Three surface molecules of mouse CD8+ dendritic cells (DCs), also found on the equivalent human DC subpopulation, were compared as targets for Ab-mediated delivery of Ags, a developing strategy for vaccination. For the production of cytotoxic T cells, DEC-205 and Clec9A, but not Clec12A, were effective targets, although only in the presence of adjuvants. For Ab production, however, Clec9A excelled as a target, even in the absence of adjuvant. Potent humoral immunity was a result of the highly specific expression of Clec9A on DCs, which allowed longer residence of targeting Abs in the bloodstream, prolonged DC Ag presentation, and extended CD4 T cell proliferation, all of which drove highly efficient development of follicular helper T cells. Because Clec9A shows a similar expression pattern on human DCs, it has particular promise as a target for vaccines of human application.

The immune system must distinguish viable cells from cells damaged by physical and infective processes. The damaged cell-recognition molecule Clec9A is expressed on the surface of the mouse and human dendritic cell subsets specialized for the uptake and processing of material from dead cells. Clec9A recognizes a conserved component within nucleated and nonnucleated cells, exposed when cell membranes are damaged. We have identified this Clec9A ligand as a filamentous form of actin in association with particular actin-binding domains of cytoskeletal proteins. We have determined the crystal structure of the human CLEC9A C-type lectin domain and propose a functional dimeric structure with conserved tryptophans in the ligand recognition site. Mutation of these residues ablated CLEC9A binding to damaged cells and to the isolated ligand complexes. We propose that Clec9A provides targeted recruitment of the adaptive immune system during infection and can also be utilized to enhance immune responses generated by vaccines.

Multipin peptide synthesis has been employed to produce biotinylated 11-mer phosphopeptides that account for every tyrosine residue in insulin receptor substrate-1 (IRS-1) and the cytoplasmic domains of the insulin-, epidermal growth factor-, platelet-derived growth factor-and basic fibroblast growth factor receptors. These phosphopeptides have been screened for their capacity to bind to the SH2 domains of Shc and Grb

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