The ETS1 gene expression co-related well with the T-cell receptor (TCR) alpha gene expression during thymic development, in different thymic development, in different thymic and T-cell subsets and during T-cell activation, indicating that ETS1 may regulate TCR alpha genes. The expression of the TCR alpha gene is regulated by a T-cell-specific enhancer located downstream of the C-alpha locus. One of the enhancer elements (T-alpha2) was used for screening the lambda-gt11 expression library made from Jurkat T-cell mRNA to identify the cDNA that programs the expression of the T-alpha2 binding proteins. One clone, upon DNA sequencing, is found to code for the entire ETS2 protein. The beta-gal-ETS1 fusion protein produced in E. coli binds to the T-alpha2 motif. Mutation in the T-alpha2 abolishes ETS1 binding and T-cell enhancer function, indicating ETS1 nuclear protein binds to DNA in a sequence-specific manner. Comparison of DNA sequences bound by ETS1 and subsequent site-specific mutational analyses indicate that centrally-located, purine-rich sequences (ETS Responsive Element) are involved in binding. Many nuclear proteins bind to ERE. Different size ERE protein complexes are seen in T and other hematopoietic and non- hematopoietic cells. In T-cells, three different complexes are seen with ERE. Complex I contains ETS1, and complex III contains an additional ETS-related protein. Since ETS1 expression and regulation are different in T and other cell types, characterization of these transacting factors are in progress. Jurkat cells have been transfected with ETS1 expression vectors in both sense and antisense orientations. Several clones have been obtained expressing antisense ETS1 mRNA, which blocks ETS1 protein formation. Characterization of these clones with respect to their growth property, mitogenic requirements and other target candidate gene expression are under investigation. The ERG gene encodes a nuclear phosphoprotein. Characterization of hematopoietic stem cells expressing the ERG gene is in progress.
