Branchio–oto–renal syndrome: detection of EYA1 and SIX1 mutations in five out of six Danish families by combining linkage, MLPA and sequencing analyses

Abstract

The branchio -oto -renal (BOR) syndrome is an autosomal-dominant disorder characterized by hearing loss, branchial and renal anomalies. BOR is genetically heterogeneous and caused by mutations in EYA1 (8q13.3), SIX1 (14q23.1), SIX5 (19q13.3) and in an unidentified gene on 1q31. We examined six Danish families with BOR syndrome by assessing linkage to BOR loci, by performing EYA1 multiplex ligationdependent probe amplification (MLPA) analysis for deletions and duplications and by sequencing of EYA1, SIX1 and SIX5. We identified four EYA1 mutations (c.920delG, IVS10À1G4A, IVS12 þ 4A4G and p.Y591X) and one SIX1 mutation (p.W122R), providing a molecular diagnosis in five out of the six families (83%). The present, yet preliminary, observation that renal and temporal bone malformations are less frequent in SIX1-related disease suggests a slightly different clinical profile compared to EYA1-related disease. Unidentified mutations impairing mRNA expression or further genetic heterogeneity may explain the lack of mutation finding in one family despite LOD score indications of EYA1 involvement.

A recurrent EYA1 mutation causing alternative RNA splicing in branchio‐oto‐renal syndrome: Implications for molecular diagnostics and disease mechanism

“…The transcriptional effects of EYA1 mutations are essentially unknown due to the difficulty of obtaining appropriate samples for EYA1 RNA analysis. To date only four other patients with BOR syndrome have been reported with EYA1 transcript analysis, two of which had unstable EYA1 transcripts [Sanggaard et al, 2007], and two of which had transcripts with aberrant splicing in the Eya domain region [Okada et al, 2006]. Clinical features of the latter two patients included renal anomalies, similar to the patients reported here with the c.867 þ 5G > A allele.…”

“…Our overall mutation detection rate is higher than the 31% reported from EYA1 testing using denaturing high performance liquid chromatography (DHPLC) prior to sequencing [Orten et al, 2008], but is comparable to a recent report that four of six BOR families had EYA1 mutations when tested by methods similar to those used in our study [Sanggaard et al, 2007]. The use of DHPLC [Migliosi et al, 2004] may have led to underestimation of the contribution of EYA1 mutations to BOR, as DHPLC has reduced sensitivity.…”

“…The effect of different EYA1 mutations on transcription is likely to be variable and not predicted from genomic mutation information, as truncating or missense mutations may affect splicing [Boyer et al, 2005;Krawczak et al, 2007] while conserved splice site mutations may produce abnormal EYA1 transcripts which are highly unstable leading to complete absence of transcript [Sanggaard et al, 2007]. The transcriptional effects of EYA1 mutations are essentially unknown due to the difficulty of obtaining appropriate samples for EYA1 RNA analysis.…”

“…BOR syndrome has variable expressivity within and among families. EYA1 mutations have been found in 30-70% of patients with BOR, with the majority of mutations novel [Abdelhak et al, 1997a;Chang et al, 2004;Sanggaard et al, 2007;Orten et al, 2008].…”

Multiple evolutionarily conserved enhancers control expression of Eya1

“…It is possible that a novel mutation that affects the activity of enhancers identified in this study causes BOR or related symptoms. Indeed, one Danish family was recently identified to lack any coding/splice site mutations despite LOD score indicative of EYA1 involvement (Sanggaard et al, 2007).…”