Abstract
Purpose :
Cyclase Associated Protein 2 (Cap2) is a highly conserved actin-binding protein that plays a critical role in regulating the assembly and disassembly of actin filaments. Cap2 facilitates the dissociation of actin monomers from the pointed ends of ADF/cofilin-bound actin filaments. The actin monomers can subsequently reassemble into actin filaments, contributing to the formation of diverse filamentous actin (F-actin) networks in conjunction with other actin-crosslinking proteins. Lens fiber cells contain a variety of F-actin networks that are dynamically rearranged during fiber cell differentiation and maturation and control of network dynamics is essential for maintaining normal lens biomechanical properties. To elucidate the function of Cap2 in regulating lens F-actin network organization and the impact on lens biomechanics, we conducted comprehensive analyses of the lens phenotype at both cellular and tissue levels in lens-specific Cap2-/- mice.
Methods :
The lens-specific Cap2-/- mouse model was created using Tg(Cryaa-cre)10Mlr mice. Western blotting, immunostaining, confocal microscopy, and mechanical testing were used to investigate the effect of Cap2 deficiency in mouse lenses.
Results :
Cap2-deficient lenses exhibit normal size, shape, and transparency. However, they are significantly stiffer under compression and demonstrated enhanced recovery upon load removal, in contrast to wild-type lenses. Immunostaining of equatorial sections of Cap2-deficient lenses revealed an increase in fiber cell disorder and misalignments. Notably, despite the overall levels of actin and the ratio of F-actin to globular (G)-actin remaining constant in the absence of Cap2, immunofluorescence staining of single fiber cells revealed an increase in the number of fluorescent puncta associated with the actin-crosslinking protein, plastin-3 (aka T-plastin, or fimbrin). These puncta were often colocalized with F-actin along the fiber cell membrane protrusions and paddle domains.
Conclusions :
Deletion of Cap2 in the lens significantly increases lens stiffness, which may be attributed to increased plastin-3 cross-linking of F-actin into compact bundles within mature lens fiber cells. This study extends our previous work showing that F-actin network composition influences lens stiffness, and presents the first evidence of Cap2's impact on cell biomechanical properties in non-muscle tissue.
This abstract was presented at the 2024 ARVO Annual Meeting, held in Seattle, WA, May 5-9, 2024.