Actinomycin D as a novel SH2 domain ligand inhibits Shc/Grb2 interaction in B104‐1‐1 (neu*‐transformed NIH3T3) and SAA (hEGFR‐overexpressed NIH3T3) cells
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Open Access
- 17 June 1999
- journal article
- Published by Wiley in FEBS Letters
- Vol. 453 (1-2) , 174-178
Abstract
Actinomycins, a family of bicyclic chromopeptide lactones with strong antineoplastic activity, were screened as inhibitors of Shc/Grb2 interaction in in vitro assay systems. To investigate the effects of actinomycin D on Shc/Grb2 interaction in cell‐based experiments, we used SAA (normal hEGFR‐overexpressed NIH3T3) cells and B104‐1‐1 (neu*‐transformed NIH3T3) cells, because a large number of the Shc/Grb2 complexes were detected. Associated protein complexes containing Shc were immunoprecipitated from actinomycin D‐treated cell lysates with polyclonal anti‐Shc antibody. Then the association with Grb2 was assessed by immunoblotting with monoclonal anti‐Grb2 antibody. The result of the immunoblotting experiment revealed that actinomycin D inhibited Shc/Grb2 interaction in a dose‐dependent manner in both B104‐1‐1 and EGF‐stimulated SAA cells. The inhibition of Shc/Grb2 interaction by actinomycin D in B104‐1‐1 cells also reduced tyrosine phosphorylation of MAP kinase (Erk1/Erk2), one of the major components in the Ras‐MAP kinase signaling pathway. These results suggest that actinomycin D could be a non‐phosphorylated natural and cellular membrane‐permeable SH2 domain antagonist.Keywords
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