Abstract
Purpose :
The removal of organelles from lens fiber cells is critical for lens clarity. Multiple congenital cataract models show retention of nuclei in the purported organelle free zone, indicating that nuclear breakdown is among essential processes in establishing and maintaining lens clarity. The pathways that control nuclear envelope breakdown during lens fiber cell development are not well understood. Identifying the steps of nuclear envelope breakdown and controlling mechanisms in lens denucleation may provide insight into cataract development.
Methods :
Nuclear morphology and localization of nuclear membrane proteins were examined during differentiation in chick and mouse lenses using immunofluorescence. Embryonic chick lenses were cultured with inhibitors to test the role of cytoskeletal motor proteins in denucleation. Nuclear structure was examined in WT mouse lenses using electron microscopy.
Results :
In chick lenses, phosphorylation of Lamin A immediately precedes nuclear condensation. Additionally, after Lamin A phosphorylation, nuclear Lamin staining becomes discontinuous in the nuclear membrane. The nuclear pore complex also relocalizes in the nuclear membrane after Lamin phosphorylation and no longer localizes with Lamins. Nuclear envelope breakdown occurs in a stepwise manner with outer nuclear membrane proteins disappearing from the nuclear envelope before Lamin reorganization. These steps occur before DNA degradation. Furthermore, autophagy markers LC3 and p62 co-localize with the nuclear membrane, suggesting that autophagy is involved in nuclear envelope breakdown. Inhibitor studies suggest that the cytoskeletal motor dynein is involved in nuclear envelope reorganization and the denucleation process.
Conclusions :
Our data indicate that phosphorylation of Lamin triggers nuclear envelope reorganization. Additionally, cytoskeletal motor proteins are involved in nuclear envelope breakdown during lens fiber cell denucleation.
This abstract was presented at the 2024 ARVO Annual Meeting, held in Seattle, WA, May 5-9, 2024.