown of CHC22 induced neurite outgrowth in neural precursor cells, which was rescued by stable overexpression of small interfering RNA-resistant CHC22, but not by mutant CHC22. Similarly, overexpression of wild-type, but not mutant, CHC22 blocked neurite outgrowth in cells treated with retinoic acid. These results reveal an essential and non-redundant role for CHC22 in neural crest development and in the genesis of pain and touch sensing neurons.
PMCID: PMC4511860  PMID: 26068709
insensitivity to pain; clathrin; endosomal trafficking; neurogenesis
5.  An Integrative Computational Approach for Prioritization of Genomic Variants 
PLoS ONE  2014;9(12):e114903.
An essential step in the discovery of molecular mechanisms contributing to disease phenotypes and efficient experimental planning is the development of weighted hypotheses that estimate the functional effects of sequence variants discovered by high-throughput genomics. With the increasing specialization of the bioinformatics resources, creating analytical workflows that seamlessly integrate data and bioinformatics tools developed by multiple groups becomes inevitable. Here we present a case study of a use of the distributed analytical environment integrating four complementary specialized resources, namely the Lynx platform, VISTA RViewer, the Developmental Brain Disorders Database (DBDB), and the RaptorX server, for the identification of high-confidence candidate genes contributing to pathogenesis of spina bifida. The analysis resulted in prediction and validation of deleterious mutations in the SLC19A placental transporter in mothers of the affected children that causes narrowing of the outlet channel and therefore leads to the reduced folate permeation rate. The described approach also enabled correct identification of several genes, previously shown to contribute to pathogenesis of spina bifida, and suggestion of additional genes for experimental validations. The study demonstrates that the seamless integration of bioinformatics resources enables fast and efficient prioritization and characterization of genomic factors and molecular networks contributing to the phenotypes of interest.
PMCID: PMC4266634  PMID: 25506935
6.  Genomic analysis of Meckel–Gruber syndrome in Arabs reveals marked genetic heterogeneity and novel candidate genes 
Meckel–Gruber syndrome (MKS, OMIM #249000) is a multiple congenital malformation syndrome that represents the severe end of the ciliopathy phenotypic spectrum. Despite the relatively common occurrence of this syndrome among Arabs, little is known about its genetic architecture in this population. This is a series of 18 Arab families with MKS, who were evaluated clinically and studied using autozygome-guided mutation analysis and exome sequencing. We show that autozygome-guided candidate gene analysis identified the underlying mutation in the majority (n=12, 71%). Exome sequencing revealed a likely pathogenic mutation in three novel candidate MKS disease genes. These include C5orf42, Ellis–van-Creveld disease gene EVC2 and SEC8 (also known as EXOC4), which encodes an exocyst protein with an established role in ciliogenesis. This is the largest and most comprehensive genomic study on MKS in Arabs and the results, in addition to revealing genetic and allelic heterogeneity, suggest that previously reported disease genes and the novel candidates uncovered by this study account for the overwhelming majority of MKS patients in our population.
PMCID: PMC3722952  PMID: 23169490
autozygome; ciliopathy; encephalocele; EVC2; EXOC4
7.  A novel mutation in DDR2 causing spondylo-meta-epiphyseal dysplasia with short limbs and abnormal calcifications (SMED-SL) results in defective intra-cellular trafficking 
BMC Medical Genetics  2014;15:42.
The rare autosomal genetic disorder, Spondylo-meta-epiphyseal dysplasia with short limbs and abnormal calcifications (SMED-SL), is reported to be caused by missense or splice site mutations in the human discoidin domain receptor 2 (DDR2) gene. Previously our group has established that trafficking defects and loss of ligand binding are the underlying cellular mechanisms of several SMED-SL causing mutations. Here we report the clinical characteristics of two siblings of consanguineous marriage with suspected SMED-SL and identification of a novel disease-causing mutation in the DDR2 gene.
Clinical evaluation and radiography were performed to evaluate the patients. All the coding exons and splice sites of the DDR2 gene were sequenced by Sanger sequencing. Subcellular localization of the mutated DDR2 protein was determined by confocal microscopy, deglycosylation assay and Western blotting. DDR2 activity was measured by collagen activation and Western analysis.
In addition to the typical features of SMED-SL, one of the patients has an eye phenotype including visual impairment due to optic atrophy. DNA sequencing revealed a novel homozygous dinucleotide deletion mutation (c.2468_2469delCT) on exon 18 of the DDR2 gene in both patients. The mutation resulted in a frameshift leading to an amino acid change at position S823 and a predicted premature termination of translation (p.S823Cfs*2). Subcellular localization of the mutant protein was analyzed in mammalian cell lines, and it was found to be largely retained in the endoplasmic reticulum (ER), which was further supported by its N-glycosylation profile. In keeping with its cellular mis-localization, the mutant protein was found to be deficient in collagen-induced receptor activation, suggesting protein trafficking defects as the major cellular mechanism underlying the loss of DDR2 function in our patients.
Our results indicate that the novel mutation results in defective trafficking of the DDR2 protein leading to loss of function and disease. This confirms our previous findings that DDR2 missense mutations occurring at the kinase domain result in retention of the mutant protein in the ER.
PMCID: PMC4001364  PMID: 24725993
DDR2; Spondylo-meta-epiphyseal dysplasia; Trafficking defect; SMED-SL; ERAD; Optic atrophy
8.  Delineation of the Clinical, Molecular and Cellular Aspects of Novel JAM3 Mutations Underlying the Autosomal Recessive Hemorrhagic Destruction of the Brain, Subependymal Calcification and Congenital Cataracts 
Human mutation  2013;34(3):498-505.
We have recently shown that the hemorrhagic destruction of the brain, subependymal calcification and congenital cataracts is caused by biallelic mutations in the gene encoding junctional adhesion molecule 3 (JAM3) protein. Affected members from three new families underwent detailed clinical examination including imaging of the brain. Affected individuals presented with a distinctive phenotype comprising hemorrhagic destruction of the brain, subependymal calcification and congenital cataracts. All patients had a catastrophic clinical course resulting in death in 7 out of 10 affected individuals. Sequencing the coding exons of JAM3 revealed three novel homozygous mutations: c.2T>G (p.M1R), c.346G>A (p.E116K) and c.656G>A (p.C219Y). The p.M1R mutation affects the start codon and therefore is predicted to impair protein synthesis. Cellular studies showed that the p.C219Y mutation resulted in a significant retention of the mutated protein in the endoplasmic reticulum, suggesting a trafficking defect. The p.E116K mutant traffics normally to the plasma membrane as the wild type and may have lost its function due to the lack of interaction with an interacting partner. Our data further support the importance of JAM3 in the development and function of the vascular system and the brain.
PMCID: PMC3951164  PMID: 23255084
brain; subependymal calcification; congenital cataract; JAM3
9.  LINS, a modulator of the WNT signaling pathway, is involved in human cognition 
Inherited intellectual disability (ID) conditions are a group of genetically heterogeneous disorders that lead to variable degrees of cognition deficits. It has been shown that inherited ID can be caused by mutations in over 100 different genes and there is evidence for the presence of as yet unidentified genes in a significant proportion of patients. We aimed at identifying the defective gene underlying an autosomal recessive ID in two sibs of an Emirati family.
A combined approach involving homozygosity mapping and whole-exome sequencing was used to identify the causative mutation. RNA analysis was performed to gain further insight into the pathogenic effect of the detected mutation.
We have identified a homozygous splicing mutation (c.1219_1222+1delAAAGG) in the LINS gene in the affected children. LINS is the human homologue of the Drosophila segment polarity gene lin that encodes an essential regulator of the wingless/Wnt signaling. The identified mutation alters the first consensus nucleotide of the 5' donor splice junction of intron 5 and the 3' end of exon 5. Transcript analysis revealed that this change leads to an exon skipping event resulting in direct splicing of exon 4 to exon 6. Another mutation in LINS has been described very briefly in an Iranian family with autosomal recessive ID and microcephaly.
Our study confirms that LINS, a modulator of the WNT pathway, is an indispensable gene to human cognition and this finding sheds further light on the importance of WNT signaling in human brain development and/or function.
PMCID: PMC3847167  PMID: 23773660
10.  Evolutionarily Assembled cis-Regulatory Module at a Human Ciliopathy Locus 
Science (New York, N.Y.)  2012;335(6071):966-969.
Neighboring genes are often coordinately expressed within cis-regulatory modules, but evidence that nonparalogous genes share functions in mammals is lacking. Here, we report that mutation of either TMEM138 or TMEM216 causes a phenotypically indistinguishable human ciliopathy, Joubert syndrome. Despite a lack of sequence homology, the genes are aligned in a head-to-tail configuration and joined by chromosomal rearrangement at the amphibian-to-reptile evolutionary transition. Expression of the two genes is mediated by a conserved regulatory element in the noncoding intergenic region. Coordinated expression is important for their interdependent cellular role in vesicular transport to primary cilia. Hence, during vertebrate evolution of genes involved in ciliogenesis, nonparalogous genes were arranged to a functional gene cluster with shared regulatory elements.
PMCID: PMC3671610  PMID: 22282472
11.  Identification of Mutations Underlying 20 Inborn Errors of Metabolism in the United Arab Emirates Population 
Inborn errors of metabolism (IEM) are frequently encountered by physicians in the United Arab Emirates (UAE). However, the mutations underlying a large number of these disorders have not yet been determined. Therefore, the objective of this study was to identify the mutations underlying a number of IEM disorders among UAE residents from both national and expatriate families. A case series of patients from 34 families attending the metabolic clinic at Tawam Hospital were clinically evaluated, and molecular testing was carried out to determine their causative mutations. The mutation analysis was carried out at molecular genetics diagnostic laboratories. Thirty-eight mutations have been identified as responsible for twenty IEM disorders, including in the metabolism of amino acids, lipids, steroids, metal transport and mitochondrial energy metabolism, and lysosomal storage disorders. Nine of the identified mutations are novel, including two missense mutations, three premature stop codons and four splice site mutations. Mutation analysis of IEM disorders in the UAE population has an important impact on molecular diagnosis and genetic counseling for families affected by these disorders.
PMCID: PMC3354585  PMID: 22106832
12.  In search of triallelism in Bardet–Biedl syndrome 
Bardet–Biedl syndrome (BBS) is a model disease for ciliopathy in humans. The remarkable genetic heterogeneity that characterizes this disease is consistent with accumulating data on the interaction between the proteins encoded by the 14 BBS genes identified to date. Previous reports suggested that such interaction may also extend to instances of oligogenic inheritance in the form of triallelism which defies the long held view of BBS as an autosomal recessive disease. In order to investigate the magnitude of triallelism in BBS, we conducted a comprehensive analysis of all 14 BBS genes as well as the CCDC28B-modifier gene in a cohort of 29 BBS families, most of which are multiplex. Two in trans mutations in a BBS gene were identified in each of these families for a total of 20 mutations including 12 that are novel. In no instance did we observe two mutations in unaffected members of a given family, or observe the presence of a third allele that convincingly acted as a modifier of penetrance and supported the triallelic model of BBS. In addition to presenting a comprehensive genotype/phenotype overview of a large set of BBS mutations, including the occurrence of nonsyndromic retinitis pigmentosa in a family with a novel BBS9 mutation, our study argues in favor of straightforward autosomal recessive BBS in most cases.
PMCID: PMC3306854  PMID: 22353939
epistasis; oligogenic; penetrance; modifiers
14.  A missense founder mutation in VLDLR is associated with Dysequilibrium Syndrome without quadrupedal locomotion 
BMC Medical Genetics  2012;13:80.
Dysequilibrium syndrome is a genetically heterogeneous condition that combines autosomal recessive, nonprogressive cerebellar ataxia with mental retardation. The condition has been classified into cerebellar ataxia, mental retardation and disequilibrium syndrome types 1 (CAMRQ1), 2 (CAMRQ2) and 3 (CAMRQ3) and attributed to mutations in VLDLR, CA8 and WDR81 genes, respectively. Quadrupedal locomotion in this syndrome has been reported in association with mutations in all three genes.
SNP mapping and candidate gene sequencing in one consanguineous Omani family from the United Arab Emirates with cerebellar hypoplasia, moderate mental retardation, delayed ambulation and truncal ataxia was used to identify the mutation. In a second unrelated consanguineous Omani family, massively parallel exonic sequencing was used.
We identified a homozygous missense mutation (c.2117 G > T, p.C706F) in the VLDLR gene in both families on a shared affected haplotype block.This is the first reported homozygous missense mutation in VLDLR and it occurs in a highly conserved residue and predicted to be damaging to protein function.
We have delineated the phenotype associated with dysequilibrium syndrome in two Omani families and identified the first homozygous missense pathogenic mutation in VLDLR gene with likely founder effect in the southeastern part of the Arabian Peninsula.
PMCID: PMC3495048  PMID: 22973972
15.  A mutation in KIF7 is responsible for the autosomal recessive syndrome of macrocephaly, multiple epiphyseal dysplasia and distinctive facial appearance 
We previously reported the existence of a unique autosomal recessive syndrome consisting of macrocephaly, multiple epiphyseal dysplasia and distinctive facial appearance mapping to chromosome 15q26.
In this manuscript, we have used whole exome sequencing on two affected members of a consanguineous family with this condition and carried out detailed bioinformatics analysis to elucidate the causative mutation.
Our analysis resulted in the identification of a homozygous p.N1060S missense mutation in a highly conserved residue in KIF7, a regulator of Hedgehog signaling that has been recently found to be causing Joubert syndrome, fetal hydrolethalus and acrocallosal syndromes. The phenotype in our patients partially overlaps with the phenotypes associated with those syndromes but they also exhibit some distinctive features including multiple epiphyseal dysplasia.
We report the first missense homozygous disease-causing mutation in KIF7 and expand the clinical spectrum associated with mutations in this gene to include multiple epiphyseal dysplasia. The missense nature of the mutation might account for the unique presentation in our patients.
PMCID: PMC3492204  PMID: 22587682
KIF7; Acrocallosal; Joubert; Sonic hedgehog; Dysmorphism; Multiple epiphyseal dysplasia; Fetal hydrolethalus
16.  Endoplasmic Reticulum Quality Control Is Involved in the Mechanism of Endoglin-Mediated Hereditary Haemorrhagic Telangiectasia 
PLoS ONE  2011;6(10):e26206.
Hereditary haemorrhagic telangiectasia (HHT) is an autosomal dominant genetic condition affecting the vascular system and is characterised by epistaxis, arteriovenous malformations and mucocutaneous and gastrointestinal telangiectases. This disorder affects approximately 1 in 8,000 people worldwide. Significant morbidity is associated with this condition in affected individuals, and anaemia can be a consequence of repeated haemorrhages from telangiectasia in the gut and nose. In the majority of the cases reported, the condition is caused by mutations in either ACVRL1 or endoglin genes, which encode components of the TGF-beta signalling pathway. Numerous missense mutations in endoglin have been reported as causative defects for HHT but the exact underlying cellular mechanisms caused by these mutations have not been fully established despite data supporting a role for the endoplasmic reticulum (ER) quality control machinery. For this reason, we examined the subcellular trafficking of twenty-five endoglin disease-causing missense mutations. The mutant proteins were expressed in HeLa and HEK293 cell lines, and their subcellular localizations were established by confocal fluorescence microscopy alongside the analysis of their N-glycosylation profiles. ER quality control was found to be responsible in eight (L32R, V49F, C53R, V125D, A160D, P165L, I271N and A308D) out of eleven mutants located on the orphan extracellular domain in addition to two (C363Y and C382W) out of thirteen mutants in the Zona Pellucida (ZP) domain. In addition, a single intracellular domain missense mutant was examined and found to traffic predominantly to the plasma membrane. These findings support the notion of the involvement of the ER's quality control in the mechanism of a significant number, but not all, missense endoglin mutants found in HHT type 1 patients. Other mechanisms including loss of interactions with signalling partners as well as adverse effects on functional residues are likely to be the cause of the mutant proteins' loss of function.
PMCID: PMC3194820  PMID: 22022569
17.  Loss of the BMP Antagonist, SMOC-1, Causes Ophthalmo-Acromelic (Waardenburg Anophthalmia) Syndrome in Humans and Mice 
PLoS Genetics  2011;7(7):e1002114.
Ophthalmo-acromelic syndrome (OAS), also known as Waardenburg Anophthalmia syndrome, is defined by the combination of eye malformations, most commonly bilateral anophthalmia, with post-axial oligosyndactyly. Homozygosity mapping and subsequent targeted mutation analysis of a locus on 14q24.2 identified homozygous mutations in SMOC1 (SPARC-related modular calcium binding 1) in eight unrelated families. Four of these mutations are nonsense, two frame-shift, and two missense. The missense mutations are both in the second Thyroglobulin Type-1 (Tg1) domain of the protein. The orthologous gene in the mouse, Smoc1, shows site- and stage-specific expression during eye, limb, craniofacial, and somite development. We also report a targeted pre-conditional gene-trap mutation of Smoc1 (Smoc1tm1a) that reduces mRNA to ∼10% of wild-type levels. This gene-trap results in highly penetrant hindlimb post-axial oligosyndactyly in homozygous mutant animals (Smoc1tm1a/tm1a). Eye malformations, most commonly coloboma, and cleft palate occur in a significant proportion of Smoc1tm1a/tm1a embryos and pups. Thus partial loss of Smoc-1 results in a convincing phenocopy of the human disease. SMOC-1 is one of the two mammalian paralogs of Drosophila Pentagone, an inhibitor of decapentaplegic. The orthologous gene in Xenopus laevis, Smoc-1, also functions as a Bone Morphogenic Protein (BMP) antagonist in early embryogenesis. Loss of BMP antagonism during mammalian development provides a plausible explanation for both the limb and eye phenotype in humans and mice.
Author Summary
Ophthalmo-acromelic syndrome (OAS) is a rare congenital genetic disorder involving complete absence of the eyes and limb malformations, with missing or fused bones in the feet and hands. In this paper we report the identification of genetic changes to both copies of the SMOC1 gene as the cause of most cases of OAS. We have identified eight different mutations in this gene in unrelated individuals, and six of these mutations are predicted to completely abolish SMOC-1 function. We have also genetically disrupted the mouse Smoc1 gene to produce only 10% of normal levels. These animals, called Smoc1tm1a/tm1a mice, have similar hindlimb malformations to those seen in the limbs of human OAS patients, resulting in missing toes in some mice and fusion of toes in others. Smoc1tm1a/tm1a embryos and pups also have eye malformations but these are milder than those seen in human cases, perhaps because, unlike the human cases, the mice still have some residual function of the gene. We suggest that the normal function of SMOC-1 may be to regulate an important class of growth factors, called Bone Morphogenetic Proteins (BMPs), which are essential for normal embryonic development.
PMCID: PMC3131273  PMID: 21750680
18.  Mutations in the Human Laminin β2 (LAMB2) Gene and the Associated Phenotypic Spectrum 
Human mutation  2010;31(9):992-1002.
Mutations of LAMB2 typically cause autosomal recessive Pierson syndrome, a disorder characterized by congenital nephrotic syndrome, ocular and neurologic abnormalities, but may occasionally be associated with milder or oligosymptomatic disease variants. LAMB2 encodes the basement membrane protein laminin β2 which is incorporated in specific heterotrimeric laminin isoforms and has an expression pattern corresponding to the pattern of organ manifestations in Pierson syndrome. Herein we review all previously reported and several novel LAMB2 mutations in relation to the associated phenotype in patients from 39 unrelated families. The majority of disease-causing LAMB2 mutations are truncating, consistent with the hypothesis that loss of laminin β2 function is the molecular basis of Pierson syndrome. While truncating mutations are distributed across the entire gene, missense mutations are clearly clustered in the N-terminal LN domain, which is important for intermolecular interactions. There is an association of missense mutations and small in frame deletions with a higher mean age at onset of renal disease and with absence of neurologic abnormalities, thus suggesting that at least some of these may represent hypomorphic alleles. Nevertheless, genotype alone does not appear to explain the full range of clinical variability, and therefore hitherto unidentified modifiers are likely to exist.
PMCID: PMC2978072  PMID: 20556798
LAMB2; Pierson syndrome; nephrotic syndrome; autosomal recessive; podocyte; laminin; ocular malformation
19.  SRD5A3 is required for the conversion of polyprenol to dolichol, essential for N-linked protein glycosylation 
Cell  2010;142(2):203-217.
N-linked glycosylation is the most frequent modification of secreted and membrane-bound proteins in eukaryotic cells, disruption of which is the basis of the Congenital Disorders of Glycosylation (CDG). We describe a new type of CDG caused by mutations in the steroid 5α-reductase type 3 (SRD5A3) gene. Patients have mental retardation, ophthalmologic and cerebellar defects. We found that SRD5A3 is necessary for the reduction of the alpha-isoprene unit of polyprenols to form dolichols, required for synthesis of dolichol-linked monosaccharides and the oligosaccharide precursor used for N-glycosylation. The presence of residual dolichol in cells depleted for this enzyme suggests the existence of an unexpected alternative pathway for dolichol de novo biosynthesis. Our results thus suggest that SRD5A3 is likely to be the long-sought polyprenol reductase and reveal the genetic basis of one of the earliest steps in protein N-linked glycosylation.
PMCID: PMC2940322  PMID: 20637498
N-glycosylation; dolichol; polyprenol; SRD5A3
20.  A novel NGF mutation clarifies the molecular mechanism and extends the phenotypic spectrum of the HSAN5 neuropathy 
Journal of Medical Genetics  2010;48(2):131-135.
Nerve growth factor β (NGFβ) and tyrosine kinase receptor type A (TRKA) are a well studied neurotrophin/receptor duo involved in neuronal survival and differentiation. The only previously reported hereditary sensory neuropathy caused by an NGF mutation, c.661C>T (HSAN5), and the pathology caused by biallelic mutations in the TRKA gene (NTRK1) (HSAN4), share only some clinical features. A consanguineous Arab family, where five of the six children were completely unable to perceive pain, were mentally retarded, did not sweat, could not discriminate temperature, and had a chronic immunodeficiency, is reported here. The condition is linked to a new homozygous mutation in the NGF gene, c.[680C>A]+[681_682delGG].
Genetic linkage and standard sequencing techniques were used to identify the causative gene. Using wild-type or mutant over-expression constructs transfected into PC12 and COS-7 cells, the cellular and molecular consequences of the mutations were investigated.
The mutant gene produced a precursor protein V232fs that was unable to differentiate PC12 cells. V232fs was not secreted from cells as mature NGFβ.
Both the clinical and cellular data suggest that the c.[680C>A]+[681_682delGG] NGF mutation is a functional null. The HSAN5 phenotype is extended to encompass HSAN4-like characteristics. It is concluded that the HSAN4 and HSAN5 phenotypes are parts of a phenotypic spectrum caused by changes in the NGF/TRKA signalling pathway.
PMCID: PMC3030776  PMID: 20978020
Clinical genetics; peripheral nerve disease
21.  Mutations in LRP2, which encodes the multiligand receptor megalin, cause Donnai-Barrow and facio-oculo-acoustico-renal syndromes 
Nature genetics  2007;39(8):957-959.
Donnai-Barrow syndrome is associated with agenesis of the corpus callosum, congenital diaphragmatic hernia, facial dysmorphology, ocular anomalies, sensorineural hearing loss and developmental delay. By studying multiplex families, we mapped this disorder to chromosome 2q23.3–31.1 and identified LRP2 mutations in six families with Donnai-Barrow syndrome and one family with facio-oculo-acoustico-renal syndrome. LRP2 encodes megalin, a multiligand uptake receptor that regulates levels of diverse circulating compounds. This work implicates a pathway with potential pharmacological therapeutic targets.
PMCID: PMC2891728  PMID: 17632512
22.  Community genetics. Its definition 2010 
Journal of Community Genetics  2010;1(1):19-22.
This paper presents a definition of the medical field of community genetics. It starts with a brief historical overview, defines the requirements for an adequate definition, presents the definition, and discusses the constituent parts of the definition.
PMCID: PMC3063846  PMID: 21475671
23.  Community genetics. Its definition 2010 
Journal of Community Genetics  2010;1(1):19-22.
This paper presents a definition of the medical field of community genetics. It starts with a brief historical overview, defines the requirements for an adequate definition, presents the definition, and discusses the constituent parts of the definition.
PMCID: PMC3063846  PMID: 21475671
24.  Trafficking defects and loss of ligand binding are the underlying causes of all reported DDR2 missense mutations found in SMED-SL patients 
Human Molecular Genetics  2010;19(11):2239-2250.
Spondylo-meta-epiphyseal dysplasia (SMED) with short limbs and abnormal calcifications (SMED-SL) is a rare, autosomal recessive human growth disorder, characterized by disproportionate short stature, short limbs, short broad fingers, abnormal metaphyses and epiphyses, platyspondyly and premature calcifications. Recently, three missense mutations and one splice-site mutation in the DDR2 gene were identified as causative genetic defects for SMED-SL, but the underlying cellular and biochemical mechanisms were not explored. Here we report a novel DDR2 missense mutation, c.337G>A (p.E113K), that causes SMED-SL in two siblings in the United Arab Emirates. Another DDR2 missense mutation, c.2254C>T (p.R752C), matching one of the previously reported SMED-SL mutations, was found in a second affected family. DDR2 is a plasma membrane receptor tyrosine kinase that functions as a collagen receptor. We expressed DDR2 constructs with the identified point mutations in human cell lines and evaluated their localization and functional properties. We found that all SMED-SL missense mutants were defective in collagen-induced receptor activation and that the three previously reported mutants (p.T713I, p.I726R and p.R752C) were retained in the endoplasmic reticulum. The novel mutant (p.E113K), in contrast, trafficked normally, like wild-type DDR2, but failed to bind collagen. This finding is in agreement with our recent structural data identifying Glu113 as an important amino acid in the DDR2 ligand-binding site. Our data thus demonstrate that SMED-SL can result from at least two different loss-of-function mechanisms: namely defects in DDR2 targeting to the plasma membrane or the loss of its ligand-binding activity.
PMCID: PMC2865377  PMID: 20223752
25.  Mutations in the inositol polyphosphate-5-phosphatase E gene link phosphatidyl inositol signaling to the ciliopathies 
Nature genetics  2009;41(9):1032-1036.
Phosphotidylinositol (PtdIns) signaling is tightly regulated, both spatially and temporally, by subcellularly localized PtdIns kinases and phosphatases that dynamically alter downstream signaling events 1. Joubert Syndrome (JS) characterized by a specific midbrain-hindbrain malformation (“molar tooth sign”) and variably associated retinal dystrophy, nephronophthisis, liver fibrosis and polydactyly 2, and is included in the newly emerging group of “ciliopathies”. In patients linking to JBTS1, we identified mutations in the INPP5E gene, encoding inositol polyphosphate-5-phosphatase E, which hydrolyzes the 5-phosphate of PtdIns(3,4,5)P3 and PtdIns(4,5)P2. Mutations clustered in the phosphatase domain and impaired 5-phosphatase activity, resulting in altered cellular PtdIns ratios. INPP5E localized to cilia in major organs affected in JS, and mutations promoted premature destabilization of cilia in response to stimulation. Thus, these data links PtdIns signaling to the primary cilium, a cellular structure that is becoming increasingly appreciated for its role in mediating cell signals and neuronal function.
PMCID: PMC2746682  PMID: 19668216

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