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The classical minor lymphocyte stimulating (Mls) antigens, which induce a strong primary T cell response in vitro, are closely linked to endogenous copies of mouse mammary tumor viruses (MMTV). Expression of Mls genes leads to clonal deletion of T cell subsets expressing specific T cell receptor (TCR) V beta chains. We describe the isolation and characterization of a new exogenous (infectious) MMTV with biological properties similar to the Mls antigen Mls-1a. In vivo administration of either Mls-1a-expressing B cells or the infectious MMTV (SW) led to an increase of T cells expressing V beta 6 followed by their deletion. Surprisingly, different kinetics of deletion were observed with the exogenous virus depending upon the route of infection. Infection through the mucosa led to a slow deletion of V beta 6+ T cells, whereas deletion was rapid after subcutaneous infection. Sequence analysis of the open reading frames in the 3' long terminal repeat of both this exogenous MMTV (SW) and of Mtv-7 (which is closely linked to Mls-1a) revealed striking similarities, particularly in the COOH terminus, which has been implicated in TCR V beta recognition. The identification of an infectious MMTV with the properties of a strong Mls antigen provides a new, powerful tool to study immunity and tolerance in vivo.

SummarySuperantigens are defined by their ability to stimulate a large fraction of T cells via interaction with the T cell receptor (TCR) V/3 domain. Endogenous superantigens, classically termed minor lymphocyte-stimulating (Mls) antigens, were recently identified as products of open reading flames (ORF) in integrated proviral copies of mouse mammary tumor virus (MMTV). We have described an infectious MMTV homologue of the classical endogenous superantigen Mls-1 ~ (Mtv-7). The ORF molecules of both the endogenous Mtv-7 and the infectious MMTV(SW) interact with T cells expressing the TCR VB6, 7, 8.1, and 9 domains. Furthermore, the COOH termini of their ORF molecules, thought to confer TCR specificity, are very similar. Since successful transport of MMTV from the site of infection in the gut to the mammary gland depends on a functional immune system, we were interested in determining the early events after and requirements for MMTV infection. We show that MMTV(SW) infection induces a massive response of V~6 + CD4 + T cells, which interact with the viral ORE Concomitantly, we observed a B cell response and differentiation that depends on both the presence and stimulation of the superantigen-reactive T cells. Furthermore, we show that B cells are the main target of the initial MMTV infection as judged by the presence of the reverse-transcribed viral genome and ORF transcripts. Thus, we suggest that MMTV infection of B cells leads to ORF-mediated B-T cell interaction, which maintains and possibly amplifies viral infection. M inor lymphocyte-stimulating (Mls) antigens are encodedby an open reading frame (ORF) z in the 3' LTR of endogenous mouse mammary tumor virus (MMTV) (1-3). They stimulate a large proportion of T cells due to the fact that T cell reactivity to Mls antigens is determined by the usage of the V segment of the TCR B chain (4, 5). The classical (and strongest) Mls determinant, Mls-1 ', which interacts with T cells expressing VB6, 7, 8.1, and 9 elements (4-7), is encoded by the ORF of the Mtv-7 proviral locus (8). We have recently found an exogenous (infectious) MMTV, MMTV(SW), with properties of Mls-1 ' (9). The ORF molecules of Mtv-7 and MMTV(SW) display an almost identical amino acid sequence, particularly in COOH termini implied to confer TCR VB specificity. In fact, both endogenous Mtv-7 and infectious MMTV(SW) interact with the same TCR VB elements. In an MMTV-infected host, ORF-reactive T cells are deleted from both the thymic and peripheral T cell pool, although at a considerably slower rate than after expression of the endogenous Mtv-7 (Mls-l') locus. Furthermore, injection of adult mice with MMTV(SW) leads to expansion of V~6 + CD4 + T cells similar to that observed after injection of . A functional immune system seems to be a prerequisite for successful transport of MMTV to the mammary gland after neonatal infection through the gut mucosa. Thus, thymectomy, absence of T cells (in nude mice), or deletion of the ORF-reactive T cells prevent virus transmission to the next generation (lO-12).Whil...

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Previous work from this laboratory has revealed that infection of mice with Leishmania major leads to an expansion of gamma delta+ T cells in the spleen. Further examination of the gamma delta+ T cells expanding in infected mice has shown that the majority of these cells in the spleen, lymph nodes, blood and liver expressed the V delta 4 gene segment. Cell cycle analysis, using propidium iodide incorporation, demonstrated that while only 1% of alpha beta+ T cells in the spleen were in either S + G2/M phase, up to 10% of the gamma delta+ T cells were in cycling phase 8 weeks after infection. Comparison of the state of activation of the two populations in different organs after infection, confirmed that gamma delta+ T cells are actively dividing in lymph nodes, liver and blood, but not in the thymus or among intraepithelial lymphocytes. Examination of the expression of different activation markers on the surface of gamma delta+ T cells in the spleen of both normal and chronically infected BALB/c mice by FACS analysis, revealed increased expression of LFA-1, CD25, CD44, 4F2, CD28 and the heat-stable antigen, whereas Thy-1 and CD5 decreased. Collectively, these results suggest an oligoclonal expansion and activation of gamma delta+ T cells in response to L. major infection.