Somatic H‐2K^k variants reveal nonidentity of serological and cytotoxic T cell‐defined K^k determinants

Abstract

The relationship of serologically defined determinants to determinants recognized by cytotoxic T cells on molecules encoded by the Kk gene of the murine major histocompatibility complex (H-2) has been analyzed. For this purpose we used three somatic variants of a Kk-expressing lymphoma line lacking individual determinants of the Kk molecule, as defined by monoclonal antibodies (mAb), as target cells for Kk-specific alloreactive and Kk-restricted cytotoxic T lymphocytes (CTL) cloned by limiting dilution. Neither alloreactive nor fluorescein isothiocyanate, influenza- or Newcastle disease virus-specific Kk-restricted CTL clones were found to distinguish between variants and wild type cells, indicating that the serologically defined determinants lost by the variants were not essential for antigen recognition of CTL with these specificities. On the other hand, two of the variants lacking either one of a pair of serological determinants were discriminated from Kk wild type cells by about 40% of Kk-restricted, trinitrophenol (TNP)-specific CTL clones. The third variant, lacking both of the determinants, however, was lysed by all CTL clones to the same extent as wild type cells. From these results we conclude that the determinants restricting the TNP-specific CTL were also not identical with those defined by mAb. In experiments performed to optimize the conditions for the limiting dilution analysis we found that the specificity of the CTL stimulation was strongly dependent on the concentration of T cell growth factor (interleukin 2) in the cultures during CTL stimulation. High concentrations of IL2 resulted in a drastic increase in the frequency of CTL clones. Part of these clones, however, were found not to be specific for antigens present on the stimulator cells.

Allospecific cytolytic T lymphocytes recognize conformational determinants on hybrid mouse transplantation antigens.

“…For example, CTL may recognize an amino acid sequence located at the junction of the ~ and ~z domains where some polymorphism has been shown (1). Although this hypothesis is consistent with other studies suggesting that T and B cells recognize distinct antigenic determinants (5,6), the folding of the MHC domains and the location of the MHC hypervariable regions to other areas of the molecule might argue against allorecognition of this part of the molecule. An alternative hypothesis that would lead to differences in the T cell and antibody repertoires was recently proposed by Gress and Hodes (24).…”

“…After incubation with mAb at 4°C for 45 rain, the cells were washed and stained with 25 #1 of fluorescein-conjugated coat F(ab')~ anti-mouse IgG (Cappel Laboratories, Cochran-~ o~2 O~3 M I CELL LINE .~' "1 I I I I Generation of K b-and Db-specific CTL Populations. Primary and secondary CTL populations were generated as previously described (6). Briefly, 5 X 106 responder cells were mixed with 2-4 x 106 irradiated (2,00 rad) stimulator cells in 2 ml final volume complete media (RPMI 1640 supplemented with 10% FCS, sodium pyruvate, nonessential amino acids, glutamine, penicillin-streptomycin, and 2-mercaptoethanol).…”

Modification of tumor cells by a low dose of Newcastle Disease Virus.

“…Hence, we first established a system for the analysis of virus-specific cytotoxic T lymphocytes in DBN2 mice. The conditions were adapted to those described by Vohr et al [35] where a frequency of H-2Kkrestricted NDV-specific (strain Italy) CTL of 1/80000 to 1/100 000 was determined. In DBN2 mice immunized with the rather high dose of 8OOO HU NDV strain Ulster we found a frequency of NDV-specific CTL of 1/32531 (Fig.…”

Somatic variation of H-2Kk expression and structure in a T-cell lymphoma: instability, stabilization, high production and structural mutation.

“…It will be of interest to see whether the mutations represent recombinatorial events resembling those recently identified in germ line mutants of class I antigens Weiss et al, 1983;Schulze et al, 1983). The structural variants have been used to study the relationship between serological Kk determinants and those defined by CTLs (Vohr et al, 1983;Wraith et al, 1983). Analyses with CTLs cloned by limiting dilution and a CTL-line revealed that the serological Kk determinants lost from the structural variants are not recognized by alloreactive CTLs and do not have a function in the restriction of CTL responses to a variety of foreign antigens.…”

“…This interpretation is strongly supported by results of functional analyses with CTLs. Neither alloreactive Kk-specific CTLs nor Kk-restricted CTLs distinguished between HK22.S2.L1 and Kk wild-type cells (Vohr et al, 1983;Wraith et al, 1983).…”