Abstract
Purpose :
To determine the role of lens-specific βA3/A1-crystallin in autophagy regulation.
Methods :
Previously, we have studied βA3/A1-complete knockout (βA3KO) mouse model that showed autophagy disruption (PLOS one, 2016, e0149027). In the present study, a lens-specific homozygous βA3 conditional knockout (βA3cKO) mouse model was generated by crossing βA3-flox mice with MLR10 Cre mice. Congenital cataract development in the βA3cKO mice was examined by Micron-IV slit-lamp microscopy. Lens RNA isolated from both control (Cre-and flip mice) and βA3cKO mice, were used for transcriptome-and Ingenuity pathway analyses.
Results :
As observed previously in the βA3KO mice, βA3cKO mice also developed congenital nuclear cataract as confirmed by slit-lamp microscopy. The βA3cKO lenses exhibited attenuation of organelle degradation, and an age-related increase in the levels of water-insoluble (WI) proteins and decreased levels of water-soluble (WS) proteins compared to age-matched control lenses. Transcriptome analysis showed that several genes from the autophagy pathway were down-regulated in the βA3cKO mice compared to control mice. These included: autophagy-related 3 (ATG3), autophagy-related 12 (ATG12), ectopic P-granules 5 (EPG5), tectonin beta-propeller repeat containing 1 (TECPR1) and FYVE and coiled-coil domain-containing 1 (FYCO1). EPG5 and TECPR1 are tethering proteins involved in the autophagosome-lysosome fusion. Whereas FYCO1, in addition to its interaction with Rab7, also binds LC3 and PI(3)P, and its depletion leads to autophagosome accumulation at the cellular perinuclear region. Ingenuity pathway analysis showed that the affected top molecular and cellular functions in βA3cKO lenses were cellular movement, cell-to-cell signaling and interactions, cellular maintenance and cell death and survival.
Conclusions :
An absence of βA3-crystallin in the lenses of βA3cKO mice leads to development of congenital nuclear cataract, attenuation of organelle degradation, and a greater insolubilization of fragmented lens proteins, which included β-tubulin and αA-crystallin. Our study shows that the genes involved in tethering complex of autophagosomes-lysosome fusion are affected. Our long term goal is to determine how βA3/A1-crystallin affects lens autophagy.
This abstract was presented at the 2023 ARVO Annual Meeting, held in New Orleans, LA, April 23-27, 2023.