MCL1 is an anti-apoptotic member of the Bcl-2 family, which has become a popular therapeutic target in cancer research over the past decade due to its propensity to promote cancer cell survival and facilitate chemoresistance. Through a phage display screen, we identified a novel MCL1 binding motif that is the reverse of the canonical BH3-binding helix that mediates interactions amongst Bcl-2 family members. Upon a BLAST sequence analysis of native proteins, we identified a putative reverse BH3 (rBH3) motif in the tumor suppressor protein, p73. Here, we show that MCL1 and p73 bind through a direct protein-protein interaction. Using fluorescence polarization assay, we characterize the strength of this interaction and reduce binding down to the rBH3-containing alpha helix in the tetramerization domain of p73. To elucidate biological function, we employ electrophoretic mobility shift assay to show that excess MCL1 negatively impacts p73 binding to DNA. Finally, using two p53-/- cancer cell lines, we demonstrate that MCL1 inhibits the transcriptional function of p73 through analysis of p73 target gene expression. In summary, we characterize a novel protein-protein interaction between MCL1 and p73 and show that MCL1 negatively impacts p73 target gene activation. This work establishes a novel function of MCL1 outside of apoptotic regulation at the mitochondria and provides new evidence of cross talk between the Bcl-2 family and the DNA damage response pathway.

Citation Format: Hayley Widden, Aneta Kaczmarczyk, Robert H. Whitaker, William J. Placzek. MCL1 binds and negatively regulates the transcriptional function of tumor suppressor p73 [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 2498.