The MUC1-C Oncoprotein Binds to the BH3 Domain of the Pro-apoptotic BAX Protein and Blocks BAX Function

Rehan Ahmad,Maroof Alam,Hasan Rajabi,Donald Kufe

doi:10.1074/jbc.m112.357293

Rehan Ahmad, Maroof Alam + Show 2 more

Open Access

https://doi.org/10.1074/jbc.m112.357293

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Abstract

The pro-apoptotic BAX protein contains a BH3 domain that is necessary for its dimerization and for activation of the intrinsic apoptotic pathway. The MUC1 (mucin 1) heterodimeric protein is overexpressed in diverse human carcinomas and blocks apoptosis in the response to stress. In this study, we demonstrate that the oncogenic MUC1-C subunit associates with BAX in human cancer cells. MUC1-C·BAX complexes are detectable in the cytoplasm and mitochondria and are induced by genotoxic and oxidative stress. The association between MUC1-C and BAX is supported by the demonstration that the MUC1-C cytoplasmic domain is sufficient for the interaction with BAX. The results further show that the MUC1-C cytoplasmic domain CQC motif binds directly to the BAX BH3 domain at Cys-62. Consistent with binding to the BAX BH3 domain, MUC1-C blocked BAX dimerization in response to (i) truncated BID in vitro and (ii) treatment of cancer cells with DNA-damaging agents. In concert with these results, MUC1-C attenuated localization of BAX to mitochondria and the release of cytochrome c. These findings indicate that the MUC1-C oncoprotein binds directly to the BAX BH3 domain and thereby blocks BAX function in activating the mitochondrial death pathway.

Highlights

The MUC1 C-terminal subunit (MUC1-C) oncoprotein attenuates activation of the intrinsic apoptotic pathway by mechanisms that are not understood
To determine whether MUC1-C interacts with the pro-apoptotic BAX protein, lysates from MCF-7 breast cancer cells were immunoprecipitated with anti-BAX antibody
In pulldown experiments, incubation of the cytoplasmic fraction with MUC1-C cytoplasmic domain (MUC1-CD) fused to GST demonstrated binding with BAX (Fig. 1E)

Summary

Introduction

The MUC1-C oncoprotein attenuates activation of the intrinsic apoptotic pathway by mechanisms that are not understood. Results: MUC1-C binds directly to the pro-apoptotic BAX protein at the critical BH3 domain. Consistent with binding to the BAX BH3 domain, MUC1-C blocked BAX dimerization in response to (i) truncated BID in vitro and (ii) treatment of cancer cells with DNA-damaging agents. In concert with these results, MUC1-C attenuated localization of BAX to mitochondria and the release of cytochrome c. These findings indicate that the MUC1-C oncoprotein binds directly to the BAX BH3 domain and thereby blocks BAX function in activating the mitochondrial death pathway

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