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Fig. 5. Regulation of E-cadherin-catenin complex by Src42A. (A)
Complex formation by E-cadherin, Arm and Src42A. Expression constructs of
E-cadherin-GFP and Src42A were transfected into S2 cells and the
E-cadherin-Arm complex was analyzed by immunoprecipitation. Blue and red boxes
mark the lanes of whole cell extract and immunoprecipitates of anti-Arm,
respectively. The expression of E-cadherin-GFP stabilized Arm in apparent high
and low molecular weight forms, indicated by H and L. Although both
Src42AACT and Src42ADN in whole cell extract were
phosphorylated at Y400, Src42ADN in complex with Arm was under
phosphorylated. (B) Src42A downregulates the cell surface level of
E-cadherin. Surface biotinylation was applied to S2 cells expressing
E-cadherin-GFP and Src42A. Western blot analyses were performed on whole cell
extracts (whole) and proteins were recovered by avidin beads (surface). Septin
and avidin-reacting proteins were used as loading controls for each blot. The
positions of marker proteins are shown in the avidin panel. (C)
Downregulation of E-cadherin-GFP by Src42AACT. Transfected S2 cells
were stained with a method to detect extracellular E-cadherin (surface,
magenta). GFP signal in the intracellular domain (intra, green) is shown.
Src42AACT reduced both signals. (D) The amount of E-cadherin
was reduced in Src mutant embryos. Control and Src42Amyr;
Src64BP1 embryos (src-/-) were collected
and analyzed for E-cadherin and pSrc expression by western blotting. Protein
extracts were analyzed in the same gels and blots. Relative amount is shown
above each band.
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