Phosphorylation-Induced Dimerization of Interferon Regulatory Factor 7 Unmasks DNA Binding and a Bipartite Transactivation Domain

Abstract

Interferon regulatory factor 7 (IRF7) is an interferon (IFN)-inducible transcription factor required for activation of a subset of IFN-α genes that are expressed with delayed kinetics following viral infection. IRF7 is synthesized as a latent protein and is posttranslationally modified by protein phosphorylation in infected cells. Phosphorylation required a carboxyl-terminal regulatory domain that controlled the retention of the active protein exclusively in the nucleus, as well as its binding to specific DNA target sequences, multimerization, and ability to induce target gene expression. Transcriptional activation by IRF7 mapped to two distinct regions, both of which were required for full activity, while all functions were masked in latent IRF7 by an autoinhibitory domain mapping to an internal region. A conditionally active form of IRF7 was constructed by fusing IRF7 with the ligand-binding and dimerization domain of estrogen receptor (ER). Hormone-dependent dimerization of chimeric IRF7-ER stimulated DNA binding and transcriptional transactivation of endogenous target genes. These studies demonstrate the regulation of IRF7 activity by phosphorylation-dependent allosteric changes that result in dimerization and that facilitate nuclear retention, derepress transactivation, and allow specific DNA binding.

ACKNOWLEDGMENTS

We thank Sarah Guadagno (Zymed Laboratories) for preparation of IRF7 antibodies, Hongyong Zheng and Adolfo García-Sastre (Mount Sinai School of Medicine, New York, N.Y.) for the gift of NDV and for helpful discussions, Charles Weissmann for communicating data before publication, Heather Harding (NYU) for advice on isoelectric focusing, Martin Seidel (Ligand) for helpful discussions, Tim Hoey (Tularik) for the gift of Gal4-UAS-luciferase, Hans Bluyssen and Li Pan for originally isolating IRF7, Doreen Ray and Regina Raz for determining its exon-intron structure, David Ron (NYU) for comments on the manuscript, and members of our laboratory for assistance and helpful discussions.

This work was supported by Public Health Services grant R01AI28900 from the National Institute of Allergy and Infectious Diseases and by a postdoctoral fellowship to E.S. from the Arthritis Foundation.