This project will investigate cutaneous antigen presenting cells by determining the function of CD1a, a non-polymorphic molecule expressed by Langerhans cells (LC). Similarities between CD1a and class I major histocompatibility (MHC) molecules suggest that CD1a can bind peptide antigen. Like class I MHC molecules, mediates cytotoxicity via the T-cell receptor (TCR) for antigen, but the role of added antigen in this interaction has not been determined. The first specific aim uses two strategies to determine whether CD1a specific T-cells recognize antigen bound to CD1a or recognize only distinct portions of CD1a. First, amino acids in the floor of the putative antigen binding groove formed by CD1a will be changed by site directed mutagenesis in order to displace any peptide normally bound int he groove. If changes of one or two amino acids alter the recognition of CD1a by multiple T-cell clones, then the hypothesis that TCRs interact with peptide antigen bound to CD1a will be supported since these amino acids should not be accessible for direct contact with the TCR. Next, T-cells from sensitized donors will be screened for recognition of specific antigens presented by cells expressing transfected CD1a but lacing classical MHC molecules. T-cells reacting only in the presence of both CD1a and antigen must be recognizing the antigen presented by CD1a. For the second specific aim, an estimate of the number of antigens which could be presented by CD1a will be made by amplifying and sequencing TCR mRNA sequences from CD1a specific T-cells. The third specific aim will test the ability of CD1a to present specific antigens in vivo by developing transgenic mice that express human CD1a controlled by murine class I MHC gene regulatory sequences. Since transgenic mice can be induced to express functional human class I MHC molecules, it is anticipated that transgenic human CD1a will also be functional. The mice will be immunized and tested for the ability to use CD1a as an antigen presenting molecule. Because presentation of specific antigens by non- polymorphic human molecules has not been previously demonstrated, these studies will provide new information about TCR recognition of antigens and about special properties of epidermal LC.
NIH ExPorter
NIH ExPorter