The IRF family of transcription factors plays critical roles in the regulation of interferons in response to viral infection, and in the development and functioning of the immune system. The IRF family includes now over ten members, characterized by a homologous DNA binding domain (DBD) at the N-terminus. We propose structural studies to explore the broader role of IRF-3 in the regulation of interferon-beta (IFN-beta) gene expression, and the role of IRF-4 in the development and functioning of the immune system. IRF-3 is activated in virally infected cells by phosphorylation of specific residues at the C-terminus, leading to nuclear translocation and binding to a so-called PRD I-III DNA element in the IFN-beta promoter. IRF-4 binds to a number of composite DNA elements in the promoters and enhancers of B-lymphoid and myeloid genes, but exclusively in association with PU.1.
Specific aims are: 1) Determine the structure of IRF-3 DBD bound to the entire PRD I-III element by crystallographic methods. 2) Determine the structure of intact, phosphorylated IRF-3 bound to the PRD I-III element. IRF-3 will be phosphorylated in vitro prior to crystallization. 3) Structurally characterize an autoinhibitory element at the N-terminus of IRF-4 by NMR methods. 4) Determine the structure of IRF-4 in complex with phosphorylated PU.I. PU.1 will be phosphorylated in vitro by casein kinase II prior to crystallization. Together, these aims explore the specificity and cooperativity of these IRFs, and the role of phosphorylation and autoinhibitory elements.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Research Project (R01)
Project #
5R01AI041706-10
Application #
7534982
Study Section
Special Emphasis Panel (ZRG1-BPC-B (02))
Program Officer
Leitner, Wolfgang W
Project Start
1998-06-01
Project End
2010-11-30
Budget Start
2008-12-01
Budget End
2010-11-30
Support Year
10
Fiscal Year
2009
Total Cost
$275,909
Indirect Cost
Name
Icahn School of Medicine at Mount Sinai
Department
Physiology
Type
Schools of Medicine
DUNS #
078861598
City
New York
State
NY
Country
United States
Zip Code
10029
Malu, Shruti; De Ioannes, Pablo; Kozlov, Mikhail et al. (2012) Artemis C-terminal region facilitates V(D)J recombination through its interactions with DNA Ligase IV and DNA-PKcs. J Exp Med 209:955-63
De Ioannes, Pablo; Escalante, Carlos R; Aggarwal, Aneel K (2011) Structures of apo IRF-3 and IRF-7 DNA binding domains: effect of loop L1 on DNA binding. Nucleic Acids Res 39:7300-7
Nistal-Villán, Estanislao; Gack, Michaela U; Martínez-Delgado, Gustavo et al. (2010) Negative role of RIG-I serine 8 phosphorylation in the regulation of interferon-beta production. J Biol Chem 285:20252-61
Joshi, Rohit; Passner, Jonathan M; Rohs, Remo et al. (2007) Functional specificity of a Hox protein mediated by the recognition of minor groove structure. Cell 131:530-43
Escalante, Carlos R; Nistal-Villan, Estanislao; Shen, Leyi et al. (2007) Structure of IRF-3 bound to the PRDIII-I regulatory element of the human interferon-beta enhancer. Mol Cell 26:703-16
James, J Anson; Aggarwal, Aneel K; Linden, R Michael et al. (2004) Structure of adeno-associated virus type 2 Rep40-ADP complex: insight into nucleotide recognition and catalysis by superfamily 3 helicases. Proc Natl Acad Sci U S A 101:12455-60
Yoon-Robarts, Miran; Blouin, Amanda G; Bleker, Svenja et al. (2004) Residues within the B' motif are critical for DNA binding by the superfamily 3 helicase Rep40 of adeno-associated virus type 2. J Biol Chem 279:50472-81
James, J Anson; Escalante, Carlos R; Yoon-Robarts, Miran et al. (2003) Crystal structure of the SF3 helicase from adeno-associated virus type 2. Structure 11:1025-35
Escalante, Carlos R; Brass, Abraham L; Pongubala, Jagan M R et al. (2002) Crystal structure of PU.1/IRF-4/DNA ternary complex. Mol Cell 10:1097-105
Escalante, Carlos R; Shen, Leyi; Thanos, Dimitris et al. (2002) Structure of NF-kappaB p50/p65 heterodimer bound to the PRDII DNA element from the interferon-beta promoter. Structure 10:383-91

Showing the most recent 10 out of 11 publications