INDUCTION OF T LYMPHOCYTES FROM PRECURSOR CELLS IN VITRO BY A PRODUCT OF THE THYMUS

A product of mouse thymus induces cells found in the spleen and bone marrow of nu/nu mice (which lack a thymus), and in 14-day embryonic mouse liver, to differentiate in vitro into T lymphocytes (defined as cells bearing TL and Thy-1 antigens). Thus the in vitro T lymphocyte induction mechanism acts on a cell that is antecedent to any thymus-mediated process

ANTIGENS OF LEUKEMIAS INDUCED BY NATURALLY OCCURRING MURINE LEUKEMIA VIRUS: THEIR RELATION TO THE ANTIGENS OF GROSS VIRUS AND OTHER MURINE LEUKEMIA VIRUSES

Leukemias can be induced in W/Fu inbred rats by neonatal inoculation of normal thymus cells of C58 mice. These leukemias are not transplantable to C58 mice or to adult W/Fu rats, but they can be kept in passage in W/Fu rats aged 0 to 7 days. Adult W/Fu rats inoculated repeatedly with these isogenic leukemias produce cytotoxic and precipitating antibodies. These antisera are of particular value in the analysis of the antigens of leukemia cells and of leukemia viruses because their mode of preparation precludes the formation of antibody against any normal constituents of the cell. Analysis based on the cytotoxic test indicates the presence of 2 distinct cell surface antigens in leukemias induced by Passage A Gross virus or occurring spontaneously in mice of high-incidence strains. All leukemias and other tissues known to contain G (Gross) leukemia antigen have both determinants, but certain leukemias of low-incidence strains have only 1 of them and so were previously classified G-. Immunoprecipitation with these antisera reveals the presence of a cellular antigen common to G+ cells and absent from G- cells; the same antigen can be demonstrated in ether-treated Gross virus, but not in intact virus. This antigen is present also in ether-treated preparations of the Friend, Moloney, and Rauscher leukemia viruses, but not in Bittner (mammary tumor) virus. Thus it may be regarded as a group-specific antigen of murine leukemia viruses, in contrast to the type-specific cellular antigens demonstrable by the cytotoxic test. Four additional antigens associated with leukemias induced by wild-type Gross virus have been demonstrated by immunoprecipitation, but their relation to viral and cellular antigens has not been determined

THE GIX SYSTEM : A CELL SURFACE ALLO-ANTIGEN ASSOCIATED WITH MURINE LEUKEMIA VIRUS; IMPLICATIONS REGARDING CHROMOSOMAL INTEGRATION OF THE VIRAL GENOME

This report concerns a cell surface antigen (GIX; G = Gross) which exhibits mendelian inheritance but which also appears de novo in cells that become productively infected with MuLV (Gross), the wild-type leukemia virus of the mouse. In normal mice, GIX is a cell surface allo-antigen confined to lymphoid cells and found in highest amount on thymocytes. Four categories of inbred mouse strains can be distinguished according to how much GIX antigen is expressed on their thymocytes. GIX- strains have none; in the three GIX+ categories, GIX3, GIX2, and GIX1, the amounts of GIX antigen present (per thymocyte) are approximately in the ratios 3:2:1. A study of segregating populations derived mainly from strain 129 (the prototype GIX3 strain) and C57BL/6 (the prototype GIX- strain) revealed that two unlinked chromosomal genes are required for expression of GIX on normal lymphoid cells. The phenotype GIX+ is expressed only when both genes are present, as in 129 mice. C57BL/6 carries neither of them. At one locus, expression of GIX is fully dominant over nonexpression (GIX fully expressed in heterozygotes). At the second locus, which is linked with H-2 (at a distance of 36.4 ± 2.7 units) in group IX (locus symbol GIX), expression is semidominant (50% expression of GIX in heterozygotes); gene order T:H-2:Tla:GIX. As a rule, when cells of GIX- mice or rats become overtly infected with MuLV (Gross), an event which occurs spontaneously in older mice of certain strains and which also commonly accompanies malignant transformation, their phenotype is converted to GIX+. This invites comparison with the emergence of TL+ leukemia cells in TL- mouse strains which has been observed in previous studies and which implies that TL- → TL+ conversion has accompanied leukemic transformation of such cells. So far the only example of GIX- → GIX+ conversion taking place without overt MuLV infection is represented by the occurrence of GCSA-:GIX+ myelomas in BALB/c (GCSA:GIX-) mice. Unlike the other Gross cell surface antigen described earlier, GCSA, which is invariably associated with MuLV (Gross) infection and never occurs in its absence, GIX antigen sometimes occurs independently of productive MuLV infection; for example, thymocytes and some leukemias of 129 mice are GCSA-:GIX+, and MuLV-producing sarcomas may be GCSA+:GIX-. The frequent emergence of cells of GIX+ phenotype in all mouse strains implies that the structural gene coding for GIX antigen is common to all mice. There is precedent for this in the TL system, in which two of the Tla genes in linkage group IX appear to be ubiquitous among mice, but are normally expressed only in strains of mice carrying a second (expression) gene. It is not yet certain whether either of the two segregating genes belongs to the MuLV genome rather than to the cellular genome. This leaves the question whether MuLV may have a chromosomal integration site still debatable. But there is a good prospect that further genetic analysis will provide the answer and so elucidate the special relationship of leukemia viruses to the cells of their natural hosts

ISOANTIGENS OF THE H-2 AND Tla LOCI OF THE MOUSE : INTERACTIONS AFFECTING THEIR REPRESENTATION ON THYMOCYTES

H-2 and TL isoantigens of the mouse are specified by the closely linked genetic loci H-2 and Tla. A. study of their representation on thymocytes was performed in order to reveal any interactions between the determinant genes or their products affecting the synthesis or disposition of these components of the thymocyte surface. The method employed was quantitative absorption of cytotoxic antibody by viable thymocytes. The phenotypic expression of TL antigens was found to reduce the demonstrable amount of certain H-2 antigens to as little as 34% of the quantity demonstrable on TL- thymocytes. A reduction was observed in all three H-2 types tested, (H-2b, H-2a, and H-2k). As antigenic modulation (change of TL phenotype from TL+ to TL-, produced by TL antibody) is known to entail a compensatory increase in H-2(D) antigen, it is concluded that the TL phenotype, rather than the Tla genotype, influences the surface representation of H-2 antigens. The two known TL+ phenotypes of thymocytes (TL.2 and TL.1,2,3) depress H-2 equally. The H-2 specificities affected are those determined by the D end of the E-2 locus, which is adjacent to Tla; antigens of the K end, which is distal to Tla, are not depressed. The reduction of demonstrable H-2 antigen on the thymocytes of TL+ x TL- progeny is half that of thymocytes of TL+ x TL+ progeny and the reduction affects equally the products of both H-2 alleles (cis and trans in relation to Tla), indicating that the mechanism of H-2 reduction by TL is extrachromosomal. Whether it involves diminished synthesis of H-2 or steric masking by TL at the cell membrane is unknown, but in either case the reciprocal relation of TL and H-2(D) antigens implies that they probably occupy adjacent positions on thymocytes and that the gene order, H-2(K): H-2(D):Tla is reflected in cell surface structure. Extrachromosomal interaction, apparently involving control of synthesis, occurs also within the TL system of antigens. Thymocytes of TL.2 x TL.1,2,3 progeny express the full homozygous quantity of antigens TL.1 and TL.3 (but not of TL.2), in contrast to the half-quantity present in thymocytes of TL- x TL.1,2,3 progeny. Another example of interaction is implicit in the finding that thymocytes of TL-1,2,3 x TL.1,2,3 progeny have more TL.2 antigen than thymocytes of TL.2 x TL.2 progeny, but in this instance there is nothing to indicate whether the mechanism is chromosomal or extrachromosomal. Thus the quantitative surface representation of at least some H-2 and TL antigens is influenced by the cellular complement of H-2:Tla genes as a whole. Comparison of H-2 heterozygous thymocytes with H-2 homozygous thymocytes in quantitative absorption tests shows (a) more than the expected 50% of each parental-type H-2 antigen on heterozygous cells, and (b) a greater suppression of H-2 by TL in H-2 heterozygotes in comparison with H-2 homozygotes. Both results may be explained on the basis of differences in the density of H-2 antigenic sites and consequent differences in the efficiency of absorption of H-2 antibody. These considerations may be useful in other contexts, e.g. in estimating the representation of Rh antigens on the red cells of human subjects homozygous and heterozygous for Rh components

PRODUCTION OF TL ANTIBODY BY MICE IMMUNIZED WITH TL- CELL POPULATIONS : A POSSIBLE ASSAY FOR THYMIC HORMONE

TL- mice make TL antibody when immunized with spleen or bone marrow cells from TL+ donors, despite the fact that these cells do not express TL antigen. This has been shown to depend on maturation of TL- precursors, contained in the inoculum, into TL+ cells under influence of the recipient's thymus; the differentiated TL+ cells then evoke production of TL antibody

SURFACE ALLOANTIGENS OF PLASMA CELLS

A serological study of immunoglobulin-forming cells of the mouse, normal and malignant, shows that they lack all known surface differentiation antigens of the thymocyte-lymphocyte axis: TL, θ, Ly-A, Ly-B, and MSLA. Two systems of normal alloantigens are expressed on these cells, H-2 and a new system named PC. The gene Pca (Plasma cell antigen) which specifies PC.1 alloantigen segregates as a mendelian dominant not closely linked with H-2. This cell surface antigen is absent from thymocytes, leukemias, and very probably from thymus-derived lymphocytes also; it is present on cells of the liver, kidney, brain, and lymph nodes as well as on hemolytic plaque-forming cells of the spleen, and on myelomas. So PC.1 is properly classified as a differentiation alloantigen. The strain distribution of PC.1 does not conform to that of any known immunoglobulin allotype or cell surface alloantigen previously described. Thus the cell surface antigens of immunoglobulin-producing cells are clearly different from those of cells belonging to the thymocyte-lymphocyte axis. Each family of cells has distinctive alloantigens, and the two families share alloantigens of only one known system, H-2. This implies that either immunoglobulin-producing cells are not derived from thymic lymphocytes, or if they are, the program responsible for the transition must include extensive revision of cell surface structure

Lyb-2 system of mouse B cells. Evidence for a role in the generation of antibody-forming cells

The Lyb-2 cell-surface alloantigens of the mouse are selectively and perhaps exclusively expressed in the B lymphocyte lineage, but not on antibody- forming cells. Thus if the Lyb-2 molecule is concerned in specific B cell function, it must participate in the generative phase of the antibody response. Accordingly, monoclonal Lyb-2 antibody was found to depress the plaque- forming cell (PFC) response to sheep erythrocytes in 5-d Mishell-Dutton assays when added within the first 3 d of culture, but not later. The rate of PFC generation was not affected, signifying an absolute reduction in the number of PFC generated. Because reduction of PFC counts by Lyb-2 antibody was not affected by exclusion of Lyt-2(+) T cells, it is unlikely that the reduction depends on augmented suppression by T cells. Augmented B cell- mediated suppression is also unlikely, because the PFC response of serial combinations of congenic Lyb-2.1 and Lyb-2.2 cells, in the presence of monoclonal Lyb-2.1 antibody, was reduced only in direct proportion to the number of Lyb-2.1 cells present. The PFC response of Lyb-2.1/Lyb-2.2 heterozygous cells was not reduced by Lyb-2.1 antibody, presumably because generation of PFC is impeded only if most Lyb-2 sites are blocked. Further evidence that the molecule identified by Lyb-2 plays a critical role in the generation of antibody-forming cells (AFC) in response to T-dependent antigen comes from the finding that Lyb-2 antibody does not reduce the PFC response to the T-independent antigens trinitrophenylated (TNP) Brucella abortus and TNP-FicolI, although elimination of Lyb-2(+) cells from the starting population by Lyb-2 antibody and complement reduces the PFC response to T- dependent and T-independent antigens alike