Bernasconi, Andrea

F.Schorderet, Daniel

Pescia, Graziano

Regli, Franco

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©1994 Oxford University Press Human Molecular Genetics, 1994, Vol. 3, No. 7 1201An additional family with Startle disease and a G1192A mutation at thesubunit of the inhibitory glycine receptor geneDaniel F.Schorderef, Graziano Pescia, Andrea Bernasconi1 and Franco Regll1Division of Medical Genetics and 1Department of Neurology, CHUV, 1011 Lausanne, SwitzerlandReceived April 8, 1994; Revised and Accepted May 3, 1994Recently, Shiang and colleagues have identified the generesponsible for Startle disease (1). This disease also known ashereditary hyperplexia or Kok disease is characterized by markedbut transient infantile hypertonia and exaggerated startle response(2). Sequence analysis of the a\ subunit of the inhibitory glycinereceptor (GLRA1) showed a mutation at the same codon 271in four out of seven tested families (Gl 192A and Gl 192T) (1).We have analyzed this gene in a family of Swiss origin and foundthe Gl 192A mutation changing an ARG to a LEU codon. Thisreport confirms that the GLRA1 gene is responsible for Startledisease and describes a simple digestion-based assay for thedetection of codon 271 mutation.The proband, a 17-year old girl of Swiss origin, was referredto our Hospital for diagnosis and treatment of numerous episodesof falling and myclonia in response to stimuli. She is the seconddaughter of non-consanguineous parents. Her older sister andmother both suffer abnormal startle reflex. The mutationsdescribed by Shiang etal. (1) at codon 271 involve a TaqI (andXhol) site which allows for their rapid detection by restrictionenzyme digestion. In the normal sequence, the TaqI site TCGAis present and complete digestion of PCR-amplified genomicDNA will produce two fragments of equal size. If, however, amutation is introduced, the mutated site will not be cut and 2bands will be seen, one from the normal and one from the mutatedchromosome. In order to test for these mutations, DNA wasextracted from peripheral lymphocytes and exon 6 of GLRA1was amplified using the described primers without the GC-clamp(1). Amplicons were digested with TaqI at 65°C in therecommended buffer, fragments were separated on a 7%poly aery lamide mini-gel for 60 min. at 60 W and the gel wasstained with ethidium bromide.Figure 1 shows the TaqI digestion of the PCR products fromthe reported family. Affected individuals reveal 2 bands whileonly one is present in the unaffected father. Because destructionof the TaqI site could be due to mutation in any of the 4 basesdefining the recognition site of the enzyme, amplicons from theproband were subcloned in pUC 18 and one clone showing lossof the TaqI site was sequenced on an A.L.F. automated sequencerto determine the exact mutation. The mutation found was a Gto A transition at position 1192.Thus, 50% of the families analyzed at a molecular level presenta Gl 192A mutation. This may arise from a common ancestoror from the presence of a mutational hot spot at that particularlocation. CpG dinucleotides are frequently involved in pointmutations probably as the result of methylation-deamination.Deamination of a 5'-methylated CpG dinucleotide in the non-sense strand would create a CpA at the sense strand and couldthus be responsible for 3 out of 4 mutations at codon 271.However, the fact that another point mutation has occurred atthe same position would also suggest that Arginine at position271 is critical for the function of the inhibitory glycine receptorgene.ACKNOWLEDGEMENTSWe are very grateful to Sylvie Luthi and Farid Ahmad for technical assistance.REFERENCES1. Shiang.R., Ryan.S.C, Zhu.Y., Hahn.A.F., O'Connell.P. and WasmuthJ.J.(1993) Nature Genet. 5, 351-357.2. Kok.O. and Bruyn.G.W. (1962) Lancet 1, 1359.i iN C N C N C N CFigure 1. Agarose gel of TaqI digested and control PCR-amplified DNA stainedwith ethidium bromide. N: not cut; C: TaqI cut. M5 is a DNA molecular weightmarker (marker V from Boehringer Mannheim GmbH, lowest band on the gelis 184 tip).• To whom correspondence should be addressed

An additional family with Startle disease and a G1192A mutation at the α1 subunit of the inhibitory glycine receptor gene

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