SYSTEMATIC QUANTITATIVE ANALYSIS OF NF-kappaB CO-ACTIVATORS

Lead Research Organisation: Imperial College London
Department Name: Unlisted

Abstract

Inflammation is a normal physiological response to infection and injury, but can lead to extensive tissue damage and disability when elicited in excess. Pathological consequences of sustained inflammatory response include variety of autoimmune diseases, such as rheumatoid arthritis, Crohn s disease, ankylosing spondylitis and multiple sclerosis. A sustained inflammatory response is often linked to the break down in regulation of production of inflammatory molecules. In a normal self-resolving body response to infection or injury inflammatory molecules are produced for a defined period of time, while in chronic autoimmune conditions their production is prolonged.
The production of many inflammatory molecules is controlled by a key family of regulatory proteins called NF-kappaB. Ways to modulate levels of these transcription factors and restore a normal cytokine balance are considered to be of a potential therapeutic importance. But NF-kappaB does not function in isolation. Many of the NF-kappaB dependent immune genes are also co-regulated by other ubiquitous transcription factors, including the IRF (interferon regulatory factor) family of transcription factors consisting of nine proteins. Which IRF proteins are able to synergise with NF-kappaB to regulate gene expression, and how this co-regulation is dependent on the sequence of genomic elements attracting these factors is the subject of this investigation.
Understanding the basic elements underlying successful inflammatory response is important for designing targeted strategies for suppressing inflammation while preserving signals required for host defence. It will also help scientists to better understand why humans have so many families of ostensibly similar proteins and what specific functions they hold.
The research will be carried out by two groups: of Dr Irina Udalova at the Kennedy Institute of Rheumatology, Imperial College, and of Dr Ioannis Ragoussis at the Wellcome Trust Centre for Human Genetics, Oxford University. It will be laboratory based and will use novel genomic techniques and computational methods. The proposed novel techniques will replace the use of animals in research. Results generated in this study will be communicated through peer-reviewed scientific journals and through annual reports from respective Institutes and presented at scientific meetings.

Technical Summary

IRF transcription factors have recently emerged as widespread cofactors of NF-kappaB, the key regulator of gene expression in inflammation. The fundamental goal of this proposal is to accurately and quantitatively annotate the minimal genomic elements that drive inflammatory response via co-recruitment of NF-kappaB and IRF transcription factors. We propose to systematically define in vitro binding specificities of various IRF dimers and to test how altering IRF-DNA interactions in vivo affects the expression of inflammatory genes in human macrophages exposed to bacterial products. The project builds on our existing expertise and recently developed genomic approaches, e.g. the protein-DNA arrays. It develops new ways for studying transcriptional gene regulation, such as the use of viruses for in situ manipulation of the chromatin in human cells, and will generate new reagents for quantitative analysis of cellular responses available to the scientific community. The mechanistic understanding of inflammatory gene control will provide an alternative path for the development of targeted therapeutic strategies for treating immune disorders.

The project is in conjunction with the MRC NIA award to Dr Udalova and solidifies the basis for her long term goal of trying to identify the molecular mechanisms that ensure successful inflammatory response. It promotes a longstanding successful collaboration between the two applicants. The promoter specific modules of NF-kappaB and IRF binding elements computationally identified during the course of this project will form a framework for future experimental studies of coordinated gene regulation in inflammation.
 
Title Photo exhibition 
Description "Scientists at work" photo exhibition in the local gallery 
Type Of Art Artwork 
Year Produced 2012 
Impact Members of the group organising a photo club and taking monthly photos of interest. The topic "Scientists at work" was presented at the local gallery in London 
 
Description CO-REGULATION OF MACROPHAGE INFLAMMATORY PHENOTYPE BY IRF5 AND RELA
Amount £332,796 (GBP)
Funding ID MR/J001899/1 
Organisation Medical Research Council (MRC) 
Sector Academic/University
Country United Kingdom
Start 01/2012 
End 01/2015
 
Description Can IRF5-expressing pro-inflammatory macrophages control allergic inflammation?
Amount £300,000 (GBP)
Organisation American Asthma Foundation 
Sector Charity/Non Profit
Country United States
Start 07/2011 
End 06/2014
 
Title EMSA-Seq 
Description novel method for analysis of protein-DNA interactions 
Type Of Material Technology assay or reagent 
Year Produced 2009 
Provided To Others? Yes  
Impact potentially better, faster, cheaper way of profiling binding specificities of transcription factors - multiple usage 
 
Title IRF binding specificities 
Description quantitative data describing binding preferences of the IRF factors to housands of DNA sequences 
Type Of Material Database/Collection of Data/Biological Samples 
Provided To Others? No  
Impact extremely valuable for teasing out whole genome ChIP data and understanding transcriptional regulation of genes 
 
Title IRF expression constructs 
Description DNA vectors encoding cDNAs of various members of the Interferon Regulatory Factor family 
Type Of Material Technology assay or reagent 
Provided To Others? No  
Impact allow to proceed with the DNA binding analysis and useful for functional studies of inflammatory gene expression: Thomas Krausgruber, David Saliba, Grigory Ryzhakov, Alessandra Lanfrancotti, Katrina Blazek, Irina A Udalova "IRF5 promotes sustained TNF production by human dendritic cells via two independent mechanisms" (submitted) 
 
Description MB 
Organisation Harvard University
Department Harvard Medical School
Country United States 
Sector Academic/University 
PI Contribution access to the data, providing with reagents
Collaborator Contribution access to the data
Impact Principles of dimer-specific gene regulation revealed by a comprehensive characterization of NF-?B family DNA binding. Siggers T, Chang AB, Teixeira A, Wong D, Williams KJ, Ahmed B, Ragoussis J, Udalova IA, Smale ST, Bulyk ML. Nat Immunol. 2011 Nov 20;13(1):95-102.
Start Year 2007
 
Description SD 
Organisation Joseph Fourier University
Department The Albert Boniot Institute
Country France 
Sector Academic/University 
PI Contribution provising selection of sequences for further analysis
Collaborator Contribution analysis of TF-FNA interactions at a signle base resolution
Impact EU FP7 Model-In grant; Extensive characterization of NF-?B binding uncovers non-canonical motifs and advances the interpretation of genetic functional traits. Wong D, Teixeira A, Oikonomopoulos S, Humburg P, Lone IN, Saliba D, Siggers T, Bulyk M, Angelov D, Dimitrov S, Udalova IA, Ragoussis J. Genome Biol. 2011 Jul 29;12(7):R70
Start Year 2009
 
Title IRF5 in macrophage differentiation 
Description This invention relates to modulating the immune system, and in particular to methods for modulating the immune system to treat disease. The invention further relates to methods for identifying agents that modulate the immune system to treat disease 
IP Reference WO2012093258 
Protection Patent application published
Year Protection Granted 2011
Licensed No
Impact potential industrial collaborations
 
Description Aggressive Lymphoma symposium 
Form Of Engagement Activity A formal working group, expert panel or dialogue
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Health professionals
Results and Impact "Functional effects of genetic polymorphisms: quantitative analysis of TF-DNA interactions" 30 min presentation at Aggressive Lymphoma symposium (Gottingen, Germany)

education of medical doctors
Year(s) Of Engagement Activity 2007
 
Description INTAS genomics and proteomics workshop 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Public/other audiences
Results and Impact presentation "Quantitative analysis of protein-DNA interactions: application to functional genomics"

education of students and public in Ukraine
Year(s) Of Engagement Activity 2007
 
Description Radio and newspapers interviews 
Form Of Engagement Activity A press release, press conference or response to a media enquiry/interview
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Media (as a channel to the public)
Results and Impact press releases on our discovery of the master regulation of mph polarization

external visibility, public education
Year(s) Of Engagement Activity 2011