The manner in which alloreactive T lymphocytes recognize determinants on class I H-2 molecules is not understood. Analysis of the reactivity of cytotoxic T lymphocytes (CTL) with cells transformed with hybrid ( or exon-shuffled) H-2 class I genes, has suggested that only the alpha-2 domains express determinants recognized by CTL. I have isolated a somatic cell mutant which expresses a Dd molecule that lacks a serological determinant residing in the alpha-3 domain but retains all of the alpha-1 and alpha-2 determinants. Sequencing of the alpha-3 domain of the mutant Dd molecule shows that there is a single point mutation resulting in the substitution of a lysine for the glutamic acid at position 227. In contrast to what is predicted from the exonshuffling experiments, this Dd mutant cell line is not killed by clone lines of anti-Dd reactive CTL. I therefore propose that integrity of at least some residues in the alpha-3 domain is required for CTL recognition because they contribute to a conserved (or monomorphic), determinant that, in cellular recognition, defines the molecule as a class I molecule. This application seeks to perform oligonucleotide directed mutagenesis in the alpha-3 domain of the cloned H-2Dd gene and analyze CTL recognition of cells transfected with the mutant genes. These studies should provide insights as to the role of the alpha-3 domain of class I molecules in CTL recognition.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Research Project (R01)
Project #
5R01AI028115-02
Application #
3142446
Study Section
Immunobiology Study Section (IMB)
Project Start
1988-09-01
Project End
1991-08-31
Budget Start
1989-09-01
Budget End
1990-08-31
Support Year
2
Fiscal Year
1989
Total Cost
Indirect Cost
Name
National Jewish Health
Department
Type
DUNS #
City
Denver
State
CO
Country
United States
Zip Code
80206
Smith, P A; Potter, T A (1998) Alloreactive T cells that do not require TCR and CD8 coengagement are present in naive mice and contribute to graft rejection. J Immunol 160:5382-9
Smith, P A; Brunmark, A; Jackson, M R et al. (1997) Peptide-independent recognition by alloreactive cytotoxic T lymphocytes (CTL). J Exp Med 185:1023-33
Cook, J L; Potter, T A; Bellgrau, D et al. (1996) E1A oncogene expression in target cells induces cytolytic susceptibility at a post-recognition stage in the interaction with killer lymphocytes. Oncogene 13:833-42
Sawyer, R T; Drevets, D A; Campbell, P A et al. (1996) Internalin A can mediate phagocytosis of Listeria monocytogenes by mouse macrophage cell lines. J Leukoc Biol 60:603-10
Zwickey, H L; Potter, T A (1996) Peptide epitopes from noncytosolic Listeria monocytogenes can be presented by major histocompatibility complex class I molecules. Infect Immun 64:1870-2
Knall, C; Smith, P A; Potter, T A (1995) CD8-dependent CTL require co-engagement of CD8 and the TCR for phosphatidylinositol hydrolysis, but CD8-independent CTL do not and can kill in the absence of phosphatidylinositol hydrolysis. Int Immunol 7:995-1004
Drevets, D A; Sawyer, R T; Potter, T A et al. (1995) Listeria monocytogenes infects human endothelial cells by two distinct mechanisms. Infect Immun 63:4268-76
Knall, C; Ingold, A; Potter, T A (1994) Analysis of coreceptor versus accessory molecule function of CD8 as a correlate of exogenous peptide concentration. Mol Immunol 31:875-83
Connolly, J M; Hansen, T H; Ingold, A L et al. (1990) Recognition by CD8 on cytotoxic T lymphocytes is ablated by several substitutions in the class I alpha 3 domain: CD8 and the T-cell receptor recognize the same class I molecule. Proc Natl Acad Sci U S A 87:2137-41