Clustering of T cell ligands on artificial APC membranes influences T cell activation and protein kinase C theta translocation to the T cell plasma membrane.
T cell activation is associated with active clustering of relevant molecules in membrane microdomains defined as the supramolecular activation cluster. The contact area between these regions on the surface of T cells and APC is defined as the immunological synapse. It has been recently shown that preclustering of MHC-peptide complexes in membrane microdomains on the APC surface affects the efficiency of immune synapse formation and the related T cell activation. Disruption of such clusters may reduce the efficiency of stimulation. We describe here an entirely artificial system for Ag-specific, ex vivo stimulation of human polyclonal T cells (artificial APC (aAPC)). aAPC are based on artificial membrane bilayers containing discrete membrane microdomains encompassing T cell ligands (i.e., appropriate MHC-peptide complexes in association with costimulatory molecules). We show here that preclustering of T cell ligands triggered a degree of T cell activation significantly higher than the one achieved when we used either soluble tetramers or aAPC in which MHC-peptide complexes were uniformly distributed within artificial bilayer membranes. This increased efficiency in stimulation was mirrored by increased translocation from the cytoplasm to the membrane of protein kinase theta, a T cell signaling molecule that colocalizes with the TCR within the supramolecular activation cluster, thus indicating efficient engagement of T cell activation pathways. Engineered aAPC may have immediate application for basic and clinical immunology studies pertaining to modulation of T cells ex vivo.
Giannoni
Francesca
F
Department of Medicine, University of California, San Diego, CA 92093, USA.
Barnett
Joellen
J
Bi
Kun
K
Samodal
Rodrigo
R
Lanza
Paola
P
Marchese
Patrizia
P
Billetta
Rosario
R
Vita
Randi
R
Klein
Mark R
MR
Prakken
Berent
B
Kwok
William W
WW
Sercarz
Eli
E
Altman
Amnon
A
Albani
Salvatore
S
D000938:Antigen-Presenting Cells / Q000276:immunology*; D001693:Biological Transport, Active; D015496:CD4-Positive T-Lymphocytes / Q000201:enzymology / Q000276:immunology; D002460:Cell Line; D002462:Cell Membrane / Q000201:enzymology; D006684:HLA-DR Antigens / Q000378:metabolism; D059811:HLA-DRB1 Chains; D019267:Hemagglutinin Glycoproteins, Influenza Virus; D006389:Hemagglutinins, Viral / Q000276:immunology; D006801:Humans; D066298:In Vitro Techniques; D007527:Isoenzymes / Q000378:metabolism*; D008024:Ligands; D008081:Liposomes; D008213:Lymphocyte Activation; D008567:Membranes, Artificial; D010446:Peptide Fragments / Q000276:immunology; D011493:Protein Kinase C / Q000378:metabolism*; D000074121:Protein Kinase C-theta; D011948:Receptors, Antigen, T-Cell / Q000378:metabolism; D013601:T-Lymphocytes / Q000201:enzymology* / Q000276:immunology*
D006684:HLA-DR Antigens; D059811:HLA-DRB1 Chains; D019267:Hemagglutinin Glycoproteins, Influenza Virus; D006389:Hemagglutinins, Viral; D007527:Isoenzymes; D008024:Ligands; D008081:Liposomes; D008567:Membranes, Artificial; D010446:Peptide Fragments; D011948:Receptors, Antigen, T-Cell; C081942:influenza hemagglutinin (307-319); C076233:PRKCQ protein, human; D011493:Protein Kinase C; D000074121:Protein Kinase C-theta