Cytoskeletal Complexes Important for Lens Function

Abstract

Abstract: : Purpose: To investigate the potential associations between the actin and intermediate filament networks in the lens Methods: Bovine lenses were fractionated by a combination of differential centrifugtion and extraction protocols to enrich for plasma membrane and cytoskeletal fractions in the lens. These fractions were then subjected to one and two dimensional gel electrophoresis. Major spots were then identified by a combination of MALDI and tandem mass spectrometry and confirmed by immunoblotting as approaches to characterise the protein complexes in each of these fractions. Where possible confocal immunofluorescence light microscopy and immunoelectron microscopy of lens sections were undertaken. Results: The major cytoskeletal fraction of the lens, the PMCC fraction, contained several high molecular weight proteins known to associate with intermediate filaments and actin, namely plectin and filamin. Both proteins are expected to cross–link their associated filament systems without necessarily bundling filaments, as well as providing vital links between both the actin and intermediate filament networks. Both filamin and plectin were also found in the plasma membrane fractions in association with spectrin, FERM and LIM–domain containing proteins. Conclusions: The data indicate the presence of a complex mat of intermediate filaments and actin filaments at the plasma membrane of the lens fibre cells. The enrichment of actin associated proteins involved in signalling and membrane organisation with these intermediate filament enriched fractions give important insight into the role that the cytoskeleton as a whole plays in plasma membrane compartmentalisation and organisation

Keywords: cytoskeleton