An alphaA-crystallin gene mutation, Arg12Cys, causing inherited cataract-microcornea exhibits an altered heat-shock response.
OBJECTIVE
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.
Zhang
Li-Yun
LY
Department of Ophthalmology and Visual Sciences, The Chinese University of Hong Kong, Hong Kong, China.
Yam
Gary Hin-Fai
GH
Tam
Pancy Oi-Sin
PO
Lai
Ricky Yiu-Kwong
RY
Lam
Dennis Shun-Chiu
DS
Pang
Chi-Pui
CP
Fan
Dorothy Shu-Ping
DS
D000328:Adult; D000368:Aged; D019943:Amino Acid Substitution; D000818:Animals; D019556:COS Cells; D002386:Cataract / Q000235:genetics* / Q000378:metabolism; D002648:Child; D002681:China; D002522:Chlorocebus aethiops; D003315:Cornea / Q000002:abnormalities*; D003459:Crystallins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D005260:Female; D008040:Genetic Linkage; D006360:Heat-Shock Proteins / Q000378:metabolism*; D018869:Heat-Shock Response; D006801:Humans; D057927:Hydrophobic and Hydrophilic Interactions; D008297:Male; D008875:Middle Aged; D020125:Mutation, Missense; D010375:Pedigree; D012995:Solubility
C511246:CRYAA protein, human; D003459:Crystallins; D006360:Heat-Shock Proteins