Domain Transcription Factor Family Research Articles

Characterization of the Elk-1 ETS DNA-binding Domain

The ETS domain family of transcription factors is comprised of several important proteins that are involved in controlling key cellular events such as proliferation, differentiation, and development. One such protein, Elk-1, regulates the activity of the c-fos promoter in response to extracellular stimuli. Elk-1 is representative of a subgroup of ETS domain proteins that utilize a bipartite recognition mechanism that is mediated by both protein-DNA and protein-protein interactions. In this study, we have overexpressed, purified, and characterized the ETS DNA-binding domain of Elk-1 (Elk-93). Elk-93 was expressed in Escherichia coli as a fusion protein with glutathione S-transferase and purified to homogeneity from both the soluble and insoluble fractions using a two-column protocol. A combination of CD, NMR, and fluorescence spectroscopy demonstrates that Elk-93 represents an independently folded domain of mixed alpha/beta structure in which the three conserved tryptophans appear to contribute to the hydrophobic core of the protein. Moreover, DNA binding studies demonstrate that Elk-93 binds DNA with both high affinity (Kd approximately 0.85 x 10(-10)M) and specificity. Circular permutation analysis indicates that DNA binding by Elk-93 does not induce significant bending of the DNA. Our results are discussed with respect to predictive models for the structure of the ETS DNA-binding domain.

New member of the winged-helix protein family disrupted in mouse and rat nude mutations.

Mutations at the nude locus of mice and rats disrupt normal hair growth and thymus development, causing nude mice and rats to be immune-deficient. The mouse nude locus has been localized on chromosome 11 (refs 3, 4) within a region of < 1 megabase. Here we show that one of the genes from this critical region, designated whn, encodes a new member of the winged-helix domain family of transcription factors, and that it is disrupted on mouse nu and rat rnuN alleles. Mutant transcripts do not encode the characteristic DNA-binding domain, strongly suggesting that the whn gene is the nude gene. Mutations in winged-helix domain genes cause homeotic transformations in Drosophila and distort cell-fate decisions during vulval development in Caenorhabditis elegans. The whn gene is thus the first member of this class of genes to be implicated in a specific developmental defect in vertebrates.

Fusion of a fork head domain gene to PAX3 in the solid tumour alveolar rhabdomyosarcoma

We have examined the structure and expression of the products associated with the t(2;13)(q35;q14) translocation associated with alveolar rhabdomyosarcoma. The chromosome 13 gene (FKHR) is identified as a member of the fork head domain family of transcription factors characterized by a conserved DNA binding motif. Polymerase chain reaction analysis demonstrates that a 5'PAX3-3' FKHR chimaeric transcript is expressed in all eight alveolar rhabdomyosarcomas investigated. Immunoprecipitation experiments detect the predicted fusion protein. These findings indicate that the t(2;13) generates a potentially tumorigenic fusion transcription factor consisting of intact PAX3 DNA binding domains, a truncated fork head DNA binding domain and C-terminal FKHR regions.