COOPERATIVE INTERACTION OF ESTROGEN-RECEPTOR ZINC-FINGER DOMAIN POLYPEPTIDES ON DNA-BINDING

The consensus oestrogen response element (ERE) contains two inverted c opies of an AGGTCA consensus hexameric half-site, spaced by three base pairs. It differs from many other hormone response elements, such as consensus thyroid (TREp) and retinoic acid (DR-5 RARE) response elemen ts, only in the relative spacing and orientation of these sequences. I n the present study we report values for cooperativity (omega) of an o estrogen receptor DNA-binding domain polypeptide upon binding to these sequences. The polypeptide binds with negative cooperativity, or with out cooperativity to retinoic acid and thyroid response elements respe ctively, but with high cooperativity to the ERE. We have also examined cooperativity upon binding of the polypeptide to an ERE variant. Sinc e naturally occurring EREs commonly contain one hexamer which is consi derably more degenerate than the other, we designed a hybrid response element in which one hexamer is a consensus ERE, while specific mutati ons were introduced into the other. We chose to mutate the second half -site to a glucocorticoid response element (GRE) half-site sequence (A GAACA), since normally no binding of the DNA-binding domain polypeptid e to a GRE hexamer alone can be detected. In the hybrid response demen t, however, the GRE half-site is recognized with relatively high affin ity, although binding to this sequence is dependent on the previous bi nding of a polypeptide to the ERE hexamer. Thus, cooperative interacti ons are capable of mediating the recognition of ERE sequence degenerac y. The ability of protein-protein interactions to mediate recognition of DNA sequence degeneracy may also have implications for transcriptio n factors in general.