title: AKT1, LKB1, and YAP1 Revealed as MYC Interactors with NanoLuc-Based Protein-Fragment Complementation Assay.
abstract: The c-Myc (MYC) transcription factor is a major cancer driver and a well-validated therapeutic target. However, directly targeting MYC has been challenging. Thus, identifying proteins that interact with and regulate MYC may provide alternative strategies to inhibit its oncogenic activity. In this study, we report the development of a NanoLuc-based protein-fragment complementation assay (NanoPCA) and mapping of the MYC protein interaction hub in live mammalian cells. The NanoPCA system was configured to enable detection of protein-protein interactions (PPI) at the endogenous level, as shown with PRAS40 dimerization, and detection of weak interactions, such as PINCH1-NCK2. Importantly, NanoPCA allows the study of PPI dynamics with reversible interactions. To demonstrate its utility for large-scale PPI detection in mammalian intracellular environment, we have used NanoPCA to examine MYC interaction with 83 cancer-associated proteins in live cancer cell lines. Our new MYC PPI data confirmed known MYC-interacting proteins, such as MAX, GSK3A, and SMARCA4, and revealed a panel of novel MYC interaction partners, such as RAC-α serine/threonine-protein kinase (AKT)1, liver kinase B (LKB)1, and Yes-associated protein (YAP)1. The MYC interactions with AKT1, LKB1, and YAP1 were confirmed by coimmunoprecipitation of endogenous proteins. Importantly, AKT1, LKB1, and YAP1 were able to activate MYC in a transcriptional reporter assay. Thus, these vital growth control proteins may represent promising MYC regulators, suggesting new mechanisms that couple energetic and metabolic pathways and developmental signaling to MYC-regulated cellular programs.
authors.affiliation: Department of Pharmacology and Emory Chemical Biology Discovery Center (X.M., Q.Q., A.A.I., Q.N., Y.L., J.H., R.G., S.B., M.A.J., Y.D., H.F.) and Department of Biomedical Informatics (L.A.D.C.), Emory University School of Medicine, Atlanta, Georgia; Departments of Hematology and Medical Oncology (F.R.K., H.F.) and Pathology and Laboratory Medicine (C.S.M.) and Winship Cancer Institute, Emory University, Atlanta, Georgia; State Key Laboratory of Pharmaceutical Biotechnology, School of Life Sciences, Nanjing University, Nanjing, China (Y.L.); and Department of Biomedical Engineering, Emory University School of Medicine/Georgia Institute of Technology, Atlanta, Georgia (L.A.D.C.).
authors.forename: Xiulei
authors.initials: X
authors.lastname: Mo
authors.affiliation: Department of Pharmacology and Emory Chemical Biology Discovery Center (X.M., Q.Q., A.A.I., Q.N., Y.L., J.H., R.G., S.B., M.A.J., Y.D., H.F.) and Department of Biomedical Informatics (L.A.D.C.), Emory University School of Medicine, Atlanta, Georgia; Departments of Hematology and Medical Oncology (F.R.K., H.F.) and Pathology and Laboratory Medicine (C.S.M.) and Winship Cancer Institute, Emory University, Atlanta, Georgia; State Key Laboratory of Pharmaceutical Biotechnology, School of Life Sciences, Nanjing University, Nanjing, China (Y.L.); and Department of Biomedical Engineering, Emory University School of Medicine/Georgia Institute of Technology, Atlanta, Georgia (L.A.D.C.).
authors.forename: Qi
authors.initials: Q
authors.lastname: Qi
authors.affiliation: Department of Pharmacology and Emory Chemical Biology Discovery Center (X.M., Q.Q., A.A.I., Q.N., Y.L., J.H., R.G., S.B., M.A.J., Y.D., H.F.) and Department of Biomedical Informatics (L.A.D.C.), Emory University School of Medicine, Atlanta, Georgia; Departments of Hematology and Medical Oncology (F.R.K., H.F.) and Pathology and Laboratory Medicine (C.S.M.) and Winship Cancer Institute, Emory University, Atlanta, Georgia; State Key Laboratory of Pharmaceutical Biotechnology, School of Life Sciences, Nanjing University, Nanjing, China (Y.L.); and Department of Biomedical Engineering, Emory University School of Medicine/Georgia Institute of Technology, Atlanta, Georgia (L.A.D.C.).
authors.forename: Andrei A
authors.initials: AA
authors.lastname: Ivanov
authors.affiliation: Department of Pharmacology and Emory Chemical Biology Discovery Center (X.M., Q.Q., A.A.I., Q.N., Y.L., J.H., R.G., S.B., M.A.J., Y.D., H.F.) and Department of Biomedical Informatics (L.A.D.C.), Emory University School of Medicine, Atlanta, Georgia; Departments of Hematology and Medical Oncology (F.R.K., H.F.) and Pathology and Laboratory Medicine (C.S.M.) and Winship Cancer Institute, Emory University, Atlanta, Georgia; State Key Laboratory of Pharmaceutical Biotechnology, School of Life Sciences, Nanjing University, Nanjing, China (Y.L.); and Department of Biomedical Engineering, Emory University School of Medicine/Georgia Institute of Technology, Atlanta, Georgia (L.A.D.C.).
authors.forename: Qiankun
authors.initials: Q
authors.lastname: Niu
authors.affiliation: Department of Pharmacology and Emory Chemical Biology Discovery Center (X.M., Q.Q., A.A.I., Q.N., Y.L., J.H., R.G., S.B., M.A.J., Y.D., H.F.) and Department of Biomedical Informatics (L.A.D.C.), Emory University School of Medicine, Atlanta, Georgia; Departments of Hematology and Medical Oncology (F.R.K., H.F.) and Pathology and Laboratory Medicine (C.S.M.) and Winship Cancer Institute, Emory University, Atlanta, Georgia; State Key Laboratory of Pharmaceutical Biotechnology, School of Life Sciences, Nanjing University, Nanjing, China (Y.L.); and Department of Biomedical Engineering, Emory University School of Medicine/Georgia Institute of Technology, Atlanta, Georgia (L.A.D.C.).
authors.forename: Yin
authors.initials: Y
authors.lastname: Luo
authors.affiliation: Department of Pharmacology and Emory Chemical Biology Discovery Center (X.M., Q.Q., A.A.I., Q.N., Y.L., J.H., R.G., S.B., M.A.J., Y.D., H.F.) and Department of Biomedical Informatics (L.A.D.C.), Emory University School of Medicine, Atlanta, Georgia; Departments of Hematology and Medical Oncology (F.R.K., H.F.) and Pathology and Laboratory Medicine (C.S.M.) and Winship Cancer Institute, Emory University, Atlanta, Georgia; State Key Laboratory of Pharmaceutical Biotechnology, School of Life Sciences, Nanjing University, Nanjing, China (Y.L.); and Department of Biomedical Engineering, Emory University School of Medicine/Georgia Institute of Technology, Atlanta, Georgia (L.A.D.C.).
authors.forename: Jonathan
authors.initials: J
authors.lastname: Havel
authors.affiliation: Department of Pharmacology and Emory Chemical Biology Discovery Center (X.M., Q.Q., A.A.I., Q.N., Y.L., J.H., R.G., S.B., M.A.J., Y.D., H.F.) and Department of Biomedical Informatics (L.A.D.C.), Emory University School of Medicine, Atlanta, Georgia; Departments of Hematology and Medical Oncology (F.R.K., H.F.) and Pathology and Laboratory Medicine (C.S.M.) and Winship Cancer Institute, Emory University, Atlanta, Georgia; State Key Laboratory of Pharmaceutical Biotechnology, School of Life Sciences, Nanjing University, Nanjing, China (Y.L.); and Department of Biomedical Engineering, Emory University School of Medicine/Georgia Institute of Technology, Atlanta, Georgia (L.A.D.C.).
authors.forename: Russell
authors.initials: R
authors.lastname: Goetze
authors.affiliation: Department of Pharmacology and Emory Chemical Biology Discovery Center (X.M., Q.Q., A.A.I., Q.N., Y.L., J.H., R.G., S.B., M.A.J., Y.D., H.F.) and Department of Biomedical Informatics (L.A.D.C.), Emory University School of Medicine, Atlanta, Georgia; Departments of Hematology and Medical Oncology (F.R.K., H.F.) and Pathology and Laboratory Medicine (C.S.M.) and Winship Cancer Institute, Emory University, Atlanta, Georgia; State Key Laboratory of Pharmaceutical Biotechnology, School of Life Sciences, Nanjing University, Nanjing, China (Y.L.); and Department of Biomedical Engineering, Emory University School of Medicine/Georgia Institute of Technology, Atlanta, Georgia (L.A.D.C.).
authors.forename: Sydney
authors.initials: S
authors.lastname: Bell
authors.affiliation: Department of Pharmacology and Emory Chemical Biology Discovery Center (X.M., Q.Q., A.A.I., Q.N., Y.L., J.H., R.G., S.B., M.A.J., Y.D., H.F.) and Department of Biomedical Informatics (L.A.D.C.), Emory University School of Medicine, Atlanta, Georgia; Departments of Hematology and Medical Oncology (F.R.K., H.F.) and Pathology and Laboratory Medicine (C.S.M.) and Winship Cancer Institute, Emory University, Atlanta, Georgia; State Key Laboratory of Pharmaceutical Biotechnology, School of Life Sciences, Nanjing University, Nanjing, China (Y.L.); and Department of Biomedical Engineering, Emory University School of Medicine/Georgia Institute of Technology, Atlanta, Georgia (L.A.D.C.).
authors.forename: Carlos S
authors.initials: CS
authors.lastname: Moreno
authors.affiliation: Department of Pharmacology and Emory Chemical Biology Discovery Center (X.M., Q.Q., A.A.I., Q.N., Y.L., J.H., R.G., S.B., M.A.J., Y.D., H.F.) and Department of Biomedical Informatics (L.A.D.C.), Emory University School of Medicine, Atlanta, Georgia; Departments of Hematology and Medical Oncology (F.R.K., H.F.) and Pathology and Laboratory Medicine (C.S.M.) and Winship Cancer Institute, Emory University, Atlanta, Georgia; State Key Laboratory of Pharmaceutical Biotechnology, School of Life Sciences, Nanjing University, Nanjing, China (Y.L.); and Department of Biomedical Engineering, Emory University School of Medicine/Georgia Institute of Technology, Atlanta, Georgia (L.A.D.C.).
authors.forename: Lee A D
authors.initials: LA
authors.lastname: Cooper
authors.affiliation: Department of Pharmacology and Emory Chemical Biology Discovery Center (X.M., Q.Q., A.A.I., Q.N., Y.L., J.H., R.G., S.B., M.A.J., Y.D., H.F.) and Department of Biomedical Informatics (L.A.D.C.), Emory University School of Medicine, Atlanta, Georgia; Departments of Hematology and Medical Oncology (F.R.K., H.F.) and Pathology and Laboratory Medicine (C.S.M.) and Winship Cancer Institute, Emory University, Atlanta, Georgia; State Key Laboratory of Pharmaceutical Biotechnology, School of Life Sciences, Nanjing University, Nanjing, China (Y.L.); and Department of Biomedical Engineering, Emory University School of Medicine/Georgia Institute of Technology, Atlanta, Georgia (L.A.D.C.).
authors.forename: Margaret A
authors.initials: MA
authors.lastname: Johns
authors.affiliation: Department of Pharmacology and Emory Chemical Biology Discovery Center (X.M., Q.Q., A.A.I., Q.N., Y.L., J.H., R.G., S.B., M.A.J., Y.D., H.F.) and Department of Biomedical Informatics (L.A.D.C.), Emory University School of Medicine, Atlanta, Georgia; Departments of Hematology and Medical Oncology (F.R.K., H.F.) and Pathology and Laboratory Medicine (C.S.M.) and Winship Cancer Institute, Emory University, Atlanta, Georgia; State Key Laboratory of Pharmaceutical Biotechnology, School of Life Sciences, Nanjing University, Nanjing, China (Y.L.); and Department of Biomedical Engineering, Emory University School of Medicine/Georgia Institute of Technology, Atlanta, Georgia (L.A.D.C.).
authors.forename: Fadlo R
authors.initials: FR
authors.lastname: Khuri
authors.affiliation: Department of Pharmacology and Emory Chemical Biology Discovery Center (X.M., Q.Q., A.A.I., Q.N., Y.L., J.H., R.G., S.B., M.A.J., Y.D., H.F.) and Department of Biomedical Informatics (L.A.D.C.), Emory University School of Medicine, Atlanta, Georgia; Departments of Hematology and Medical Oncology (F.R.K., H.F.) and Pathology and Laboratory Medicine (C.S.M.) and Winship Cancer Institute, Emory University, Atlanta, Georgia; State Key Laboratory of Pharmaceutical Biotechnology, School of Life Sciences, Nanjing University, Nanjing, China (Y.L.); and Department of Biomedical Engineering, Emory University School of Medicine/Georgia Institute of Technology, Atlanta, Georgia (L.A.D.C.).
authors.forename: Yuhong
authors.initials: Y
authors.lastname: Du
authors.affiliation: Department of Pharmacology and Emory Chemical Biology Discovery Center (X.M., Q.Q., A.A.I., Q.N., Y.L., J.H., R.G., S.B., M.A.J., Y.D., H.F.) and Department of Biomedical Informatics (L.A.D.C.), Emory University School of Medicine, Atlanta, Georgia; Departments of Hematology and Medical Oncology (F.R.K., H.F.) and Pathology and Laboratory Medicine (C.S.M.) and Winship Cancer Institute, Emory University, Atlanta, Georgia; State Key Laboratory of Pharmaceutical Biotechnology, School of Life Sciences, Nanjing University, Nanjing, China (Y.L.); and Department of Biomedical Engineering, Emory University School of Medicine/Georgia Institute of Technology, Atlanta, Georgia (L.A.D.C.) hfu@emory.edu.
authors.forename: Haian
authors.initials: H
authors.lastname: Fu
mesh_terms: D001681:Biological Assay; D045744:Cell Line, Tumor; D057809:HEK293 Cells; D057166:High-Throughput Screening Assays; D006801:Humans; D008156:Luciferases / Q000378:metabolism*; D053758:Nanoparticles / Q000737:chemistry*; D010750:Phosphoproteins / Q000378:metabolism*; D011485:Protein Binding; D025941:Protein Interaction Mapping / Q000379:methods*; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism*; D016271:Proto-Oncogene Proteins c-myc / Q000378:metabolism*; D015203:Reproducibility of Results
chemical_list: D010750:Phosphoproteins; D016271:Proto-Oncogene Proteins c-myc; D008156:Luciferases; D017346:Protein Serine-Threonine Kinases; D051057:Proto-Oncogene Proteins c-akt