Collaborative Competition Mechanism for Gene Activation In Vivo

Abstract

The mechanism by which gene regulatory proteins gain access to their DNA target sites is not known. In vitro, binding is inherently cooperative between arbitrary DNA binding proteins whose target sites are located within the same nucleosome. We refer to such competition-based cooperativity as collaborative competition. Here we show that arbitrarily chosen foreign DNA binding proteins, LexA and Tet repressor, cooperate with an adjacently binding endogenous activator protein, Gcn4, to coactivate expression of chromosomal reporter genes in Saccharomyces cerevisiae. Coactivation requires that the cooperating target sites be within a nucleosome-length distance; it leads to increased occupancy by Gcn4 at its binding site; and it requires both Gcn5 and Swi/Snf which, at an endogenous Gcn4-dependent promoter, act subsequent to Gcn4 binding. These results imply that collaborative competition contributes to gene regulation in vivo. They further imply that, even in the presence of the cell's full wild-type complement of chromatin remodeling factors, competition of regulatory proteins with histone octamer for access to regulatory target sites remains a quantitative determinant of gene expression levels. We speculate that initial target site recognition and binding may occur via spontaneous nucleosomal site exposure, with remodeling factor action required downstream to lock in higher levels of regulatory protein occupancy.

ACKNOWLEDGMENTS

This work was supported by an NIH grant (J.W.) and a Howard Hughes Medical Institute (HHMI) Predoctoral Fellowship (J.A.M.).

We thank Peggy Lowary for valuable scientific discussions and Eileen Chang for cloning the 154-bp spacer plasmid. We thank Paul Mason and Kevin Struhl for reagents and helpful advice in the chromatin immunoprecipitation assay and Randall Morse for the suggestion to use the HIS4 locus as a control. We gratefully acknowledge the use of the flow cytometer in the laboratory of Robert Lamb (HHMI) and instruments in the Keck Biophysics Facility at Northwestern University (http://www.biochem.northwestern.edu/Keck/keckmain.html ).