Synthesis of S100 Protein on Free and Membrane-Bound Polysomes of the Rabbit Brain

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Abstract
Free and membrane-bound polysomes were isolated from the cerebral hemispheres and cerebellum of the young adult rabbit. The 2 polysomal populations were translated in an mRNA-dependent cell-free system derived from rabbit reticulocytes. Analysis of the [35S]methionine-labeled translation products on 2-dimensional polyacrylamide gels indicated an efficient separation of the 2 classes of brain polysomes. The relative synthesis of S100 protein by free and membrane-bound polysomes was determined by direct immunoprecipitation of the cell-free translation products in the presence of detergents to reduce nonspecific trapping. Synthesis of S100 protein was found to be 2-fold greater on membrane-bound polysomes compared with free polysomes isolated from either the cerebral hemispheres or the cerebellum. The proportion of poly(A+)mRNA coding for S100 protein was also 2-fold greater in membrane-bound polysomes compared with free polysomes isolated from the cerebral hemispheres. The cytoplasmic S100 protein is probably synthesized predominantly on membrane-bound polysomes in the rabbit brain. The nascent S100 polypeptide chain translation complex is probably attached to the rough endoplasmic reticulum by an ionic interaction involving a sequence of 13 basic aminoacids in S100 protein.