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An alphaA-crystallin gene mutation, Arg12Cys, causing inherited cataract-microcornea exhibits an altered heat-shock response.

Zhang LY, Yam GH, Tam PO, Lai RY, Lam DS, Pang CP, Fan DS - Mol. Vis. (2009)

Bottom Line: When expressed in COS-7 cells, the R12C mutant CRYAA resembled the wild type protein in its solubility when extracted with 0.5% Triton X-100 and with its cytoplasmic localization.However, mutant cells exhibited an altered heat-shock response, evidenced by the delayed expression of HSP70, when compared to cells expressing wild type CRYAA.The altered heat-shock response of mutant cells suggested a change of chaperoning capacity and networking, which could be associated with the pathogenesis of hereditary cataract-microcornea syndrome.

View Article: PubMed Central - PubMed

Affiliation: Department of Ophthalmology and Visual Sciences, The Chinese University of Hong Kong, Hong Kong, China.

ABSTRACT

Purpose: To investigate the clinical features and molecular basis of inherited cataract-microcornea caused by an alphaA-crystallin gene (CRYAA) mutation in a Chinese family.

Methods: A three-generation Chinese family with members having autosomal dominant cataract and microcornea was recruited. Genomic DNA from peripheral blood or buccal swab samples of five affected and five unaffected members were obtained. Based on 15 genes known to cause autosomal dominant cataract, single nucleotide polymorphisms (SNPs) or microsatellite markers were selected and genotyped for two-point linkage analysis. Direct sequencing was performed to identify the disease-causing mutation. The expression construct coding for recombinant COOH-terminal myc-His-tagged wild type or R12C alphaA-crystallin protein (CRYAA) was expressed in COS-7 cells. Detergent solubility and subcellular distribution of wild type and R12C CRYAA were examined by western blotting and immunofluorescence, respectively. Heat-shock response was monitored by quantitative polymerase chain reaction (qPCR) of heat-shock proteins 70 and 90alpha (HSP70 and HSP90alpha).

Results: The five affected family members showed variable lens opacities and microcornea. Clinical features of cataract were asymmetric in two eyes of some affected subjects. A heterozygous missense substitution, c.34C>T, in CRYAA, which is responsible for the R12C amino acid change, segregated with autosomal dominant cataract (ADCC) in this family. This substitution was absent in 103 unrelated controls. When expressed in COS-7 cells, the R12C mutant CRYAA resembled the wild type protein in its solubility when extracted with 0.5% Triton X-100 and with its cytoplasmic localization. However, mutant cells exhibited an altered heat-shock response, evidenced by the delayed expression of HSP70, when compared to cells expressing wild type CRYAA.

Conclusions: The R12C mutation in CRYAA was responsible for a variable type of inherited cataract associated with microcornea in this Chinese family. The altered heat-shock response of mutant cells suggested a change of chaperoning capacity and networking, which could be associated with the pathogenesis of hereditary cataract-microcornea syndrome.

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Related in: MedlinePlus

A Chinese autosomal dominant cataract family. A: Pedigree of a three-generation Chinese family with autosomal dominant cataract is shown. The asterisk indicates the family members whose DNA samples were analyzed. B: The lens image was taken from subject III5 following pupillary dilation. Both lenses showed the lamellar opacity within the fetal nucleus with a clear peripheral cortex. C: Subject II3 displayed similar lamellar opacity in the right lens and a central dense opacity with sharp margins in the left lens. D: Both images were from subject II5. A single spherical opacity (indicated by black arrows) was evident in the left lens, and only a smaller dot was found in the right lens.
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f1: A Chinese autosomal dominant cataract family. A: Pedigree of a three-generation Chinese family with autosomal dominant cataract is shown. The asterisk indicates the family members whose DNA samples were analyzed. B: The lens image was taken from subject III5 following pupillary dilation. Both lenses showed the lamellar opacity within the fetal nucleus with a clear peripheral cortex. C: Subject II3 displayed similar lamellar opacity in the right lens and a central dense opacity with sharp margins in the left lens. D: Both images were from subject II5. A single spherical opacity (indicated by black arrows) was evident in the left lens, and only a smaller dot was found in the right lens.

Mentions: A three-generation Chinese family with inherited cataract (Figure 1A) was recruited to the University Eye Center of The Chinese University of Hong Kong (Hong Kong, China). The research protocol followed the tenets of the Declaration of Helsinki and was approved by the ethics committee of The Chinese University of Hong Kong. Ten family members underwent complete ophthalmic examination including slit lamp biomicroscopy and ophthalmoscopy. All affected individuals (I1, II3, II5, II9, and III5) were followed up for eight years until 2008 by the same senior ophthalmologist in our eye center. During their last visit, additional ophthalmic examinations were performed including corneal diameter measurement, keratometry (Autokerato-refractometer KR-7100; Topcon Optical, Tokyo, Japan), anterior segment optical coherence tomography (1310 nm, Visante OCT Model 1000; Carl Zeiss Meditec, Dublin, CA), and ultrasonography (Ultrasonic Pachymeter SP-3000 and Biometer AL-100; Tomey Corporation, Nagoya, Japan). Slit lamp photography was also performed. Microcornea was defined as the horizontal corneal diameter being shorter than 11 mm [20,21] and microphthalmia as the antero-posterior axial length of the adult (normally elder than three) eye globe being shorter than 20 mm [22,23].


An alphaA-crystallin gene mutation, Arg12Cys, causing inherited cataract-microcornea exhibits an altered heat-shock response.

Zhang LY, Yam GH, Tam PO, Lai RY, Lam DS, Pang CP, Fan DS - Mol. Vis. (2009)

A Chinese autosomal dominant cataract family. A: Pedigree of a three-generation Chinese family with autosomal dominant cataract is shown. The asterisk indicates the family members whose DNA samples were analyzed. B: The lens image was taken from subject III5 following pupillary dilation. Both lenses showed the lamellar opacity within the fetal nucleus with a clear peripheral cortex. C: Subject II3 displayed similar lamellar opacity in the right lens and a central dense opacity with sharp margins in the left lens. D: Both images were from subject II5. A single spherical opacity (indicated by black arrows) was evident in the left lens, and only a smaller dot was found in the right lens.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2690964&req=5

f1: A Chinese autosomal dominant cataract family. A: Pedigree of a three-generation Chinese family with autosomal dominant cataract is shown. The asterisk indicates the family members whose DNA samples were analyzed. B: The lens image was taken from subject III5 following pupillary dilation. Both lenses showed the lamellar opacity within the fetal nucleus with a clear peripheral cortex. C: Subject II3 displayed similar lamellar opacity in the right lens and a central dense opacity with sharp margins in the left lens. D: Both images were from subject II5. A single spherical opacity (indicated by black arrows) was evident in the left lens, and only a smaller dot was found in the right lens.
Mentions: A three-generation Chinese family with inherited cataract (Figure 1A) was recruited to the University Eye Center of The Chinese University of Hong Kong (Hong Kong, China). The research protocol followed the tenets of the Declaration of Helsinki and was approved by the ethics committee of The Chinese University of Hong Kong. Ten family members underwent complete ophthalmic examination including slit lamp biomicroscopy and ophthalmoscopy. All affected individuals (I1, II3, II5, II9, and III5) were followed up for eight years until 2008 by the same senior ophthalmologist in our eye center. During their last visit, additional ophthalmic examinations were performed including corneal diameter measurement, keratometry (Autokerato-refractometer KR-7100; Topcon Optical, Tokyo, Japan), anterior segment optical coherence tomography (1310 nm, Visante OCT Model 1000; Carl Zeiss Meditec, Dublin, CA), and ultrasonography (Ultrasonic Pachymeter SP-3000 and Biometer AL-100; Tomey Corporation, Nagoya, Japan). Slit lamp photography was also performed. Microcornea was defined as the horizontal corneal diameter being shorter than 11 mm [20,21] and microphthalmia as the antero-posterior axial length of the adult (normally elder than three) eye globe being shorter than 20 mm [22,23].

Bottom Line: When expressed in COS-7 cells, the R12C mutant CRYAA resembled the wild type protein in its solubility when extracted with 0.5% Triton X-100 and with its cytoplasmic localization.However, mutant cells exhibited an altered heat-shock response, evidenced by the delayed expression of HSP70, when compared to cells expressing wild type CRYAA.The altered heat-shock response of mutant cells suggested a change of chaperoning capacity and networking, which could be associated with the pathogenesis of hereditary cataract-microcornea syndrome.

View Article: PubMed Central - PubMed

Affiliation: Department of Ophthalmology and Visual Sciences, The Chinese University of Hong Kong, Hong Kong, China.

ABSTRACT

Purpose: To investigate the clinical features and molecular basis of inherited cataract-microcornea caused by an alphaA-crystallin gene (CRYAA) mutation in a Chinese family.

Methods: A three-generation Chinese family with members having autosomal dominant cataract and microcornea was recruited. Genomic DNA from peripheral blood or buccal swab samples of five affected and five unaffected members were obtained. Based on 15 genes known to cause autosomal dominant cataract, single nucleotide polymorphisms (SNPs) or microsatellite markers were selected and genotyped for two-point linkage analysis. Direct sequencing was performed to identify the disease-causing mutation. The expression construct coding for recombinant COOH-terminal myc-His-tagged wild type or R12C alphaA-crystallin protein (CRYAA) was expressed in COS-7 cells. Detergent solubility and subcellular distribution of wild type and R12C CRYAA were examined by western blotting and immunofluorescence, respectively. Heat-shock response was monitored by quantitative polymerase chain reaction (qPCR) of heat-shock proteins 70 and 90alpha (HSP70 and HSP90alpha).

Results: The five affected family members showed variable lens opacities and microcornea. Clinical features of cataract were asymmetric in two eyes of some affected subjects. A heterozygous missense substitution, c.34C>T, in CRYAA, which is responsible for the R12C amino acid change, segregated with autosomal dominant cataract (ADCC) in this family. This substitution was absent in 103 unrelated controls. When expressed in COS-7 cells, the R12C mutant CRYAA resembled the wild type protein in its solubility when extracted with 0.5% Triton X-100 and with its cytoplasmic localization. However, mutant cells exhibited an altered heat-shock response, evidenced by the delayed expression of HSP70, when compared to cells expressing wild type CRYAA.

Conclusions: The R12C mutation in CRYAA was responsible for a variable type of inherited cataract associated with microcornea in this Chinese family. The altered heat-shock response of mutant cells suggested a change of chaperoning capacity and networking, which could be associated with the pathogenesis of hereditary cataract-microcornea syndrome.

Show MeSH
Related in: MedlinePlus