BINDING OF ETS FAMILY MEMBERS IS IMPORTANT FOR THE FUNCTION OF THE C-SIS PLATELET-DERIVED GROWTH FACTOR-B TATA NEIGHBORING SEQUENCE IN 12-O-TETRADECANOYLPHORBOL-13-ACETATE-TREATED K562 CELLS/

The c-sis/platelet-derived growth factor (PDGF)-B TATA neighboring seq uence (TNS) is a promoter element that is required for the full induct ion of this gene in K562 erythroleukemia cells undergoing 12-O-tetrade canoylphorbol-13-acetate-mediated megakaryoblastic differentiation. Nu clear factors from K562 cells can bind to the c-sis/PDGF-B TNS, genera ting four complexes in electrophoretic mobility shift assays. One of t hese complexes was previously shown to contain Sp family members. In t his work, we provide evidence implicating two of the remaining complex es as belonging to the ETS family of transcription factors. This inclu des the identification of a novel constitutive TNS-binding complex con taining the ETS family member ELK-1, The binding of both ETS-like comp lexes was disrupted by mutations in a central CCGGAA core within the T NS and, for one of the complexes, could be promoted by bringing the se quences flanking the core closer to a consensus ETS binding site. The molecular weights of these TNS-binding factors were estimated by UV cr oss-linking analysis and found to be consistent with those of several ETS family transcription factors, including ELK-1, A consensus ELK-1 b inding site could compete for the binding of both putative ETS-like fa ctors, and the novel complex could be disrupted by the addition of an antibody raised against ELK-1. Transient transfection analysis using m utant TNS promoter-reporter constructs demonstrated a strong correlati on between the binding of the ETS-like factors and the transcriptional activity of the TNS, In contrast, mutations that prevented the bindin g of Sp family transcription factors had no effect on promoter activit y. Thus, ETS family members, such as ELK-1, are not only capable of bi nding to the TNS but seem to be necessary for the function of this ele ment in differentiating K562 cells.