A novel CISD2 intragenic deletion, optic neuropathy and platelet aggregation defect in Wolfram syndrome type 2
- Enza Mozzillo†1Email author,
- Maurizio Delvecchio†2,
- Massimo Carella3,
- Elvira Grandone4,
- Pietro Palumbo3,
- Alessandro Salina5,
- Concetta Aloi5,
- Pietro Buono1,
- Antonella Izzo6,
- Giuseppe D’Annunzio5,
- Gennaro Vecchione4,
- Ada Orrico7,
- Rita Genesio6,
- Francesca Simonelli7 and
- Adriana Franzese1
© Mozzillo et al.; licensee BioMed Central Ltd. 2014
Received: 6 May 2014
Accepted: 11 July 2014
Published: 24 July 2014
Wolfram Syndrome type 2 (WFS2) is considered a phenotypic and genotypic variant of WFS, whose minimal criteria for diagnosis are diabetes mellitus and optic atrophy. The disease gene for WFS2 is CISD2. The clinical phenotype of WFS2 differs from WFS1 for the absence of diabetes insipidus and psychiatric disorders, and for the presence of bleeding upper intestinal ulcers and defective platelet aggregation. After the first report of consanguineous Jordanian patients, no further cases of WFS2 have been reported worldwide. We describe the first Caucasian patient affected by WFS2.
The proband was a 17 year-old girl. She presented diabetes mellitus, optic neuropathy, intestinal ulcers, sensorineural hearing loss, and defective platelet aggregation to ADP. Genetic testing showed a novel homozygous intragenic deletion of CISD2 in the proband. Her brother and parents carried the heterozygous mutation and were apparently healthy, although they showed subclinical defective platelet aggregation. Long runs of homozygosity analysis from SNP-array data did not show any degree of parental relationship, but the microsatellite analysis confirmed the hypothesis of a common ancestor.
Our patient does not show optic atrophy, one of the main diagnostic criteria for WFS, but optic neuropathy. Since the “asymptomatic” optic atrophy described in Jordanian patients is not completely supported, we could suppose that the ocular pathology in Jordanian patients was probably optic neuropathy and not optic atrophy. Therefore, as optic atrophy is required as main diagnostic criteria of WFS, it might be that the so-called WFS2 could not be a subtype of WFS. In addition, we found an impaired aggregation to ADP and not to collagen as previously reported, thus it is possible that different experimental conditions or inter-patient variability can explain different results in platelet aggregation. Further clinical reports are necessary to better define the clinical spectrum of this syndrome and to re-evaluate its classification.
KeywordsCISD2 Optic neuropathy Non-autoimmune diabetes mellitus Novel mutation Platelet aggregation Sensorineural hearing loss SNP-array Upper intestinal ulcers Wolfram syndrome
Wolfram syndrome (WFS; MIM 222300) is a rare neurodegenerative disease with autosomal recessive inheritance and incomplete penetrance, whose diagnosis requires diabetes mellitus (DM) and optic atrophy (OA). WFS includes a wide spectrum of other possible disorders, such as diabetes insipidus, sensorineural deafness, genitourinary tract problems, male hypogonadism, neurological or psychiatric disorders, and less frequently, bowel dysfunction. Clinical wide spectrum is likely sustained by a genetic heterogeneity . To date, 2 types of WFS, type 1 (WFS1; MIM 606201) and type 2 (WFS2; MIM 604928) characterized by different disease genes have been identified. The disease gene for WFS2 is CISD2, encoding for a highly conserved zinc-finger protein of the Endoplasmic Reticulum Intermembrane Small (ERIS), playing a pivotal role in calcium homeostasis (Ca2H) . It was identified in three consanguineous Jordanian families carrying a point mutation in exon 2 . In WFS2, diabetes insipidus and psychiatric disorders are not described, whereas bleeding upper intestinal ulcers (b-UIU), not reported in WFS1 , and defective platelet aggregation (PA) may be considered diagnostic criteria. After the first report [3, 5], no further CISD2 mutations have been reported. We describe the first Caucasian patient with WFS2 with a novel homozygous CISD2 intragenic deletion.
Ophtalmological abnormalities, DM and deafness were not found in the parents and brother, whereas a reversible PA in response to ADP was found. Currently, FS is 22 years old and presents DM, ON with preserved visual acuity and electrophysiologic tests, no OA, tinnitus due to SHLHF triggered by loud environments, EGBR, slow intestinal transit, reduced and reversible defective PA to ADP, and b-UIU.
Matherials and methods
DNA from the proband, parents, brother and controls were extracted from whole blood using High Pure PCR Template Preparation Kit (Roche, Mannheim, Germany). The three exons and the flanking region of CISD2 gene were amplified by PCR (PrimeStar GXL DNA Polymerase, Takara Bio Inc, Japan) using three couples of primers. Amplicons were purified and then sequenced for both sense and antisense strands. CISD2 cDNA sequencing was performed using known cDNA primers and conditions . SNP-array analysis on trios was performed using the Affymetrix CytoScan HD Array (Affymetrix, Santa Clara, CA) as previously described .
Patients were enrolled after obtaining appropriate informed consent and approval by the local ethics committee (#08-CE) of the IRCCS Casa Sollievo della Sofferenza (San Giovanni Rotondo) where genetics studies were carried out.
Identification of novel mutation in CISD2
In the patient the amplification of CISD2 exon 2 was not detected, while exons 1 and 3 were regularly present. Moreover exons 1, 2 and 3 were amplified in parents, in brother and in controls. CISD2 cDNA sequencing revealed a novel homozygous deletion affecting the whole exon 2 of CISD2 (see Additional file 1: Figure S1). To further confirm the exon 2 deletion, to map boundaries, and to confirm that her parents and brother were carriers, a SNP-array analysis on trios was performed. Log2Ratio values of markers encompassing the CISD2 deletion were of approximately -0.45 to -1, suggestive for a heterozygous deletion (mother, father and brother) while Log2Ratio values from -1 to higher negative values were suggestive for homozygous deletion.
Log2Ratio value of markers encompassing the CISD2 deletion on the CytoScan HD array
The percentage of estimate consanguinity expressed in term of Coefficient of Inbreedeing and Identical by Descent
Coefficient of inbreeding
IBD (Identical by Descent %)
We describe the first European Caucasian girl with WFS2, carrying a novel intragenic exon 2 CISD2 homozygous deletion. Only a homozygous mutation of the CISD2, a single nucleotide substitution that leads to a missense change Glu37Gln, was previously associated to WFS2 . This gene, mapping on chromosome 4q24 , encodes an extremely small zinc-finger protein, named ERIS, which displays a fairly wide cellular expression profile, including pancreas, brain, blood and platelets . Conlan et al.  showed that the domain 2Fe-2S is essential for the activity of this zinc-finger protein, and is located from aminoacid 99 (Cys) to aminoacid 112 (Gly). Genomic data from our subjects show that the triplets coding for aminoacids WRSKT, from the position 102 to 106 of the polypeptide, are within the deleted region, and this may result in a non functional protein because of zinc-finger domain lost. The ERIS protein is highly conserved and has 70% identity and 82% similarity across all the vertebrate species . It plays a pivotal role in Ca2H , as well as wolframin, another endoplasmic reticulum (ER)  protein encoded by WFS1 gene. Also wolframin is involved in the regulation of ER stress and Ca2H [11, 12], without any interaction with ERIS. The ER function influences some crucial secretory proteins, such as insulin, as well as cell-surface receptors and integral membrane proteins. Imbalance in Ca2H of ER elicits stress in this organelle, leading to the accumulation of misfolded and unfolded protein in the organelle, a state called ‘ER stress’, which plays, in WFS, a key role in β-cells, leaving these cells more prone to oxidative stress and consequently to apoptosis. Activation of the unfolded protein response (UPR), an adaptive response that counteracts the ER stress, serves to restore ER homeostasis. Moreover, UPR maintains pancreatic β-cell function and promotes β-cell survival. Exaggerated ER stress, which normally triggers the UPR, has been hypothesized in the β-cells impairing the cell progression and increasing apoptosis . Neuronal cells have highly developed ER, whose "stress" is related with ganglion cells apoptosis, as demonstrated in a glaucoma chronic injury model . The degeneration of ganglion and glial retinal cells could be related to the ER stress probably accountable for the ocular involvement in our patient that presented moderate ON but not OA. Few and non-specific studies on ERIS localization and expression, in particular in the ganglion and glial cells of the optic nerve, may explain the different optic nerve involvement and a favourable outcome of ocular pathology. In WFS1, early OA causes alterations of the optic nerve caliber due to the loss of a considerable number of ganglion cells . These patients show severe progressive visual acuity reduction and non recordable p-VEP or increased latency or greatly reduced amplitudes [16, 17]. In Jordanian WFS2 patients OA was observed after childhood or was described as “asymptomatic”, but opportune investigations were not reported . Since our patient does not show OA at the age of 22 years, and since “asymptomatic” OA described in Jordanians is not completely supported, we suppose that the ocular pathology in those patients was probably ON and not OA. This is just a case report and thus great caution is needed, even if the absence of OA, one of the main diagnostic criteria of WFS, suggests that the so-called WFS2 could not be a WFS subtype.
Clinical features of WFS1, WFS2, and of our case
WFS2 our case
Neurological and Psychiatric disorders
Genitourinary tract problems
Upper intestinal ulcers
Platelet aggregation defects to collagen
Platelet aggregation defects to ADP
Furthermore, in Table 3 we show the difference in the clinical spectrum of Jordanian cases  and our patient. Firstly, our girl presented ON and not OA even though as previously discussed it is possible that the ocular pathology in those patients was probably ON and not OA; secondly, she showed an impaired platelet aggregation to ADP and not to collagen; however collagen- and ADP- induced aggregation are both calcium-mediated, thus it is possible that different experimental conditions or inter-patient variability can explain different results in PA.
The new findings of this report could better characterize the WFS2 phenotype.
We describe the first Caucasian patient with so-called WFS2. She carried a novel mutation in CISD2. Our data debate the previous description of WFS2 pointing out new data about eye and platelets involvement. The proband presented optic neuropathy and not optic atrophy, impaired platelet aggregation to ADP and not to collagen. Further clinical reports are necessary to better define the clinical spectrum of this syndrome and to re-evaluate its classification.
This study was approved by institutional ethics committee and was done in accordance with Declaration of Helsinki. Informed consent for genetic analysis was obtained from the study participants. Moreover, as the patient is a girl, written informed consent was obtained from her parents for publication of this Case report. A copy of the written consent is available for review by the Editor of this journal.
WFS type 1
WFS type 2
Endoplasmic reticulum intermembrane small
Best corrected visual acuity
Pattern visual evoked potentials
Optical coherence tomography
brain Magnetic Resonance Imaging
Bleeding upper intestinal ulcers
Sensorineural hearing loss in the high frequencies
Entero-gastric bile reflux
Unfolded protein response.
The authors thank The Genomic Disorder Biobank, member of the Telethon Network of Genetic Biobanks funded by Telethon Italy (project no. GTB12001) and of the EuroBioBank network for providing us with the specimen used in this study. The study was partially supported both by Italian Ministry of Health grant RC2013 to MC and by POR Campania FSE 2007-20013, Project CREME from Campania Region to Prof. L. Nitsch (Department of Molecular Medicine and Medical Biotechnology, University of Naples "Federico II", Italy). Prior publication of the study in abstract form: Diabetologia (2013) 56: Suppl 1 S1-S566; 360. An abstract with preliminary genetic findings was presented as poster presentation at the 49th EASD Annual Meeting of the European Association for the Study of Diabetes. Diabetologia (2013) 56: Suppl 1 S1-S566; 360
- El-Shanti H, Lidral AC, Jarrah N, Druhan L, Ajlouni K: Homozygosity mapping identifies an additional locus for Wolfram syndrome on chromosome 4q. Am J Hum Genet. 2000, 66 (4): 1229-1236.View ArticlePubMedPubMed CentralGoogle Scholar
- Solovyova N, Veselovsky N, Toescu EC, Verkhratsky A: Ca(2+) dynamics in the lumen of the endoplasmic reticulum in sensory neurons: direct visualization of Ca(2+)-induced Ca(2+) release triggered by physiological Ca(2+) entry. EMBO J. 2002, 21 (4): 622-630.View ArticlePubMedPubMed CentralGoogle Scholar
- Amr S, Heisey C, Zhang M, Xia XJ, Shows KH, Ajlouni K, Pandya A, Satin LS, El-Shanti H, Shiang R: A homozygous mutation in a novel zinc-finger protein ERIS, is responsible for Wolfram syndrome 2. Am J Hum Genet. 2007, 81 (4): 673-683.View ArticlePubMedPubMed CentralGoogle Scholar
- Al-Sheyyab M, Jarrah N, Younis E, Shennak MM, Hadidi A, Awidi A, el-Shanti H, Ajlouni K: Bleeding tendency in Wolfram syndrome: a newly identified feature with phenotype genotype correlation. Eur J Pediatr. 2001, 160 (4): 243-246.View ArticlePubMedGoogle Scholar
- Ajlouni K, Jarrah N, El-Khateeb M, El-Zaheri M, El Shanti H, Lidral A: A Wolfram syndrome: identification of a phenotypic and genotypic variant from Jordan. Am J Med Genet. 2002, 115 (1): 61-65.View ArticlePubMedGoogle Scholar
- Odom JV, Bach M, Brigell M, Holder GE, McCulloch DL, Tormene AP, Vaegan : ISCEV standard for clinical visual evoked potentials (2009 update). Doc Ophthalmol. 2010, 120 (1): 111-119.View ArticlePubMedGoogle Scholar
- Tosetto A, Rodeghiero F, Castaman , Goodeve A, Federici AB, Batlle J, Meyer D, Fressinaud E, Mazurier C, Goudemand J, Eikenboom J, Schneppenheim R, Budde U, Ingerslev J, Vorlova Z, Habart D, Holmberg L, Lethagen S, Pasi J, Hill F, Peake I: A quantitative analysis of bleeding symptoms in type 1 von Willebrand disease: results from a multicenter European study (MCMDM-1VVD). J Thromb Haemost. 2006, 4 (4): 766-773.View ArticlePubMedGoogle Scholar
- Palumbo O, Fichera M, Palumbo P, Rizzo R, Mazzolla E, Cocuzza DM, Carella M, Mattina T: TBR1 is the candidate gene for intellectual disability in patientswith a 2q24.2 interstitial deletion. Am J Med Genet A. 2014, 164 (3): 828-833.View ArticleGoogle Scholar
- Kearney HM, Kearney JB, Conlin LK: Diagnostic implications of excessive homozygosity detected by SNP-based microarrays: consanguinity, uniparental disomy, and recessive single-gene mutations. Clin Lab Med. 2011, 31 (4): 595-613.View ArticlePubMedGoogle Scholar
- Conlan AR, Axelrod HL, Cohen AE, Abresch EC, Zuris J, Yee D, Nechushtai R, Jennings PA, Paddock ML: Crystal structure of Miner1: the redox-active 2Fe-2SProtein causative in Wolfram syndrome 2. J Mol Biol. 2009, 392 (1): 143-153.View ArticlePubMedPubMed CentralGoogle Scholar
- Fonseca SG, Fukuma M, Lipson KL, Nguyen LX, Allen JR, Oka Y, Urano F: WFS1 is a novel component of the unfolded protein response and maintains homeostasis of the endoplasmic reticulum in pancreatic b-cells. J Biol Chem. 2005, 280 (47): 39609-39615.View ArticlePubMedGoogle Scholar
- Zatyka M, Ricketts C, da Silva XG, Minton J, Fenton S, Hofmann-Thiel S, Rutter GA, Barrett TG: Sodium–potassium ATPase 1 subunit is a molecular partner of Wolframin, an endoplasmic reticulum protein involved in ER stress. Hum Mol Genet. 2008, 17 (2): 190-200.View ArticlePubMedGoogle Scholar
- Yamada T, Ishihara H, Tamura A, Takahashi R, Yamaguchi S, Takei D, Tokita A, Satake C, Tashiro F, Katagiri H, Aburatani H, Miyazaki J, Oka Y: WFS1-deficiency increases endoplasmic reticulum stress, impairs cell cycle progression and triggers the apoptotic pathway specifically in pancreatic beta-cells. Hum Mol Genet. 2006, 15 (10): 1600-1609.View ArticlePubMedGoogle Scholar
- Doh SH, Kim JH, Lee KM, Park HY, Park CK: Retinal ganglion cell death induced by endoplasmic reticulum stress in a chronic glaucoma model. Brain Res. 2010, 1308: 158-166.View ArticlePubMedGoogle Scholar
- Galluzzi P, Filosomi G, Vallone IM, Bardelli AM, Venturi C: MRI of Wolfram syndrome (DIDMOAD). Neuroradiology. 1999, 41 (10): 729-731.View ArticlePubMedGoogle Scholar
- Simsek E, Simsek T, Tekgül S, Hosal S, Seyrantepe V, Aktan G: Wolfram (DIDMOAD) syndrome: a multidisciplinary clinical study in nine Turkish patients and review of the literature. Acta Paediatr. 2003, 92 (1): 55-61.View ArticlePubMedGoogle Scholar
- Langwińska-Wośko E, Broniek-Kowalik K, Szulborski K: A clinical case study of a Wolfram syndrome-affected family: pattern-reversal visual evoked potentials and electroretinography analysis. Doc Ophthalmol. 2012, 124 (2): 133-141.View ArticlePubMedGoogle Scholar
- Peri S, Navarro JD, Amanchy R, Kristiansen TZ, Jonnalagadda CK, Surendranath V, Niranjan V, Muthusamy B, Gandhi TK, Gronborg M, Ibarrola N, Deshpande N, Shanker K, Shivashankar HN, Rashmi BP, Ramya MA, Zhao Z, Chandrika KN, Padma N, Harsha HC, Yatish AJ, Kavitha MP, Menezes M, Choudhury DR, Suresh S, Ghosh N, Saravana R, Chandran S, Krishna S, Joy M, et al: Development of human protein reference database as an initial platform for approaching systems biology in humans. Genome Res. 2003, 13 (10): 2363-2371.View ArticlePubMedPubMed CentralGoogle Scholar
- The pre-publication history for this paper can be accessed here:http://www.biomedcentral.com/1471-2350/15/88/prepub
This article is published under license to BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly credited. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.