Abstract
Purpose :
The mutations in the βB2-crystallin gene cause congenital cataract, but the mechanisms are not fully understood. We investigated the molecular mechanisms of cataract formation using gene knock-in mice expressing the tryptophan 151 to cysteine mutation (W151C) in βB2-crystallin (βB2-W151C), which is linked with autosomal dominant hereditary cataract in a four-generation Chinese family.
Methods :
Wild-type (WT), heterozygote (βB2-W151C +/−), and homozygote (βB2-W151C +/+) mouse lenses were assessed for lens morphology with slit-lamp biomicroscopy, dissecting microscope, and histological analysis. To investigate the mechanisms of the development of congenital cataract, the whole genome transcriptomics of WT and homozygous mice at 3 months old were performed. The transcript levels of representative endoplasmic reticulum stress (ERS) related genes were validated using real-time PCR. The activation of unfolded protein response (UPR) involves the expression of PERK, p-PERK, ATF4, IRE1, eif2α, and p-eif2α were determined by western blot.
Results :
βB2-W151C +/+ homozygous mice developed punctate lens opacities at the peripheral cortex at 18 days of age, while βB2-W151C +/− heterozygotes develop opacities at 21 days old. Punctate peripheral lens opacities extended gradually and became denser with aging. The whole lens became opaque by 2.5 months of age in homozygous mice, and 3 months in heterozygous mice. Lens histology for heterozygous and homozygous mice at 3 months old displayed abnormalities, the fiber cell migration and elongation pattern is found to be defective and is distinctly different from that observed in WT lenses. Microarray analysis revealed that 12 ERS-related genes were significantly up-regulated in homozygous mice. The validation results from real-time PCR also demonstrated ERS-related genes were dramatically increased in W151C +/+ and W151C +/− mice, including Trib3, chac1, ATF3, ATF5, ATF4, CHOP, Ddit3, Hspb8, Hsph1, Dnajc15, and Cebpb. Furthermore, W151C +/+ and W151C +/− mice showed increased expression for ATF4, p-PERK, p-eif2α, and IRE1 in the lens, indicating the activation of UPR.
Conclusions :
The W151C βB2-crystallin mutation develops progressive bilateral congenital cataract after birth. Mutant βB2-crystallin induced-ERS plays a vital role in the progression of congenital cataract, and inhibition of ERS may be a target for treatment in human patients.
This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.