Control of lens fiber cell denucleation by CDK1 regulation in mouse and chick

Abstract

Purpose : The lens of the eye undergoes a regulated differentiation process whereby organelle removal in the fiber cells is necessary for lens clarity. A critical step in this process is the removal of the fiber cell nucleus. The mechanisms by which this occurs are unclear. Previous work in our lab demonstrated that impaired lens fiber cell denucleation (LFCD) was associated with increased levels of the CDK inhibitor p27. Additionally, deletion of CDK1 in the lens impaired LFCD indicating that CDK activity is crucial for LFCD. In this study, we test CDK activators and inhibitors in the denucleation process.

Methods : We examined lens differentiation in the mouse and embryonic chick by histology and immunofluorescence. We used three different mouse models to ask whether increased p27 levels affects lens differentiation. The first model expresses a drug inducible p27 transgene. We induced expression of the transgene during pregnancy to increase p27 during a crucial phase of lens differentiation. The second model is a Skp2-/-. Skp2 is part of a ubiquitin ligase complex that is known to ubiquitinate p27 and target it for proteasomal degradation; these mice constitutively express high levels of p27. The third model expresses a degradation resistant mutant p27. To determine how CDK1 regulators are involved in LFCD, we used an organ culture system and treated isolated chick lenses with small molecules that inhibit the activities of CDK1 and Cdc25, which activates CDK1. Additionally, we examined LFCD in lenses in which we inhibited CDK1 inhibitors, Wee1 and PP2A.

Results : Expression of increased levels of p27 led to impaired LFCD in all three mouse models. Inhibition of the CDK1 activator Cdc25 inhibits LFCD, whereas inhibition of the CDK1 inhibitors Wee1 and PP2A potentiate LFCD in chick lenses. PP2A inhibits CDK1 activity: activation of PP2A inhibits LFCD while inhibition of PP2A increases LFCD.

Conclusions :
Increasing p27 levels in lenses or inhibiting CDK1 in lenses by genetic and pharmacologic mechanisms results in impaired LFCD. CDK1 activity is a crucial regulator of LFCD in the chick as well as the mouse.

This is a 2021 ARVO Annual Meeting abstract.