Protein 4.1 in forebrain postsynaptic density preparations - Enrichment of4.1 gene products and detection of 4.1R binding proteins

4.1 Proteins are a family of multifunctional cytoskeletal components (4.1R, 4.1G, 4.1N and 4.1B) derived from four related genes, each of which is exp ressed in the nervous system. Using subcellular fractionation, we have inve stigated the possibility that 4.1 proteins are components of forebrain post synaptic densities, cellular compartments enriched in spectrin and actin, w hose interaction is regulated by 4.1R. Antibodies to each of 4.1R, 4.1G, 4. 1N and 4.1B recognize polypeptides in postsynaptic density preparations. Of these, an 80-kDa 4.1R polypeptide is enriched 11-fold in postsynaptic dens ity preparations relative to brain homogenate. Polypeptides of 150 and 125 kDa represent 4.1B; of these, only the 125 kDa species is enriched (threefo ld). Antibodies to 4.1N recognize polypeptides of approximate to 115, 100, 90 and 65 kDa, each enriched in postsynaptic density preparations relative to brain homogenate. Minor 225 and 200 kDa polypeptides are recognized sele ctively by specific anti-4.1G antibodies: the 200 kDa species is enriched 2 .5-fold. These data indicate that specific isoforms of all four 4.1 protein s are components of postsynaptic densities. Blot overlay analyses indicate that, in addition to spectrin and actin, postsynaptic density polypeptides of 140. 115, 72 and 66 kDa are likely to be 4.1R-interactive. Of these, 72 kDa and 66 kDa polypeptides were identified as neurofilament L, and alpha - internexin, respectively. A complex containing 80 kDa 4.1R, alpha -internex in and neurofilament L was immunoprecipitated with anti-4.1R antibodies fro m brain extract. We conclude that 4.1R interacts with the characteristic in termediate filament proteins of postsynaptic densities, and that the 4.1 pr oteins have the potential to mediate the interactions of diverse components of postsynaptic densities.