title: Localization of ras antigenicity in rat hepatocyte plasma membrane and rough endoplasmic reticulum fractions.
abstract: We have examined the antigenicity of plasma membrane (PM) and rough microsomal (RM) fractions from rat liver using anti-ras monoclonal antibodies 142-24EO5 and Y13-259 and immunochemistry as well as electron microscope immunocytochemistry. Proteins immunoprecipitated with monoclonal antibody 142-24E05 were separated using single-dimensional gradient-gel electrophoresis. The separated proteins were then blotted onto nitrocellulose sheets and incubated with [alpha-32P]GTP. Radioautograms of blots indicated the presence of specific 21.5- and 22-kDa labeled proteins in the PM fraction. A 23.5-kDa [alpha-32P] GTP-binding protein was detected in immunoprecipitates of both PM and RM fractions. Monoclonal antibody Y13-259 reacted only with the 21.5-kDa [alpha-32P] GTP-binding protein in the plasma membrane fraction. When anti-ras monoclonal antibody 142-24E05 and the immunogold technique were applied to membrane fractions using a preembedding immunocytochemical method, specific labeling was observed in association with both vesicular structures and membrane sheets in the PM fraction but only with electron-dense vesicular structures in the RM fraction. Thus ras antigenicity is associated with hepatocyte plasma membranes and ras-like antigenicity is probably associated with vesicular (secretory/endocytic) elements in both plasma membrane and rough microsomal preparations.
authors.affiliation: Département d'anatomie, Faculté de Médecine, Université de Montréal, Québec, Canada.
authors.forename: J M
authors.initials: JM
authors.lastname: Dominguez
authors.forename: J
authors.initials: J
authors.lastname: Lanoix
authors.forename: J
authors.initials: J
authors.lastname: Paiement
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal; D002462:Cell Membrane / Q000737:chemistry*; D003429:Cross Reactions; D004721:Endoplasmic Reticulum / Q000737:chemistry*; D019204:GTP-Binding Proteins / Q000276:immunology; D007150:Immunohistochemistry; D008099:Liver / Q000737:chemistry*; D008970:Molecular Weight; D015689:Oncogene Protein p21(ras) / Q000032:analysis*; D010761:Phosphorus Radioisotopes; D011233:Precipitin Tests; D051381:Rats; D013347:Subcellular Fractions / Q000737:chemistry
chemical_list: D000911:Antibodies, Monoclonal; D010761:Phosphorus Radioisotopes; D019204:GTP-Binding Proteins; D015689:Oncogene Protein p21(ras)