Abstract A42: In vitro and in vivo studies of c-Myc amplifiers function

c-Myc, the product of a proto-oncogene Myc originally was identified as a transcriptional factor, which interacts with MAX protein through the basic helix-loop-helix leucine zipper (bHLHZip) domain and binds to the DNA containing the E-box (CACGTG) to regulate global gene expression. Recently, we and others found that c-Myc is a genome-wide non-linear amplifier of all expressed genes. The purpose of this study is to confirm previous results that c-Myc targets active promoters and enhancers to amplify the gene expression and to define the contributions of different Myc boxes (Mbs) in the regulation of gene expression and to identify the protein partners of Mbs. A Gibson Assembly system was used to create the A to N mutations of conserved amino acids in different Mbs. The wild type (WT) and mutated Myc –Pd4-EGFP fusion genes were transiently co-transfected to U2OS cells with Tk-, GRE-TK-, PR-TK-Luc vectors and analyze the reporter activities with a Dual-Luciferase Reporter Assay System, respectively. Our preliminary data demonstrated that WT-c-Myc does amplify reporter gene activity non-linearly. However, constructs with mutatied MbI and MbII were handicapped for amplification. Interestingly, mutated MbIII-b dramatically enhanced reporter gene amplification especially at higher Myc concentrations. Using reporter constructs with and without E-boxes, we found that this element is not necessary to enable c-Myc 9s amplification function. To understand c-Myc9s function in vivo, we established several Tet-inducible WT and mutated c-Myc stable cell lines with lenti-virus transduced system. The gene expression profiles of these cell lines are being compared. The screening of the partners of different Mb domains is on the way. Citation Format: Zuqin Nie, Chunhua Guo, Stoney S. Simons, David L. Levens. In vitro and in vivo studies of c-Myc amplifiers function. [abstract]. In: Proceedings of the AACR Special Conference on Myc: From Biology to Therapy; Jan 7-10, 2015; La Jolla, CA. Philadelphia (PA): AACR; Mol Cancer Res 2015;13(10 Suppl):Abstract nr A42.