Abstract
Purpose :
The deletion G-91 in βA3 (βA3ΔG91) causes pediatric congenital cataract in several families worldwide. The purpose was to determine phenotypic characteristics of lenses in βA3ΔG91 mice to delineate molecular mechanism of pediatric congenital nuclear cataract development.
Methods :
We have generated βA3ΔG91 mouse model by the Crisper/cas9 methodology using UAB Transgenic & Genetically Engineered Models Core (TGEMs) facility. Lenses from 1-month-old βA3ΔG91- and wild-type mice were examined for cataract development using Micron V slit-lamp analysis. The insolubilization of the lens proteins were examined following isolation of water soluble (WS) - and water insoluble (WI) - proteins and their analysis by SDS-PAGE. The protein structure and function prediction were done using iTASSER (Interactive Threading ASSEmbly Refinement) in both wild-type and mutant protein. The amyloidogenic regions was analyzed using FoldAmyloid analysis software.
Results :
Relative to wild-type lenses, the lenses of 1-month-old βA3ΔG91 mice showed polar nuclear cataract, which became progressively more pronounced at 2-months of age. A two-fold greater insolubilization of proteins in 1- and 2-month-old βA3ΔG91 lenses relative to the age-matched wild-type lenses was observed. βA3ΔG91 protein also exhibited eight amyloidogenic sites compared seven amyloidogenic sites in native βA3-crystallin. The additional amyloidogenic sites in βA3ΔG91 had QFILER sequence comprising of G91 deletion site, and structural analysis (iTASSER) of the mutated βA3ΔG91 protein showed random structure compared to an organized structure of native βA3.
Conclusions :
The findings suggest that on G-91 deletion causes an altered structure in βA3ΔG91 protein relative to native βA3-crystallin, and the development of an additional amyloidogenic region comprising G91-deleted region, which might be responsible for its relatively greater insolubilization and pediatric congenital cataract development.
This is a 2020 ARVO Annual Meeting abstract.