Molecular genetic cause of X-linked retinitis pigmentosa in a Czech family

Abstract

Editor, A branch of a large eight generation Caucasian Czech family diagnosed with presumable X-linked retinitis pigmentosa (XLRP) was investigated (Fig. 1). This study adhered to the tenets of the Declaration of Helsinki. Men were severely affected in this family with early onset of night blindness and myopia, usually at the beginning of the second decade, and progressive loss of visual function resulting in blindness in the fourth decade. Women were either asymptomatic or developed symptoms with later onset (Otradovec et al. 1979). There is no evidence of male to male transmission. After obtaining informed consent, blood samples were drawn and DNA extracted. Polymorphic microsatellite markers located on the X chromosome (Table 1) were amplified using fluorescently labelled oligonucleotides and analysed on an ABI-377 DNA sequencer (Applied Biosystems, Darmstadt, Germany) using GeneScan software (Applied Biosystems). Haplotype analysis indicated that the disease segregates with markers encompassing both the RPGR and RP2 genes as RPGR lies between markers DXS1214 and DXS1068 and RP2 between DXS1055 and DXS991 (Fig. 1). The chromosomal region between the most proximal and most distal marker used in this study encompassed approximately 200 known protein-coding genes (http://www.ensembl.org). Investigated branch of an eight generation Czech family with X-linked retinitis pigmentosa. The affected haplotype is shown as a black bar and encompasses the RPGR and RP2 genes located between DXS1214–DXS1068 and DXS1055–DXS991, respectively. Sequencing of the RP2 gene, as previously described (Hardcastle et al. 1999), did not reveal any pathogenic changes. ORF15 of the RPGR gene (Vervoort et al. 2000) was then amplified and bidirectionally sequenced in four overlapping fragments using the ABI Prism® BigDye™ dGTP Terminator Ready Reaction Cycle Sequencing Kit (Applied Biosystems). A pathogenic mutation, c.2426_2427 delAG (g.ORF15+673_674delAG), leading to p.Glu809fs (ORF15Glu224fs) was identified in all affected males and three female obligate carriers (Fig. 2). The change was also detected in one other female family member (individual IV:6) and was absent in females IV:2 and IV:4 (1, 2). This mutation has been observed previously in different XLRP patient populations (http://rpgr.hgu.mrc.ac.uk), including one Japanese XLRP families (Jin et al. 2006). Interestingly, no other sequence variants were identified in the highly polymorphic RPGR ORF15 in our family. Sequence chromatogram showing the c.2426_2427delAG mutation in ORF15 of the RPGR gene in an obligate carrier III:1 (A) and an affected male IV:5 (B). The mutation was not detected in an unaffected female IV:4 (C). Reference sequence NM_001034853 was used. All three affected men were documented to have bone spicules on fundus examination. There were marked interindividual differences between IV:1 and IV:5 despite similar age; IV:5 was much more severely affected with night blindness, whereas IV:1 reported no symptoms. II:3 was registered blind in the fourth decade. Examined female carriers (III:1, III:5, IV:6) had some peripheral pigmentary changes but not classic bone spicules. RPRG mutations are the most frequent cause of XLRP (Vervoort et al. 2000; http://rpgr.hgu.mrc.ac.uk). The pedigree we have investigated here represents the first Czech family with an identified molecular genetic cause of RP. Although in Western European countries the prevalence and mutational spectrum of various RP types has been extensively studied, little is known about this heterogeneous group of disorders and their molecular genetic cause in the restructured countries of Central and Eastern Europe. To the best of our knowledge, prior to this study, only one other report describes a pathogenic mutation causing XLRP in a family of Bulgarian Gypsy origin (Chakarova et al. 2006). Here, we would like to propose the necessity for service offering molecular genetic testing for RP patients, and also for patients with other inherited ocular disorders, in all European countries. This should be supported by continuous education in the field of ophthalmic genetics among medical practitioners which would lead to improved counselling. However, as it is technically challenging to establish mutation analysis for every gene causing inherited ocular disorders in every country, we suggest that where X-linked inheritance cannot be excluded, the RPGR gene is screened for mutations using already established diagnostic services. This work was in part supported by the Czech Ministry of Education, Youth and Sports research project 0021620806/20610011.