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Activating Receptors of Natural Killer cells

Reference	BBS/E/B/32001153
Principal Investigator / Supervisor	Dr James Ross Miller
Co-Investigators / Co-Supervisors
Institution	Babraham Institute
Department	Babraham Institute Department
Funding type	Research
Value (£)	137,151
Status	Completed
Type	Institute Project
Start date	01/01/1999
End date	31/03/2000
Duration	15 months

Abstract

Natural Killer (NK) cells express surface receptors which recognise MHC Class 1 molecules and inhibit NK cell cytotoxicity. The primary structure of inhibitory receptors has now been described in considerable detail. Activating receptors appear to show considerable, but not total sequence homology to inhibitory receptors. In humans, killer cell activation has been shown to be dependent on association between an activating receptor and DAP-12, a small disulphide bonded dimeric protein which contains a tyrosine based activation motif. A DAP-12 cDNA clone can be marked with a FLAG epitope permitting detection with an anti-FLAG antibody. When human cells are transfected with DAP-12/Flag, expression of FLAG is not detected on the cell surface, but can be detected by Western blotting. However, when DAP-12/Flag cells are transfected with an activating receptor, the FLAG epitope is expressed on the cell surface. The intention is thus to use a stably transformed DAP12- Flag cell line to search for activating receptors by transformation with cDNA libraries. Positive cells will be detected by flow cytometry, episomal DNA isolated, amplified in bacteria, then reselected in human cells. Several such cycles should result in isolation of a cDNA encoding a protein which binds DAP-12 and can cause NK cell activation. This strategy is open-ended, with no assumptions being made about the activating receptor, other than that it can interact with DAP-12.This work is especially relevant to the H&LS Foresight specific priority for Manipulation of the Immune System. This project will permit isolation and characterisation of novel NK cell activating receptors. Further study of these receptors will provide greater detail of the process of NK cell activation. In turn this will increase our understanding of mechanisms for evasion of NK cell killing. Receptor molecules and pathways in both activation and inhibition of NK cells will be likely targets for therapeutic intervention. Consequently, detailed knowledge of the structure of these molecules will be a pre-requisite for design of therapeutic reagents.

Summary

unavailable

Committee	Closed Committee - Biochemistry & Cell Biology (BCB)
Research Topics	X – not assigned to a current Research Topic
Research Priority	X – Research Priority information not available
Research Initiative	X - not in an Initiative
Funding Scheme	X – not Funded via a specific Funding Scheme

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