Quantitation of endogenous mouse mammary tumor virus superantigen expression by lymphocyte subsets

Abstract

Superantigens (SAg) encoded by endogenous mouse mammary tumor viruses (Mtv) interact with the V beta domain of the T cell receptor (TcR-V beta). Presentation of Mtv SAg can lead to stimulation and/or deletion of the reactive T cells, but little is known about the quantitative aspects of SAg presentation. Although monoclonal antibodies have been raised against Mtv SAg, they have not been useful in quantitating SAg protein, which is present in very low amounts in normal cells. Alternative attempts to quantitate Mtv SAg mRNA expression are complicated by the fact that Mtv transcription occurs from multiple loci and in different overlapping reading frames. In this report we describe a novel competitive polymerase chain reaction assay which allows the locus-specific quantitation of SAg expression at the mRNA level in lymphocyte subsets from mouse strains with multiple endogenous Mtv loci. In B cells as well as T cells (CD4+ or CD8+), Mtv-6 SAg is expressed at the highest levels, followed by Mtv-7 SAg and (to a much lesser extent) Mtv-8,9. Consistent with functional Mtv-7 SAg presentation studies, we find that Mtv-7 SAg expression is higher in B cells than in CD8+ T cells and very low in the CD4+ subset. The overall hierarchy in Mtv SAg expression (i.e. Mtv-6 > Mtv-7 > Mtv 8,9) was also observed for mRNA isolated from neonatal thymus. Furthermore, the kinetics of intrathymic deletion of the corresponding TcR-V beta domains during ontogeny correlated with the levels of Mtv SAg expression. Collectively our data suggest that T cell responses to Mtv SAg are largely controlled by SAg expression levels on presenting cells.

Negative thymocyte selection to HERV-K18 superantigens in humans

“…They are that donors differed in their capacity to undergo SAg-mediated negative selection despite the rigorous selection of thymocyte samples for consistent biologic behavior and that thymic HERV-K18 SAg expression levels are anticipated to influence the efficiency and the kinetics of deletion, analogous to MMTV SAgs. 9,10 Our data indeed lend support to this idea (Figure 1). We are in the process of dissecting the respective role of each parameter by large-scale genetic analyses of the interactions among HERV-K18, MHC class 2, and TCR polymorphisms and by repopulation assays in immunodeficient mice reconstituted with human hematopoietic precursors that can give rise to thymocytes plus APCs capable of negative selection.…”

“…6 Thymic deletion reflects the susceptibility of immature and semimature thymocytes to undergo negative selection at the corticomedullary junction and in the thymic medulla replete of professional antigen-presenting cells (APCs). 7,8 It requires that SAg expression exceed a threshold level during thymocyte development 9,10 and that SAgs be presented by the appropriate thymic cell type, which includes medullary epithelial cells and hematopoietic APCs. 11 Expression of some MMTV SAgs is restricted to the thymocyte lingeage, 12 and efficient deletion in these instance relies on paracrine transfer to major histocompatibility complex (MHC) class 2 ϩ thymic APCs.…”

Receptor revision in CD4 T cells is influenced by follicular helper T cell formation and germinal-center interactions

“…Our results showing no effect of LPS stimulation on Mtv-8 superAg RNA expression are surprising in light of evidence that LPS stimulation enhances expression of Mtv-8 envelope RNA (44). However, superAg and envelope are encoded by different portions of the Mtv-8 locus and are differentially regulated (19,45).…”

“…Vβ5 + TCRs interact with an extrathymic superantigen (superAg) encoded by mouse mammary tumor virus 8 (Mtv-8), a defective retrovirus (17,18). Mtv-8 is very poorly expressed and only weakly stimulates T cells (17)(18)(19). Most Mtv-8-reactive Vβ5 + CD4 T cells become anergic and are deleted, leading to an age-dependent decline in the CD4:CD8 T-cell ratio in Vβ5 Tg B6 mice (16,20).…”

META-Controlled env-Initiated Transcripts Encoding Superantigens of Murine Mtv29 and Mtv7 and Their Possible Role in B Cell Lymphomagenesis

“…These transcripts clearly come from Mtv7, because Mtv8, the only other MMTV in that strain, does not show any RT-PCR products in either of the two lymphomas examined in detail. In a previous study in which a quantitation was made of RsaI-digested RT-PCR products obtained from 5Ј LTR initiated (presumably vSAg) transcripts from Mtv6, Mtv7, Mtv8, and Mtv9 in normal B and T cells from BALB-D2.Mls-1a mice, such transcripts for Mtv7 were found to be highly expressed, particularly in B cells (26). These LTR-initiated transcripts were thought to encode vSAg7, which is known to be highly expressed in B cells and to cause efficient deletion of Mtv7-responsive V␤6 ϩ , V␤7 ϩ , V␤8.1 ϩ , and V␤9 ϩ T cells in I-E ϩ strains that possess Mtv7 in their genome (27)(28)(29).…”