ALBERT

Notes: We established the organization of the AKR Qa region and determined the sequence of the 44 and Q5 genes. Restriction mapping and genomic Southern blot analysis revealed that the AKR strain codes for only three H-2K homologous genes in this region. The AKR Q5 gene is not homologous to the Q5 gene of the C57BL strain, but is presumably allelic to the Q5 gene isolated from Balb/c. The organization and structure of the AKR Qa family is virtually identical to the Qa genes of the C3H mouse. The AKR Q5 gene, in contrast to other H-2K homologous Qa region genes, codes for a typical transmembrane region, and upon transfection into BHK cells, a 1.6 kb Q5 transcript is detected.

Notes: The polymorphism of histocompatibility antigens is usually explained by classical Mendelian laws, which allow the inheritance of one allele per locus. Thus each individual would express two antigens for each locus of the pair of relevant homologous chromosomes, one from each parent.Another explanation is that the structural genes of all the different haplotypes are all in tandem. Regulatory genes would then determine the polymorphism by mimicking Mendelian inheritance–allowing only the relevant gene products for a particular haplotype to be expressed. If correct, one might expect the mechanism to fail sometimes, and silent genes of foreign haplotypes to be derepressed, allowing H-2 antigens of the wrong haplotype to become expressed.In the light of this hypothesis (Martin, 1975; Amos, 1971; Bodmer, 1975; Festenstein, 1978; Garrido et al., 1976a,b), the original findings in our laboratory of extra foreign ‘H-2-like’ determinants on tumour cells passaged in vitro and in vivo with and without virus are considered rather important (Festenstein, 1978; Garrido et al., 1976a,b); not only because they could lead to a better understanding of the genetic basis of MHS polymorphism, but also because of its implication for the surveillance of tumours (Garrido et al., 1976b; Gomard et al., 1977; Blank et al., 1976; Parmiani & Invernizzi, 1975) and microbially damaged cells (Doherty & Zinkernagel, 1975). These extra determinants were tested by various serological techniques and assays of cell mediated immunity.

Notes: Maize varieties with improved nitrogen(N)-use efficiency under low soil N conditions can contribute to sustainable agriculture. Tests were carried to see whether selection of European elite lines at low and high N supply would result in hybrids with differential adaptation to these contrasting N conditions. The objective was to analyze whether genotypic differences in N uptake and N-utilization efficiency existed in this material and to what extent these factors contributed to adaptation to low N supply. Twenty-four hybrids developed at low N supply (L × L) were compared with 25 hybrids developed at high N supply (H × H). The N uptake was determined as total above-ground N in whole plants, and N-utilization efficiency as the ratio between grain yield and N uptake in yield trials at four locations and at three N levels each. Highly significant variations as a result of hybrids and hybrids × N-level interaction were observed for grain yield as well as for N uptake and N-utilization efficiency in both hybrid types. Average yields of the L × L hybrids were higher than those of the H × H hybrids by 11.5% at low N supply and 5.4% at medium N level. There was no significant yield difference between the two hybrid types at high N supply. The L × L hybrids showed significantly higher N uptake at the low (12%) and medium (6%) N levels than the H × H hybrids. In contrast, no differences in N-utilization efficiency were observed between the hybrid types. These results indicate that adaptation of hybrids from European elite breeding material to conditions with reduced nitrogen input was possible and was mainly the result of an increase in N-uptake efficiency.

Notes: Cellular cytotoxicity experiments were done to test the potential of the extra H-2 antigenic specificities on the K36 spontaneous leukaemia as targets. In addition, experiments were performed to rule out the possibility that the target determinants could be normal cross-reacting alloantigens, e.g., T1, Qa, non-H-2 and previously undetected H-2 public specificities, which are more readily detectable on tumours than on normal cells.We used F1 hybrid mice, in which one parent was of the strain of origin of the tumour (K36) and the other parent of the B10 congenic series, i.e., (AKR x B10)F1. These cells were stimulated by lymphoid cells from other B10 congenic strains, B10.A and B10.D2, and tested against the test tumour K36 and several PHA blast controls. Several K36 sublines as well as a cloned line of K36 (K36.16) were used and significant cytotoxicity against. an H-2d-like target on these tumour cells was obtained. These data exclude the possibility of a corss-reactive alloantigen, e.g., undetected H-2 public specificity, or differentiation antigens. These results with the K36 tunour were consistent with our immunochemical studies (see Schmidt & Festenstein, 1980) and were confirmed and extended by cold target inhibition experiments. In these experiments, B10.BR cells were sensitized by B10.D2 lymphoid cells and tested against B10.D2 (51chromium-labelled PHA balsts). Two kinds of normal unlabelled lymph node suspensions as well as the K36.16 tumour cell suspension were used. Significant specific inhibition of between 19% and 40% was obtained using K36 and between 23% and 37% using B10.D2 (positive conrol). AKR cells (negative control) in contrast were unable to reduce the precentage specific cytotoxicity.Since it was already known that the H-2Kk gene products are missing from this tumour (Schmidt & Festenstein, 1980), it was of interest to test whether cytotoxic effectors directed against the H-2kk gene products were able to kill the K36 tumour. Accordingly, B10.D2 lymphoid cells were sensitized to B10.BR (C3H.0H) and B10.A targets, respectively, and tested against K36 and appropriate controls. Only weak killing was observed when sensitization was effected against the K end of the H-2k haplotype (i.e., using B10.A as the sensitizing cell) but strong and significant cytolysis was found when the sensitization was against the whole H-2k haplotype or against the H-2Dk gene product. These results were confirmed by cold target inhibition studies.These experiments provide further indications for the H-2d-like characteristics of these allodeterminants. We have already excluded some of the possible explanations for these findings (i.e., corss-reactions with H-2 and non-H-2 normal specificities). The cold target inhibition experiments rule out non-specific viral effects. Thus, we favour an alteration in regulatory genes leading to repression of the H-2Kk product and derepression of the H-2Dd product, but cannot formally rule out highly corss-reactive H-2-like virally encoded determinants.

Notes: From a series of mouse sarcomata, newly induced by Rous sarcoma virus (RSV), the DOH cell line was shown to lack expression of syngeneic H-2Kd and Dk antigens (No11 et al., 1986). It unexpectedly displayed determinants specific for H-2Kk molecules. Interferon treatment of DOH stimulated the expression of H-2Kk determinants and also the display of some, but not all, determinants of the syngeneic H-2Kd molecules. H-2Dk expression was not stimulated. Southern blot hybridization of genomic DNA digests from DOH cells confirmed the identity of the H-2K region with that from syngeneic C3H.OH liver cells, but also showed changes in the pattern of restriction fragments that contain class I genes from the D and Qa regions. These results suggest that aberrant MHC class I molecules that carry H-2Kd- and H-2Kk-like determinants are expressed by DOH sarcoma cells. These molecules may act as target antigens for turnour-specific cytotoxic T cells elicited by injection of DOH cells into syngeneic mice.

Notes: Expression of H-2 antigenic specificities on K36, a spontaneous leukaemia originating from AKR (H-2k) mice, was studied by serology and immunochemistry. Two ascites lines of the tumour, as well as a tissue culture adjusted and cloned tumour line, were used in these studies with similar results being obtained.K36 expresses on its cell surface D-region encoded H-2K antigens but does not express K-region encoded H-2K alloantigens. It also expresses on its cell membrane, H-2 specificities of foreign haplotypes not present on normal AKR lymphoid cells. The molecular basis of the H-2Dd specificity on K36 (H-2k was analysed by immunoprecipitation and polyacrylamide gel electrophoresis. The specificity was shown to be present on a glycoprotein of apparent molecular weight 45,000. However, antisera against the H-2Dd private specificity (H-2.4) precipitate additional glycoprotein of 45,000D and also 70,000D.In tryptic peptide maps of the isolated 45,000D fraction precipitated by anti-H-2.4 serum from radiolabelled K36 glycoprotein, all H-2Dd specific peptides were present in the same quantitative ratio. This is consistent with the structural identity of the foreign H-2Dd from the K36 tumour with normal H-2Dd and supports the hypothesis of a regulator system controlling the H-2 allelism. Under certain circumstances such a system could cause suppression of one and derepression of the other H-2 gene products.

Notes: The expression of H-2 antigenic specificities on Meth-A and meth-A-vaccinia were compared. Their ability to inhibit complement dependent cytotoxicity of 51Cr-labelled normal lymphoid target cells was tested by anti H-2 sera of restricted specificity. Normal lymphocytes positive or negative for a particular specificity, were used as controls. With this assay the expression of H-2-like specificities of foreign haplotypes on Meth-A cells passaged together with vaccinia virus was confirmed.In addition, using an immunofluorescence technique the expression of some foreign H-2 antigens on non-infected Meth-A tumour cells was shown. Anti-vaccinia virus sera were used to compare the reactivity of Meth-A and Meth-A-vaccinia. Unexpectedly, anti-vaccinia sera stained non-infected Meth-A and other tumour cells of different H-2 origin and absorption of these sera with normal Meth-A left little activity behind when testing against Meth-A-vaccinia.Finally, the rǒle that the foreign H-2 antigenic specificities present in Meth-A after vaccinia virus infection may have in the growth of these tumour cells in syngeneic recipients was studied. A powerful inhibitory effect was observed and regressor mice were found to be resistant to a challenge of non-infected meth-A. In contrast, vaccinia virus was irrelevant to the rate of growth of P815 Y in DBA/2 mice.These results are discussed in relation to the origin of the foreign H-2 antigens on tumour cells after virus infection (derepression) and the role that this new antigenicity could have in immunopotentiation.