studproj023

Localization of costimulatory receptor CD6 in the immunological synapse

Student project 3-9 months (both bachelor and master project is possible)

Department of Tumor Immunology, NCMLS, UMC St. Radboud, Nijmegen, in collaboration with Nanobiophysics, MIRA, University of Twente, Enschede

Theoretical background:

The immune system protects our body against disease caused by microbial infection and cancer. A part of this defence system are the T cells, which can specifically attack and kill pathogens like, virus infected cells and cancer cells. To be activated, T cells need to recognize parts of these pathogens (antigens) that are presented by specialized antigen presenting cells (APCs). When the T cell receptor (TCR) on a T cell recognizes antigen presented on major histocompatibility complex (MHC) molecules by an antigen presenting cell a specialized contact between these two cells, the immunological synapse (IS), is formed.

The mature immunological synapse is composed of a central region (central supramolecular activation cluster, cSMAC) and a peripheral SMAC (pSMAC). The cSMAC is enriched in TCR molecules and costimulatory molecules on the T cell site and MHC-antigen complexes on the APC site. The pSMAC contains the integrin LFA-1 on the T cell side and its major ligand intracellular adhesion molecule-1 (ICAM-1) on the APC site (figure 1).

Figure 1. The Immunological synapse (Huppa & Davis, 2003)

In this project you are going to investigate the costimulatory molecule CD6. CD6 is expressed by T cells, and through its interaction with its ligand ALCAM, which is expressed on antigen presenting cells, it is thought to act both as a costimulatory molecule and an adhesion molecule, involved in T cell activation [3-5].

Project outline:

In this project you are going to investigate where CD6 is localized in the immunological synapse. A tool which is often used to study the IS is an artificial lipid bilayer, which is an artificially prepared bilayer of fatty acids on a glass support in which proteins can be incorporated. In this way the APC site of the IS can be mimicked on a glass surface. In this project you will prepare artificial lipid bilayers in which we incorporate antibodies that mimic the peptide-MHC complex and ALCAM, the ligand for CD6. We seed T cells onto these bilayers and let the cells form a synapse. Subsequently we will image the T cells to see how the synapse is organized, with specific interest in the localization of CD6.

The first part of this project will be to set up the preparation of lipid bilayers and the incorporation of antibodies and ALCAM into these bilayers. For this purpose we cooperate with the university of Twente, where they have experience with this method. In the second part of the project you will use immunohistochemistry and confocal and TIRF microscopy to study the localization of CD6 in the mature synapse. If the project lasts longer (6-9 months) you will also perform live cell imaging during synapse formation to investigate the dynamics of CD6 during synapse formation.

Most part of this project will be performed at the Tumor Immunology department in Nijmegen, but some experimental work will be done at the Nanobiophysics department in Twente

Techniques:

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Preparation of lipid bilayers

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Sterile cell culture

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Immunohistochemistry

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Confocal microscopy

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TIRF microscopy

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Live cell imaging

Literature:

1. Grakoui, A., et al., The immunological synapse: a molecular machine controlling T cell activation. Science, 1999. 285(5425): p. 221-7.

2. Dustin, M.L., Insights into function of the immunological synapse from studies with supported planar bilayers, in Immunological Synapse. 2010. p. 1-24.

3. Zimmerman, A.W., et al., Long-term engagement of CD6 and ALCAM is essential for T-cell proliferation induced by dendritic cells. Blood, 2006. 107(8): p. 3212-20.

4. Gimferrer, I., et al., Relevance of CD6-mediated interactions in T cell activation and proliferation. J Immunol, 2004. 173(4): p. 2262-70.

5. Gimferrer, I., et al., The lymphocyte receptor CD6 interacts with syntenin-1, a scaffolding protein containing PDZ domains. J Immunol, 2005. 175(3): p. 1406-14.

For more information:

Marjolein Meddens, PhD student

Department of Tumor Immunology, NCMLS, UMC St. Radboud, Nijmegen

m.meddens@ncmls.ru.nl

Tel: 024 3653717

Alessandra Cambi, Associate Professor

a.cambi@ncmls.ru.nl

Nanobiophysics Department, MIRA, University of Twente, Enschede