Effects of arecoline on expression of ZO-1 and HER2 protein in Ishikawa cells. (A) Subconfluent cultures of Ishikawa cells were treated with DMSO (vehicle control), 0.1 mM, 0.3 mM or 0.5 mM arecoline for 24 and 48 hrs, and total cell extracts were fractionated in SDS polyacrylamide gels. The arecoline regulation of ZO-1 and HER2 protein production was determined by western blot analysis and compared to the levels of actin protein; (B) To determine if the arecoline mediated downregulation of ZO-1 and HER2 protein was due to induced ubiquitination and 26 S proteasome mediated degradation, Ishikawa cells were treated with or without 0.3 mM arecoline for 48 hrs and in the presence or absence of MG132, an inhibitor of proteasome peptidase enzymatic activity. Total cell extracts were analyzed by Western blotting for ZO-1 and HER2 in comparison to actin.
Giri et al. BMC Cell Biology 2010 11:53 doi:10.1186/1471-2121-11-53