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Functional characterization of a novel c.614-622del rhodopsin mutation in a French pedigree with retinitis pigmentosa.

Maubaret C, Kosmaoglou M, Low S, Chakarova CF, Bidot S, Thauvin-Robinet C, Robson AG, Waseem N, Cheetham ME, Bhattacharya SS - Mol. Vis. (2012)

Bottom Line: The deleted mutant protein localized to the endoplasmic reticulum and formed inclusion bodies.This novel deletion in exon 3 of the RHO gene, c.614-622del results in a classical form of adRP in a multi-generation French family.Protein expression analyses confirmed that the deletion led to protein misfolding and suggest this is a class II mutation, similar to P23H, the most common class II mutation seen in North America.

View Article: PubMed Central - PubMed

Affiliation: UCL Institute of Ophthalmology, London, United Kingdom. rmhacm1@ucl.ac.uk

ABSTRACT

Purpose: To identify and functionally characterize the mutation responsible for autosomal dominant retinitis pigmentosa (adRP) in a large, six-generation French family.

Methods: Twenty individuals from this family participated in the genetic investigation. Six affected and 14 unaffected individuals from three-generations were available for linkage analysis using microsatellite markers flanking the rhodopsin (RHO) gene. A two-point logarithm of odds (LOD) score calculation was undertaken using GENEMARKER and MLINK software. Sanger sequencing of RHO was performed. Cellular localization of the mutant protein was performed by transforming SK-N-SH cells with pEGFP-N1-Rho, pEGFP-N1-Rho(P23H), and pEGFP-N1-Rho(c.614-622del).

Results: The proband had nyctalopia, visual field constriction, peripheral bone spicule pigmentation of the fundus, central acuity (6/24 RE; 6/12 LE) at 55 years of age. Linkage analysis of this family suggested RHO as a possible candidate since the flanking marker D3S1292 yielded a LOD score of 2.43 at θ=0. Cloning of an exon 3 PCR product and direct sequencing of single clones identified a novel deletion in the third exon of RHO, c.614-622del (p.Y206-F208del). The deleted mutant protein localized to the endoplasmic reticulum and formed inclusion bodies.

Conclusions: This novel deletion in exon 3 of the RHO gene, c.614-622del results in a classical form of adRP in a multi-generation French family. Protein expression analyses confirmed that the deletion led to protein misfolding and suggest this is a class II mutation, similar to P23H, the most common class II mutation seen in North America.

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Rhodopsin mutation in exon 3. Detection of the novel c.614–622del (p.Y206-F208del) mutation in the RHO gene from a sequence analysis after cloning of both alleles. The electropherogram representing the normal sequence is shown on top while the deleted one is presented below.
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f3: Rhodopsin mutation in exon 3. Detection of the novel c.614–622del (p.Y206-F208del) mutation in the RHO gene from a sequence analysis after cloning of both alleles. The electropherogram representing the normal sequence is shown on top while the deleted one is presented below.

Mentions: Cloning of both PCR fragments (wild type and mutant) from individual 9 identified a novel mutation in the RHO gene: a nine base-pair deletion from nucleotide 614 to nucleotide 622 in exon three, c.614–622del (Figure 3). The in silico analysis of the mutant sequence predicts deletion of amino acid 206 to 208, which are tyrosine, methionine, and phenylalanine (p.Y206-F208del). The mutation co-segregated with the disease: it was identified in all six participants with a positive history or clinical signs of RP, but none of the unaffected individuals were carriers of this change.


Functional characterization of a novel c.614-622del rhodopsin mutation in a French pedigree with retinitis pigmentosa.

Maubaret C, Kosmaoglou M, Low S, Chakarova CF, Bidot S, Thauvin-Robinet C, Robson AG, Waseem N, Cheetham ME, Bhattacharya SS - Mol. Vis. (2012)

Rhodopsin mutation in exon 3. Detection of the novel c.614–622del (p.Y206-F208del) mutation in the RHO gene from a sequence analysis after cloning of both alleles. The electropherogram representing the normal sequence is shown on top while the deleted one is presented below.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3298422&req=5

f3: Rhodopsin mutation in exon 3. Detection of the novel c.614–622del (p.Y206-F208del) mutation in the RHO gene from a sequence analysis after cloning of both alleles. The electropherogram representing the normal sequence is shown on top while the deleted one is presented below.
Mentions: Cloning of both PCR fragments (wild type and mutant) from individual 9 identified a novel mutation in the RHO gene: a nine base-pair deletion from nucleotide 614 to nucleotide 622 in exon three, c.614–622del (Figure 3). The in silico analysis of the mutant sequence predicts deletion of amino acid 206 to 208, which are tyrosine, methionine, and phenylalanine (p.Y206-F208del). The mutation co-segregated with the disease: it was identified in all six participants with a positive history or clinical signs of RP, but none of the unaffected individuals were carriers of this change.

Bottom Line: The deleted mutant protein localized to the endoplasmic reticulum and formed inclusion bodies.This novel deletion in exon 3 of the RHO gene, c.614-622del results in a classical form of adRP in a multi-generation French family.Protein expression analyses confirmed that the deletion led to protein misfolding and suggest this is a class II mutation, similar to P23H, the most common class II mutation seen in North America.

View Article: PubMed Central - PubMed

Affiliation: UCL Institute of Ophthalmology, London, United Kingdom. rmhacm1@ucl.ac.uk

ABSTRACT

Purpose: To identify and functionally characterize the mutation responsible for autosomal dominant retinitis pigmentosa (adRP) in a large, six-generation French family.

Methods: Twenty individuals from this family participated in the genetic investigation. Six affected and 14 unaffected individuals from three-generations were available for linkage analysis using microsatellite markers flanking the rhodopsin (RHO) gene. A two-point logarithm of odds (LOD) score calculation was undertaken using GENEMARKER and MLINK software. Sanger sequencing of RHO was performed. Cellular localization of the mutant protein was performed by transforming SK-N-SH cells with pEGFP-N1-Rho, pEGFP-N1-Rho(P23H), and pEGFP-N1-Rho(c.614-622del).

Results: The proband had nyctalopia, visual field constriction, peripheral bone spicule pigmentation of the fundus, central acuity (6/24 RE; 6/12 LE) at 55 years of age. Linkage analysis of this family suggested RHO as a possible candidate since the flanking marker D3S1292 yielded a LOD score of 2.43 at θ=0. Cloning of an exon 3 PCR product and direct sequencing of single clones identified a novel deletion in the third exon of RHO, c.614-622del (p.Y206-F208del). The deleted mutant protein localized to the endoplasmic reticulum and formed inclusion bodies.

Conclusions: This novel deletion in exon 3 of the RHO gene, c.614-622del results in a classical form of adRP in a multi-generation French family. Protein expression analyses confirmed that the deletion led to protein misfolding and suggest this is a class II mutation, similar to P23H, the most common class II mutation seen in North America.

Show MeSH
Related in: MedlinePlus