A novel splice site mutation in gene C2orf37 underlying Woodhouse–Sakati syndrome (WSS) in a consanguineous family of Pakistani origin

Gene 490 (2011) 26–31

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A novel splice site mutation in gene C2orf37 underlying Woodhouse–Sakatisyndrome (WSS) in a consanguineous family of Pakistani origin

Rabia Habib, Sulman Basit, Saadullah Khan, Muhammad Nasim Khan, Wasim Ahmad ⁎Department of Biochemistry, Faculty of Biological Sciences, Quaid-i-Azam University, Islamabad, Pakistan

Abbreviations: ALX4, aristaless like 4; ANE, alopeendocrinopathy; APMR, alopecia with mental retardationsearch tool; CT, computed tomography; C2orf37, chromosDNA, deoxyribonucleic acid; ECG, electrocardiogram; EPS,institutional review board; LOD, logarithm of odds; MRMIM, Mendelian inheritance in man; PCR, polymerase chaCalifornia, Santa Cruz; WSS, Woodhouse–Sakati syndrome.⁎ Corresponding author. Tel.: +92 51 90643003; fax:

E-mail address: [email protected] (W. Ahmad).

0378-1119/$ – see front matter © 2011 Elsevier B.V. Aldoi:10.1016/j.gene.2011.09.002

a b s t r a c t

Article history:Accepted 8 September 2011Available online 22 September 2011

Received by Takashi Gojobori

Keywords:Woodhouse–Sakati syndromeC2orf37Splice site mutation

Woodhouse–Sakati Syndrome (WSS) is a rare autosomal recessive multisystemic disorder that is marked byhypogonadism, alopecia, intellectual disability, deafness, diabetes mellitus and progressive extrapyramidaldefects. Mutations in the gene C2orf37 are the cause of Woodhouse–Sakati syndrome. In the present study,a four-generation consanguineous family with clinical manifestations of WSS was ascertained from a remoteregion of Pakistan. Linkage in the family was tested using microsatellite markers linked to several genesinvolved in producing WSS related phenotypes. Linkage in the family was established to the gene C2orf37,mapped on chromosome 2q22.3–2q35. DNA sequence analysis revealed a novel splice site mutationinvolving a homozygous G→A transition in the splice donor site of intron 3 (c.321+1 GNA) of C2orf37.This study presents a first report of Woodhouse–Sakati syndrome identified in Pakistani population.

cia, neurological defects and; BLAST, basic local alignmentome 2 open reading frame 37;extrapyramidal symptoms; IRB,I, magnetic resonance imaging;in reaction; UCSC, University of

+92 51 9205753.

l rights reserved.

© 2011 Elsevier B.V. All rights reserved.

1. Introduction

Woodhouse–Sakati syndrome (WSS; Mendelian inheritance in man241080) is an autosomal recessive multisystemic disorder character-ized by alopecia, hypogonadism, intellectual disability, diabetes melli-tus, extrapyramidal symptoms (EPS), deafness and decreased insulin-like growth factor 1 (IGF-1) levels (Al-Semari and Bohlega, 2007;Woodhouse and Sakati, 1983). In fewpatients withWSS, seizures, poly-neuropathy,whitematter lesions in the brain, thyroid dysfunction, elec-trocardiogram (ECG) abnormalities, keratoconus and syndactyly ofhand and/or feet have been reported as well (Al-Swailem et al., 2006;Schneider and Bhatia, 2008).

Alazami et al. (2008)mappedWSS locus, in a family originated fromSaudi Arabia, on chromosomes 2q22.3–2q35 and identified mutationsin the gene C2orf37 responsible for generating WSS phenotypes. Todate, eight disease causing mutations including nonsense, deletionsand splicing have been reported in the gene C2orf37. A deletion muta-tion (c.436delC), representing a founder mutation, was identified in af-fected individuals of eight Saudi Arabian families (Alazami et al., 2010).

The C2orf37 shows high level expression in brain, liver and skintissues that is consistent with the disease phenotype observed in

these organs (Alazami et al., 2008). The number of organs affectedclinically and the progressive nature of their involvement is indicativeof the wide role that C2orf37 plays in the development and/or main-tenance of these organs.

In the study, presented here, we report the first Pakistani familywith clinical manifestations of WSS. DNA sequence analysis revealeda novel splice site mutation (c.321+1 GNA) in the gene C2orf37,mapped on chromosomes 2q22.3–2q35.

2. Materials and methods

2.1. Human subjects

A four-generation consanguineous family with clinical manifesta-tions of WSS was recruited from a remote region of Pakistan. The ped-igree (Fig. 1) provided convincing evidence of an autosomal recessivemode of inheritance. Seven individuals including a deceased (III-5) inthe family were affected with WSS. Six affected individuals (IV-1, IV-2, IV-7, IV-8, IV-9, IV-10) underwent careful clinical examination at alocal government hospital. The study was approved by InstitutionalReview Board (IRB) of Quaid-i-Azam University, Islamabad, Pakistan.Informed written consent for the study, including presentation ofphotographs for publication, was obtained from elders includingparents of the affected individuals in the family.

2.2. Isolation of genomic DNA and genotyping

Venous blood sampleswere collected from4 affected (IV-1, IV-2, IV-8,IV-9) and 6 unaffected individuals (III-3, III-6, III-7, III-8, 1V-3, IV-11) inEDTA containing blood vacutainer set. Genomic DNA was extracted

Fig. 1. Pedigree of a consanguineous Pakistani family segregating Woodhouse-Sakati syndrome. Circles and squares represent females and males, respectively. Clear symbols representunaffected individuals whilefilled symbols represent affected individuals. For genotyped individuals, haplotypes are shown beneath each symbol. Genetic distances in centiMorgans (cM)are depicted according to the Rutgers combined linkage-physical map (build 36.2) (Matise et al., 2007).

27R. Habib et al. / Gene 490 (2011) 26–31

from peripheral-blood lymphocytes using standard procedures. Poly-merase chain reaction (PCR) was carried out in 25 μl reaction volumescontaining 40 ng genomic DNA, 20 pmol of each primer, 200 μM ofeach deoxynucleoside triphosphate (dNTP), 2.5 μl reaction buffer (KCl50 mM, Tris–Cl pH 8.3, 1.5 mMMgCl2) and 1 unit of Taq DNA polymer-ase (MBI Fermentas, Life Sciences, York, UK). The thermal cycling condi-tions used included 95 °C for 5 min, followed by 40 cycles of 95 °C for1 min, 55–57 °C for 1 min, 72 °C for 1 min and a final extension at72 °C for 10 min. The PCR was performed by GeneAmp® PCR system9700 obtained from Applied Biosystems (Applera Corp, Foster City,CA, USA). The amplified product was resolved on 8% non-denaturingpolyacrylamide gel and visualized by ethidium bromide staining toscore the alleles by manual inspection.

The family was tested first for linkage by genotyping highly polymor-phic microsatellite markers linked to genes involved in alopecia withmental retardation, and alopecia associated with hypogonadismand other phenotypes. These included three alopecia and mentalretardation syndromes APMR1 at chromosome 3q26.33 (D3S2314,D3S3578, D3S3583, D3S3592, D3S1530, D3S1617, D3S1602, D3S1262),APMR2 at chromosome 3q26.2 (D3S3656, D3S2433, D3S3053,D3S1556, D3S2425, D3S2421, D3S2309) and APMR3 at chromosome18q11.2–q12.2 (D18S1151, D18S877, D18S847, D18S536, D18S1100,D18S1102, D18S811), human aristaless like 4 (ALX4, MIM 605420)at chromosome 11p11.2–q12.3 (D11S1993, D11S1393, D11S903,D11S986, D11S4103) and chromosome 2 open reading frame 37(C2orf37, MIM 612515) involved in Woodhouse Sakati syndrome atchromosomes 2q22.3–2q35 (D2S1281, D2S2284, D2S2302, D2S1267,D2S2307, D2S138, D2S2978, D2S364, 2S426, D2S118, D2S389,D2S280, D2S117, D2S2318, D2S2392, D2S309).

Haplotypes were constructed using SIMWALK2 (Sobel and Lange,1996). Two point and multipoint LOD score was calculated usingALLEGRO version 2 (Gudbjartsson et al., 2005). Two-point LOD scoreswere computed for recombination fraction values of θ=0.00, 0.01,0.05, 0.10, 0.20, 0.30 and 0.40. Autosomal recessive mode of inheri-tance with complete penetrance and a disease allele frequency of0.001 were used. Order of markers was based on Rutgers combinedlinkage physical map of the human genome (Matise et al., 2007).

2.3. Sequencing C2orf37

After establishing linkage of the family to C2orf37, mapped on chro-mosome 2q22.3–2q35, entire coding region and splice junction sites ofthe gene were sequenced for potential sequence variants. Primersequences were designed for each exon using Primer3 software(Rozen and Skaletsky, 2000) and checked for specificity using basiclocal alignment search tool (Basic local alignment search tool; http://www.ncbi.nlm.nih.gov/blast). DNA sequence of the gene C2orf37 wasobtained from UCSC Human Genome browser (UCSC genome bio-informatics website; http://genome.ucsc.edu/cgi-bin/hgGateway).DNA from all four affected and six unaffected individuals of the familywere amplified by PCR, purified and sequenced under standard condi-tions. Amplified DNA fragments were purified using the Rapid PCR Pu-rification System (Marligen Biosciences, Ijamsville, MD, USA) andwere sequenced in an ABI Prism 310 automated DNA sequencer usingthe BigDye® Terminator Cycle Sequencing Kits v3.1 (Applied Biosys-tems, Foster City, CA, USA) under standard conditions. Sequence vari-ants were identified via BIOEDIT sequence alignment editor version6.0.7.

28 R. Habib et al. / Gene 490 (2011) 26–31

3. Results

3.1. Clinical findings

Six affected individuals (IV-1, IV-2, IV-7, IV-8, IV-9, IV-10) of thefamily presented features compatible with those reported earlier in pa-tients of WSS. Medical officials at the local government hospitalreported the presence of hypogonadism, intellectual disability, mildsensory neural hearing loss, alopecia and extrapyramidal disorder inaffected individuals of our family (Table 1). Alopecia in the affectedindividuals was characterized by the presence of thin fragile hair onscalp, sparse eyebrows and eyelashes, and complete absence of body,axillary and pubic hair. Triangular facial appearance with prominentnasal roots, prominent ears with large earlobes and wrinkled skin inprogeric fashion were observed in the affected individuals (Fig. 2).

The affected female subjects (IV-1, IV-2, IV-9) reported onset ofhypogonadism in early adolescence. During puberty, all three femalesubjects experienced a condition of primary amenorrhea and absenceof secondary sexual features including breast tissue. At the age of22 years, an affected female patient (IV-1) displayed completeedentulism while her sister (IV-2) who was 16 years old at the timeof the study showed partial edentulism. Blepharospasmwas observedin an affected female individual (IV-2). Another affected female indi-vidual (IV-9), aged 17, experienced generalized dystonic and involun-tary choreoathetoid movements of the body leading to walking andgait difficulties. The dystonic movements of the limbs started at theage of 13 years, which later progressed to arm and leg tremors,head shaking, postural impairment and freezing of gait.

Table 1Variable clinical manifestations reported in six patients with Woodhouse–Sakati syndrome

Affected individuals IV-1 IV-2

Gender Female FemaleAge(years) 22 16

Ectodermal featuresAlopecia + +Edentalism (toothless) Complete Partial

Neurological featuresSensorineural deafness + +Intellectual disability Moderate ModerateExtrapyramidal symptoms − +

• Dystonia − −• Choreoathetosis − −• Blepharospasm − +

Spastic quadriplegia − −Dysarthria − −Dysphagia − −

Endocrine featuresHypogonadism + +

• primary amenorrhea + +• Failure of secondary sexualcharacters development

+ +

Other featuresDysmorphic facial features + +

• High forehead,Triangular face, Prominentnasal root, hypertelorism,.

+ +

Precocious skin aging + +

Laboratory tests• IGF-1 (ng/μl) NA NA• LH (mIU/ml) NA NA• FSH (mIU/ml) NA NA• Testosterone (ng/dl) NA NA

+ indicated feature present, − indicated feature absent, NA not available.

In three affected male individuals (IV-7, IV-8, IV-10) secondarysexual characteristics and genitalia were underdeveloped. In two ofthese individuals (IV-7, IV-8), levels of IGF-1, LH, FSH and testoster-one were decreased (Table 1). Signs of movement disorder were ob-served only in one of the three male affected individuals. Thisindividual (IV-10), at the age of 14 years, showed clinical featuressimilar to those reported in patients with spastic quadriplegia. He dis-played muscle stiffness, postural impairment, unsteady gait, inabilityto walk, dysarthria and dysphagia.

Magnetic resonance imaging (MRI) and computed tomography(CT) scan images of the affected individuals were not available forthe study. History of cardiac, diabetic and immunological problemswas not reported in any of the affected individuals. Glycated hemo-globin (HbA1C) levels measured in two individuals (IV-7, IV-8)were in the range of 30–33 nmol/mol. Obligate heterozygous carriersin the family were normal and were clinically indistinguishable fromgenotypically normal individuals.

3.2. Genotyping and mutation analysis

Linkage of the family was tested to genes including APMR1(3q26.33), APMR2 (3q26.2), APMR3 (18q11.2–q12.2), ALX4(11p11.2–q12.3) and C2orf37 (2q22.3–2q35). Haplotype analysisshowed linkage of the family to C2orf37, mapped on chromosome2q22.3–2q35. Parametric two-point and multipoint LOD scores werecalculated using ALLEGRO version 2 (Gudbjartsson et al., 2005). Max-imum two-point LOD score of 2.58 was obtained with marker D2S138and maximum multipoint LOD score of 3.64 was obtained with three

.

IV-7 IV-8 IV-9 IV-10

Male Male Female Male23 18 17 14

+ + + +− − − −

+ + + +Moderate Moderate Moderate Moderate− − + +− − + +− − + +− − − −− − − +− − − +− − − +

− + + +− − + −+ + + +

+ + + ++ + + +

− − + −

272 315 NA NA0.5 0.8 NA NA1.0 0.8 NA NA90 110 NA NA

Fig. 2. Clinical features of patients with Woodhouse-Sakati syndrome. Phenotypic appearance of affected individual IV-2 at 16 years of age (a), IV-9 at 17 years of age (b), IV-10 at14 years of age (c) IV-8 at 18 years of age (d). Note thin hair on scalp, sparse eyebrows and eyelashes, triangular facial shape with the large earlobes and precocious skin ageing.

29R. Habib et al. / Gene 490 (2011) 26–31

markers (D2S1267, D2S2307, D2S138) along the disease interval(Table 2). C2orf37 was then sequenced in all those affected and unaf-fected individuals of the family for whom the DNA was available.Sequence analysis of the gene C2orf37 in affected subjects revealeda novel splice site mutation involving a homozygous G→A transitionin the splice donor site of intron 3 (c.321+1 GNA) (Fig. 3).

Table 2Two-point and multipoint LOD score between Woodhouse–Sakati syndrome and microsate

Marker Mba cMb mptc Two-point LOD score

0.00 0.01

D2S354 163.60 170.72 −4.77 − Inf −2.D2S124 165.85 171.59 −6.14 − Inf −2.D2S1379 167.58 172.25 0.04 − Inf −2.D2S1281 168.98 174.19 3.18 −1.21 −0.D2S2284 171.20 177.92 3.61 2.18 2.13D2S2302 172.26 178.56 3.63 2.18 2.13D2S1267 174.02 181.57 3.64 2.18 2.13D2S2307 175.16 182.48 3.64 2.18 2.13D2S138 177.45 184.67 3.64 2.58 2.52D2S2978 180.82 188.07 3.24 2.18 2.13D2S364 182.74 189.47 −2.67 − Inf 0.49D2S426 190.06 192.53 − Inf − Inf 0.53

a Physical distance (mega base pairs).b Genetic distance (centiMorgans) according to the second-generation combined linkagec Multi-point LOD Score.

The pathogenic sequence variant reported here was found in theheterozygous state in the obligate carriers and segregated with thedisease in the family. To exclude the possibility that the novel splicesite mutation do not represent a non-pathogenic polymorphism, apanel of 100 unrelated ethnically matched control individuals werescreened and this mutation was not identified outside the family.

llite markers of chromosome 2q22.3–q35.

at θ=

0.05 0.10 0.20 0.30 0.40

10 −0.84 −0.36 −0.04 0.02 0.0048 −1.18 −0.66 −0.24 −0.08 −0.0110 −0.84 −0.36 −0.04 0.02 0.0041 0.19 0.38 0.39 0.26 0.12

1.93 1.68 1.18 0.70 0.271.93 1.68 1.18 0.70 0.271.93 1.68 1.18 0.70 0.271.93 1.68 1.18 0.70 0.272.29 2.00 1.42 0.87 0.361.93 1.68 1.18 0.70 0.270.98 1.01 0.80 0.50 0.201.02 1.06 0.84 0.53 0.23

physical map of the human genome (Matise et al., 2007).

Fig. 3. Sequence analysis of a novel splice site mutation (c.321+1 GNA) in C2orf37. Theupper panel (a) represents nucleotide sequences in the control unaffected individual,the middle panel (b) in the heterozygous carrier and the lower panel (c) in the affectedindividual. Arrows represent position of the mutations.

30 R. Habib et al. / Gene 490 (2011) 26–31

4. Discussion

In the present investigation,we have reported clinical andmolecularanalysis of a four generation consanguineous Pakistani family present-ing clinical manifestations of Woodhouse–Sakati syndrome (WSS). Af-fected individuals of the family displayed clinical features such asintellectual disability, hypogonadotropic hypogonadism, mild sensoryneural hearing loss, partial alopecia and extra-pyramidal disorder. Dia-betesmellituswas not found in any of the affected individual. Intrafami-lial phenotypic variability including the extent of extra-pyramidalmanifestations and edentulism was observed among the affected indi-viduals of the family. Edentulism, a condition of being toothless, wasobserved only in two affected females (IV-1, IV-2) of the present family.Apart from one report which described the presence of anodontia(Steindl et al., 2010), dental anomalies have not been reported in anyother case of WSS. The extra-pyramidal manifestations observed inthree affected individuals of the present family present chreoathetoidand dystonic movements leading to walking and gait disturbance.Similar features of extra-pyramidal have been reported earlier inother familieswithWWS (Al-Semari and Bohlega, 2007). These authorshave also reported interfamilial and intrafamilial phenotypic variabilityof WWS in twelve families they have studied.

Genotyping analysis established linkage of the family, presentedhere, to the gene C2orf37 at WSS locus on chromosome 2q22.3–2q35. Subsequently, sequence analysis of the gene C2orf37 revealeda novel donor splice site mutation (c.321+1 GNA) in all affected in-dividuals of the family (Fig. 3). Up till now eight different disease-causing mutations including three deletions, three nonsense andtwo splicing errors have been reported in the gene C2orf37 (Alazamiet al., 2008, 2010; Steindl et al., 2010). All of these mutations lead toprotein truncations.

The mutation (c.321+1 GNA), identified in our family, is the thirdsplice sitemutation in the gene C2orf37, reported to date. Mutations atsplice junction sites of the genes make a significant contribution tohuman genetic diseases, since approximately 15% of disease-causingpoint mutations affect pre-mRNA splicing (Krawczak et al., 1992).Such splice site mutations may result in exon skipping, activation ofa cryptic splice site, creation of a pseudo-exon within an intron,and intron retention (Nakai and Sakamoto, 1994). According to theBerkeley Drosophila Genome Project (Berkeley Drosophila GenomeProject; http://www.fruitfly.org/), change in the splice donor sequencefrom GT to AT in intron 3 (c.321+1 GNA) of C2orf37, detected in thepresent family, is predicted to result in a dramatic drop in splice siterecognition by splicing factors from 0.95 to 0.0, essentially leading toabolition of the normal splice donor site. A possible outcome of suchalteration in the splice donor sitewould be either loss of C2orf37 expres-sion, possibly due to nonsense mediated mRNA decay (Weischenfeldtet al., 2005) or synthesis of premature truncated protein, in eithercase resulting in loss-of-function effect on the final gene product.

The C2orf37 encodes a nucleolar protein with two major isoforms,called alpha and beta. The protein lacks any recognizable domain orlocalization signal and shares no significant homology with other pro-teins (Alazami et al., 2008). The amino acid sequence is well con-served across species, present in the mouse, rat, chimpanzee, cow,fowl and other higher animals (Alazami et al., 2008). Expression pro-file of C2orf37 in various adult human tissues revealed low ubiquitousexpression of both protein isoforms. Immunohistochemical analysisshowed nearly ubiquitous nucleolar expression of C2orf37 in mouseembryos, with increased expression in brain, liver, and skin which isrelevant to the fact that disease phenotype reported in human is con-fined to these organs (Alazami et al., 2008). Despite of its ubiquitousexpression, C2orf37 deficiency results in clinical manifestation con-fined to limited number of organs, potentially displaying functionalredundancy, which is also true for mutations affecting other nucleolarproteins (Nousbeck et al., 2008).

The precise function performed by C2orf37 is poorly understood.According to Alazami et al. (2008) nucleolar defect may underliethe pathogenesis of WSS. The nucleolus is associated with the regula-tion of a number of major physiological cellular processes includingribosome biogenesis, regulation of cell cycle, cellular aging, signalrecognition- particle biosynthesis, small-RNA processing, mRNAtransport, and even apoptosis (Boisvert et al., 2007). Disruption inany of these nucleolar processes may underlie the pathogenesis ofWSS, most favored hypothesis being defective ribosome biogenesisas possible pathogenic mechanism in WSS (Alazami et al., 2008).The discovery of other defective nucleolar protein like RBM28 in-volved in another multisystem disorder ANE (alopecia, neurologicaldefects, and endocrinopathy) syndrome (Nousbeck et al., 2008) high-lights the vital role nucleolar proteins play in regulation of variouskey physiological processes. Further studies of functions of thesenucleolar genes will shed light on the physiological important roleplayed by individual constituents of nucleolus.

Acknowledgments

We are grateful to all members of the family for their invaluableparticipation and cooperation. The work presented here was fundedby Higher Education Commission (HEC), Islamabad, Pakistan. RabiaHabib was supported by indigenous PhD fellowships fromHEC, Islam-abad, Pakistan.

References

Alazami, A.M., et al., 2008. Mutations in C2orf37, encoding a nucleolar protein, causehypogonadism, alopecia, diabetes mellitus, mental retardation, and extrapyrami-dal syndrome. Am. J. Hum. Genet. 83, 684–691.

Alazami, A.M., et al., 2010. C2orf37 mutational spectrum in Woodhouse–Sakatisyndrome patients. Clin. Genet. 78, 585–590.

31R. Habib et al. / Gene 490 (2011) 26–31

Al-Semari, A., Bohlega, S., 2007. Autosomal-recessive syndrome with alopecia,hypogonadism, progressive extra-pyramidal disorder, white matter disease,sensory neural deafness, diabetes mellitus, and low IGF1. Am. J. Med. Genet. A143, 149–160.

Al-Swailem, S.A., Al-Assiri, A.A., Al-Torbak, A.A., 2006. Woodhouse Sakati syndromeassociated with bilateral keratoconus. Br. J. Ophthalmol. 90, 116–117.

Boisvert, F.M., van Koningsbruggen, S., Navascués, J., Lamond, A.I., 2007. The multifunc-tional nucleolus. Nat. Rev. Mol. Cell Biol. 80, 574–585.

Gudbjartsson, D.F., Thorvaldsson, T., Kong, A., Gunnarsson, G., Ingolfsdottir, A., 2005.Allegro version 2. Nat. Genet. 37, 1015–1016.

Krawczak, M., Reiss, J., Cooper, D.N., 1992. The mutational spectrum of single base-pairsubstitutions in mRNA splice junctions of human genes: causes and consequences.Hum. Genet. 90, 41–54.

Matise, T.C., et al., 2007. A second-generation combined linkage physical map of thehuman genome. Genome Res. 17, 1783–1786.

Nakai, K., Sakamoto, H., 1994. Construction of a novel database containing aberrantsplicing mutations of mammalian genes. Gene 41, 171–177.

Nousbeck, J., et al., 2008. Alopecia, neurological defects, and endocrinopathy syndromecaused by decreased expression of RBM28, a nucleolar protein associated withribosome biogenesis. Am. J. Hum. Genet. 82, 1114–1121.

Rozen, S., Skaletsky, H., 2000. Primer3 on the WWW for general users and for biologistprogrammers. Methods Mol. Biol. 132, 365–386.

Schneider, S.A., Bhatia, K.P., 2008. Dystonia in the Woodhouse–Sakati syndrome: a newfamily and literature review. Mov. Disord. 23, 592–596.

Sobel, E., Lange, K., 1996. Descent graphs in pedigree analysis: applications to haplotyping,location scores, and marker-sharing statistics. Am. J. Hum. Genet. 58, 1323–1337.

Steindl, K., et al., 2010. A novel C2orf37 mutation causes the first Italian cases ofWoodhouse–Sakati syndrome. Clin. Genet. 78, 594–597.

Weischenfeldt, J., Lykke-Andersen, J., Porse, B., 2005. Messenger RNA surveillance: neu-tralizing natural nonsense. Curr. Biol. 15, 559–562.

Woodhouse, N.J., Sakati, N.A., 1983. A syndrome of hypogonadism, alopecia, diabetesmel-litus,mental retardation, deafness, and ECG abnormalities. J.Med. Genet. 20, 216–219.

Web resources

Mendelian inheritance in man (MIM): http://www.ncbi.nlm.nih.gov/omim/.UCSC genome bioinformatics website: http://genome.ucsc.edu/cgi-bin/hgGateway.Basic local alignment search tool (BLAST): http://www.ncbi.nlm.nih.gov/blast.Berkeley Drosophila Genome Project: http://www.fruitfly.org/.