Phorbol mynstate acetate (PMA) induced the scattering and morphological changes of HepG2 human hepatoma cells Studies concerning the expression, down-regulation and selective inhibition of the individual protein kinase C (PKC> tsoforms suggested that Ca-independent subtypes, especially PKC-e were involved in the PMA-induced effects Electron microscopic analysis based on preembeddmg tmmunocytocheniistry demonstrated the presence of E-cadherin in the plasma membranes of neighbouring cells. The monoclonal antibody applied for the detection of adherent junctions was developed against tlie extracellular part of E-cadlierm When this antibody was used for Western blotting it recognized a 120 kDa protein found exclusively in the cytoskeletal fraction In PMA-treated cells the anti-E-cadherin imnumoreactive structures disappeared from the plasma membranes However, at the same time, the amount of the 120 kDa protein was not reduced in the cytoskeletal fraction We investigated the subcellular distribution of E-cadlierin with a second monoclonal antibody, as well This antibody was developed against the intracellular part of E-cadherin The dominant protein recognized by the second antibody was slightly larger than thai recognized by the first one and was distributed between the cytoskeletal and TntonXlOO extractable fractions of HepG2 cells With the aid of this antibody we observed a reduction of the amount of E-cadhenn in (he TntonXlOO extractable fraction of PMA-treated cells however, in the cytoskeletal fraction the amount of E-cadherin was not decreased. Oui data suggest that in HepG2 cells three pools of E-cadhenn exist The disappearance of E-cadhenn from the plasma membrane upon the permanent activation of PK.C is probably mediated by the rearrangement of the actm-cytoskeleton.
|Publication status||Published - Dec 1 1997|
ASJC Scopus subject areas
- Molecular Biology