Immunolocalization of the 110,000 molecular weight cytoskeletal protein of intestinal microvilli |
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Authors: | Evelyne Coudrier Hubert Reggio Daniel Louvard |
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Affiliation: | European Molecular Biology Laboratory Postfach 10.2209 6900 Heidelberg, Federal Republic of Germany |
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Abstract: | The core structures of microvilli from absorptive cells of the intestinal epithelium are primarily composed of calmodulin (Mr 16,000), actin (Mr 43,000), villin (Mr 95,000) and a protein of Mr 110,000. We have isolated this protein and raised antibodies against it. The antibodies interact specifically with villin and Mr 110,000 polypeptides present in isolated microvilli or brush borders. However, after absorption on an immobilized villin preparation, these antibodies still immunoprecipitate the Mr 110,000 protein but not villin. Thus, these two proteins appear to share some antigenic determinants but also contain other determinants specific for each protein. Immunolocalization studies have been performed using specific antibodies against the Mr 110,000 protein. Immunofluorescent studies on thin frozen sections of intestinal cells show that this protein is located in the brush border and at the basolateral faces of these polarized cells. Immunoferritin studies on rat brush borders demembranated with the detergent Triton X-100 show the association of the Mr 110,000 protein with core filaments of microvilli, as well as with some filaments localized in the terminal web network.Using sealed, right-side-out vesicles prepared from pig intestinal mucosa in the presence of Ca2+ and Mg2+, a polypeptide of Mr 140,000 was found to be a major component of the Triton X-100 insoluble pellet. This protein is a minor component of an equivalent pellet obtained from isolated microvilli prepared in the presence of EDTA. The significance of this Mr 140,000 polypeptide associated with the core residue of intestinal microvilli is discussed. |
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