Abstract
Purpose: :
Differentiation of lens cells involves the sequential expression of two different intermediate filament (IF) systems: vimentin in the lens epithelium and younger fiber cells, and the fiber cell-specific Beaded Filaments as fiber cells mature. Both IF systems show a shift in cellular distribution as differentiation progresses. Because IF function can be modulated by differential expression of IF "linker proteins" which target IFs to a range of cellular structures, we sought to identify candidate linkers that may govern IF regulation in lens.
Methods: :
The Cre-Lox approach was used to specifically delete plectin from lens cells. Wild Type and lens plectin-deficient mice were compared by immunofluorescence, co-immunoprecipitation, and both transmission and scanning electron microscopy.
Results: :
Microscopy revealed that the elimination of plectin from lens cells did not alter the subcellular distribution of the Beaded Filament. In contrast, the loss of plectin was sufficient to sever the linkage of vimentin to the fiber cell plasma membrane, producing a dramatic shift in subcellular localization. Co-immunprecipitation studies showed a similar shift in vimentin from the more insoluble fraction to the more soluble fraction of sequentially extracted lens fiber cells.
Conclusions: :
Vimentin IFs show a strong localization to the basal compartment of the lens epithelial cells. However, as the cell initiates differentiation into a fiber cell, that localization shifts to a predominantly membrane associated form, suggesting a change in function. These studies establish that the IF linker protein plectin is required for tethering vimentin IFs, but not BFs, to the fiber cell plasma membrane.
Keywords: cytoskeleton • cell membrane/membrane specializations • differentiation