DISSECTING THE FUNCTION OF INDIVIDUAL NF-kappaB SUBUNITS IN INFLAMMATION

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. All five NF-kappaB proteins are present at the site of inflammation and are active, but at different times. Why there are five similar but not identical NF-kappaB proteins, why they come to and leave the site of inflammation at different times and how they control the level and the length of production of inflammatory molecules is a subject of this investigation.
Understanding how each NF-kappaB protein functions and what molecules it regulates is important for the development of a new generation of therapeutic approaches. These would only target specific cellular events without interfering with the global host defence mechanisms. It will also help scientists to better understand why we have so many families of ostensibly similar proteins and what specific functions they hold.
The research will be carried out by Dr Irina Udalova at the Kennedy Institute of Rheumatology, Imperial College, and will be laboratory based using 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 the Kennedy Institute and presented at scientific meetings.

Technical Summary

NF-kappaB plays a pivotal role in immunity and inflammation and is considered to be a prime candidate for drug development. The fundamental goal of this proposal is to identify NF-kappaB subunit specific regulatory events in a human immune response. I propose to systematically determine the genes whose expression is dependent on specific NF-kappaB dimeric complexes in primary human dendritic cells exposed to bacterial products and to characterise the functional genetic elements which select these complexes. The project builds on my existing expertise and available genomic and computational approaches and develops new ways for studying transcriptional gene regulation, such as the genetic manipulation of human cells, previously achievable only in animal models. It will generate new reagents for quantitative analysis of cellular responses available to the scientific community. The identification of NF-kappaB subunit specific regulatory events will provide an alternative path for the development of targeted therapeutic strategies for treating immune disorders.

Publications

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Field S (2007) Accuracy and reproducibility of protein-DNA microarray technology. in Advances in biochemical engineering/biotechnology

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Ragoussis J (2006) Quantitative profiling of protein-DNA binding on microarrays. in Methods in molecular biology (Clifton, N.J.)

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Thomson SJ (2009) The role of transposable elements in the regulation of IFN-lambda1 gene expression. in Proceedings of the National Academy of Sciences of the United States of America

 
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 EC FP7 (HEALTH-2.1.2-5)
Amount £425,000 (GBP)
Organisation European Commission 
Department Seventh Framework Programme (FP7)
Sector Public
Country European Union (EU)
Start 12/2008 
End 06/2012
 
Description MRC Collaborative Grant
Amount £208,121 (GBP)
Organisation Medical Research Council (MRC) 
Sector Academic/University
Country United Kingdom
Start 09/2007 
End 08/2009
 
Title NF-kappaB expression plasmids 
Description DNA constructs expressing various NF-kappaB subunits 
Type Of Material Technology assay or reagent 
Year Produced 2007 
Provided To Others? Yes  
Impact useful tools for analysis of gene regulation 
 
Title NF-kappaB genes 
Description database of the genes regulated by NF-kappaB 
Type Of Material Database/Collection of Data/Biological Samples 
Provided To Others? No  
Impact framework for genome-wide analysis of gene expression in inflammation 
 
Title RNAi 
Description optimized reagents for NF-kappaB inhibition in human cells; cell lines with depleted NF-kappaB levels 
Type Of Material Technology assay or reagent 
Year Produced 2008 
Provided To Others? Yes  
Impact useful tool for gene regulation studies 
 
Description GN 
Organisation European Institute of Oncology (IEO)
Country Italy 
Sector Charity/Non Profit 
PI Contribution Intellectual contribution to DNA-protein interactions and genome analysis
Collaborator Contribution Intellectual contribution to the aspects of gene regulation; transferring chromatin immunoprecipitation and 3C techniques
Impact Grants: 1. FP7-EU grant 222008 "Genomic determinants of inflammation" 2. Royal Society joint grant "Examining DNA sequence selectivity by alternative NF-kappaB proteins" 3. Horizon 2020 PCH03 proposal "Unifying basis of Chronic Inflammatory Diseases" submitted
Start Year 2006
 
Description JR 
Organisation University of Oxford
Department Wellcome Trust Centre for Human Genetics
Country United Kingdom 
Sector Academic/University 
PI Contribution I have initially set up and developed high throughput analysis of transcription factor - DNA interactions in vitro, further optimised and developed by the collaborator's team.
Collaborator Contribution Using the genomic platform developed in this collaboration, Ragoussis team has produced our analysis of their functional properties and conservation.
Impact Publications: 1. PMID: 17785540 2. PMID: 17785540 3. PMID: 16888364 4. Linnell J, Mott R, Field S, Kwiatkowski DP, Ragoussis J, Udalova IA. Quantitative high-throughput analysis of transcription factor binding specificities. Nucleic Acids Res. 2004 Feb 27;32(4):e44. PMID: 14990752 Grants: 1. FP7-EU grant 222008 "Genomic determinants of inflammation" 2. MRC collaborative grant G0700818
 
Description KM 
Organisation University of Oxford
Department Kennedy Institute of Rheumatology
Country United Kingdom 
Sector Academic/University 
PI Contribution Providing with expertise in gene regulation
Collaborator Contribution Expansion of our research scope into the field of extracellular matrix
Impact Publications: 1. Fui G. Goh, Anna M. Piccinini, Thomas Krausgruber, Irina A. Udalova and Kim S. Midwood. "Transcriptional regulation of the endogenous danger signal tenascin-C: a novel autocrine loop in inflammation". Journal of Immunology. In press. Grants: 1. Wellcome Trust grant application WT14378 "Controlling the expression of endogenous danger signals during inflammation" - decision pending
Start Year 2007
 
Description MP 
Organisation University of Cologne
Country Germany 
Sector Academic/University 
PI Contribution analysis of gene expression in provided cells
Collaborator Contribution providing with reagents (cells with targeted disruption of NFkB RelA)
Impact Grants: 1. FP7-EU grant 222008 "Genomic determinants of inflammation"
Start Year 2008
 
Description RC 
Organisation University of Oxford
Department Wellcome Trust Centre for Human Genetics
Country United Kingdom 
Sector Academic/University 
PI Contribution Setting up the question; generating experimental data
Collaborator Contribution bioinformatic support
Impact Publications: 1. PMID: 17785540 Grants 1. MRC grant application G0900965 - scores received 9, 9, 7, 7 - marked as of high priority - invited to re-submit in the next round 2. Wellcome Trust grant application WT090847 "THE ROLE OF TRANSPOSABLE ELEMENTS IN REGULATION OF NF-KAPPAB DEPENDENT GENES" - decision pending
Start Year 2006
 
Description RM 
Organisation University of Oxford
Department Wellcome Trust Centre for Human Genetics
Country United Kingdom 
Sector Academic/University 
PI Contribution Setting up the question; generating experimental data
Collaborator Contribution Computational support
Impact Grants: 1. MRC NIA G0501087 2. MRC collaborative grant G0700818
Start Year 2006
 
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