Abstract
Purpose: :
Establishment and maintenance of lens cytoarchitecture involves a variety of cell-cell junctions. We now have identified a novel N-cadherin junction which becomes linked to the vimentin intermediate filament cytoskeleton through γ-catenin and desmoplakin in differentiating lens fiber cells.
Methods: :
Co-precipitation analysis was used to determine the composition of N-cadherin junctions in microdissected E10 chick embryo lenses. A new double IP technique was developed to separate N-cadherin/γ-catenin from N-cadherin/β-catenin junctions. Immunofluorescence of E10 chick lenses was used to determine the localization of N-cadherin/γ-catenin and N-cadherin/β-catenin junctions.
Results: :
We have previously shown that proper morphogenesis of the lens is dependent upon the formation of mature N-cadherin junctions. These classical cadherin junctions can be stabilized through interactions with the actin cytoskeleton, thought to be mediated by both β- and γ-catenin. Unlike β-catenin, γ-catenin can also bind to desmosomal cadherins, linking them to intermediate filaments through desmoplakin. We now present the first biochemical evidence that, in vivo, γ-catenin also mediates interactions between a classical cadherin and the intermediate filament cytoskeleton, linked through desmoplakin. In the developing lens, which has no desmosomes, we discovered that vimentin became linked to N-cadherin complexes in a differentiation-state specific manner. This newly identified junctional complex was tissue specific but not unique to the lens. Our novel double IP technique made possible, for the first time, the separation of N-cadherin/γ-catenin from N-cadherin/β-catenin complexes and the identification of multiple members of each of these protein complexes. This study revealed that vimentin was associated exclusively with N-cadherin/γ-catenin junctions. Immunolocalization studies confirmed these biochemical results. β-catenin localized along lateral cell-cell interfaces of the hexagonally-packed lens fiber cells, co-localizing with F-actin. Both β-catenin and F-actin were absent from the cell vertices, which is the site of localization of the N-cadherin/γ-catenin junction. Formation of this novel junction occurred coincident with the establishment of the unique cytoarchitecture of lens fiber cells.
Conclusions: :
We believe this novel vimentin-linked N-cadherin/γ-catenin junction is important to the establishment of the unique cytoarchitecture of the lens and provides the tensile strength necessary to maintain structural integrity of the lens.
Keywords: cell adhesions/cell junctions • cytoskeleton • development