Fluorescent tagged NK cell receptors expressed in vivo using retrogenic
mice to study the delivery of signals in NK cells
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1
Imperial College London, Department of Immunology, United Kingdom
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2
Imperial College London, Department of Life Sciences, United Kingdom
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3
Imperial College London, Division of Immunology and Inflammation, United Kingdom
NK cells display both activating and inhibitory receptors which recognize
different ligands, and it is the integration of signals from these two classes of
receptors that ultimately determines NK cell effector activity. Nevertheless,
how exactly the signal integration occurs is still not well understood.
Our approach is to employ retrogenic mice to generate authentic primary NK
cells expressing two fluorescent tagged receptors, NKG2D (short or long
isoform) and Ly49A, prototypical activating and inhibitory NK cell receptors,
respectively. The aim is to use NKG2D-GFP/Ly49A-RFP Forster resonance
energy transfer (FRET) to study the co-localisation of these receptors in
synapses at NK-target cell-cell contact interfaces. Also, we will test the
consequences of altering ligand dimensions in the synapse formation and
NK cell activation.
Our results show that GFP-NKG2D (short and long isoform) interacts with
both adaptor molecules DAP10 and DAP12 in vitro, allowing efficient cell
surface expression of fluorescent NKG2D. Also, we reconstituted irradiated
C57BL/6 recipients with GFP-NKG2D transduced bone marrow cells from
NKG2D-deficient C57BL/6 donor mice. Two weeks post reconstitution, we
detected cell surface expression of GFP-NKG2D predominantly in NK cells.
The functionality of the fluorescent receptors and their interaction with the
adaptor proteins DAP10 and DAP12 are now being investigated in NK cells
generated in vivo. Data on the functionality of the fluorescent receptors and
the generation of NK cells in retrogenic mice will be presented.
Keywords:
NK cells,
innate immunity,
NKG2D,
ly49a,
retrogenic mice
Conference:
15th International Congress of Immunology (ICI), Milan, Italy, 22 Aug - 27 Aug, 2013.
Presentation Type:
Abstract
Topic:
Immune receptors and signaling
Citation:
Tomaz
D,
Guerra
N,
Dyson
J and
Gould
K
(2013). Fluorescent tagged NK cell receptors expressed in vivo using retrogenic
mice to study the delivery of signals in NK cells.
Front. Immunol.
Conference Abstract:
15th International Congress of Immunology (ICI).
doi: 10.3389/conf.fimmu.2013.02.01194
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Received:
22 Jul 2013;
Published Online:
22 Aug 2013.
*
Correspondence:
Dr. Keith Gould, Imperial College London, Department of Immunology, London, United Kingdom, k.gould@ic.ac.uk