Molecular insight into the mechanism of antigen presentation: Role of transmembrane domains in MHC Class-II assembly

Lead Research Organisation: University of Warwick
Department Name: Chemistry

Abstract

The acquired immune system enables an organism to defend itself against assaults from invading pathogens, such as bacteria or viruses, by selectively attacking infected or cancerous cells. In order to locate and destroy these infected cells, a complex symphony of events takes place which is directed by the most important of immune cells, the helper T-cells. Helper T-cells respond to the presence of a foreign pathogen by releasing a range of chemicals that regulate all aspects of the immune system. Helper T-cells are themselves controlled by the Class II Major Histocompatibility Complex (MHC Class II), a complex composed of two membrane proteins found on the surface of antigen presenting cells. MHC Class II is responsible for binding and presenting small peptide antigens, derived from foreign proteins, on the surface of cells. This process of antigen presentation by MHC Class II activates helper T-cells and triggers an immune response.

The failure of MHC Class II to activate helper T-cells in a normal way results in a severe collapse of the immune system. This is a key factor in autoimmune diseases as well as other serious human diseases. Therefore, correct transport of peptide-loaded Class II molecules to the cells surface is essential to healthy immune response. This transport is initiated by a chaperone molecule called the MHC Class II-associated invariant chain (Ii). A trimer of Ii binds three Class II heterodimers in the endoplasmic reticulum to form a nine-chain complex, and only as part of this complex are Class II molecules targeted to the cells surface. The assembly of the nine-chain complex is therefore an essential first step in antigen presentation.

Assembly of the nine-chain complex involves a large number of specific protein-protein interactions, and recent evidence indicates that specific interactions between the transmembrane domains of all three proteins are of significant biological importance. The focus of this research is investigation of the structures and interactions of these domains in order to better understand the assembly of the complex and its role in cellular processes and disease. Work at this stage is primarily fundamental, however the interactions we will characterise can be used in design of molecules that target TM domain interactions and could be applied to design of drugs and therapeutics that target the large number of autoimmune diseases and cancers in which MHC Class II and MHC Ii are implicated.

Technical Summary

The Class II Major Histocompatibility Complex (MHC Class II) is a membrane protein heterodimer responsible for activating the most important cells in the acquired immune system, CD4+ helper T-cells. Helper T-cells recognise foreign peptides presented by MHC Class II on the surface of antigen presenting cells, and respond by releasing chemicals that regulate all aspects of the immune system. Therefore, correct transport of peptide-loaded MHC Class II to the cells surface is essential for healthy immune response. This transport is initialised by a membrane protein chaperone called the MHC Class II-associated invariant chain (Ii). In the endoplasmic reticulum (ER), a trimer of Ii binds three Class II heterodimers to form a nine-chain complex. Only as part of this complex will MHC Class II be released from the ER, protected from degradation, and targeted to the cells surface. The assembly of the nine-chain complex is an essential first step in antigen presentation and is the focus of this research.

The assembly of the nine-chain complex involves a large number of specific protein-protein interactions, and recent evidence indicates that the transmembrane (TM) domains of all three proteins are of significant biological importance. Despite this fact, the transmembrane domains of these proteins have been severely overlooked in MHC Class II studies. A detailed structural knowledge of key interactions in the membrane is the next step in understanding the function and fate of this complex. The objective of this research is to systematically investigate TM helix-helix interactions involved in assembly of the nine-chain complex using state-of-the-art biochemical, biophysical and structural methods, including NMR spectroscopy. We will determine structural features that drive association and investigate the role of known interaction motifs. The end product of this research will be a solution structure of the Ii-TM trimer, dissociation constants for all transmembrane domain interactions in the nine-chain complex, and a detailed model revealing specific residues/motifs that control formation and stabilisation of the complex. The interactions we will characterise in this study have broad ranging implications for the design of molecules that target TM domain interactions and, further downstream, could be applied to design of drugs and therapeutics that target the large number of autoimmune diseases and cancers in which MHC Class II and MHC Ii are implicated.
 
Description British Heart Foundation Project Grant
Amount £243,000 (GBP)
Organisation British Heart Foundation (BHF) 
Sector Charity/Non Profit
Country United Kingdom
Start 03/2009 
End 03/2012
 
Description Warwick Research and Development Fund
Amount £14,000 (GBP)
Organisation University of Warwick 
Sector Academic/University
Country United Kingdom
Start 01/2010 
End 06/2010
 
Title AUC methods established 
Description The sample preparation, measurement, and data processing protocols required to perform and interpret analytical untracentrifugation experiments for determination of oligomeric state were established, as were long term collaborations with labs at Nottingham, Birmingham, and Oxford Universities to obtain high quality AUC data, as a direct result of this funding. 
Type Of Material Improvements to research infrastructure 
Year Produced 2008 
Provided To Others? No  
Impact The ability to obtain high quality AUC data has and will continue to allow us to more quantitatively understand the stoichiometry and binding energies of the systems we study. The technique is unique in its ability to provide such information, and invaluable to our work. 
 
Title FRET methods established 
Description The sample preparation, purification, and fluorescence measurement protocols required to perform Forster resonance energy transfer (FRET) measurements on TM helices within my research team were established as a direct result of this work. 
Type Of Material Improvements to research infrastructure 
Year Produced 2008 
Provided To Others? No  
Impact This method is very broadly applicable and will be applied to other research projects taking place within my group. Development of this technique will support our research, providing another tool to allow us to better understand these difficult to study systems. 
 
Title GALLEX methods established 
Description The plasmids and cells lines required to perform the GALLEX assay for TM helix-helix homo- and hetero-oligomerization are now well established within my research lab, as are the protocols required to perform measurement of the all controls and the b-gal repression. This is the first time these measurements have been successfully performed outside of two other leading labs, and our success in getting this method to work is a direct result of the funding received by the MRC. 
Type Of Material Improvements to research infrastructure 
Year Produced 2008 
Provided To Others? Yes  
Impact This method is also very broadly applicable and will be applied to other research projects taking place within my group. Development of this technique will support our research, providing another tool to allow us to better understand these difficult to study systems. 
 
Title New model of MHC II-Ii complex 
Description Cannot disclose details as data have not been made public (publication in preparation for invited submission Feb. 2010) 
Type Of Material Model of mechanisms or symptoms - human 
Year Produced 2010 
Provided To Others? Yes  
Impact Will result in publications as well as new research directions 
 
Description Mock Panel Meeting 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? Yes
Geographic Reach Regional
Primary Audience Schools
Results and Impact We hosted a mock panel meeting for secondary school children, in order for them to better understand the way research is conceived, communicated and funded in today's society. In this activity, students were given a problem near our research interests, they research the problem, and they have to defend their research in a panel. During this process, we (as academics) were asked to join one group of students and help them through the tasks, as well as describing our work to them in everyday terms. I discussed the work I am doing with MRC funding.

Encouraging the next generation of scientific researchers
Year(s) Of Engagement Activity 2008