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M J Soloski

Publications and source records attributed to M J Soloski.

At least 37 records · Page 2Linked to original sources
PubMed1993

HIV-1 envelope protein is expressed on the surface of infected cells before its processing and presentation to class II-restricted T lymphocytes.

T lymphocytes are activated upon binding of their Ag receptors to a complex of Ag-derived peptides and MHC class I or class II molecules expressed on the surface of APC. It is now well established that APC degrade exogenous Ag in acidic endosomal compartments, and that Ag fragments bind to class II molecules moving through these compartments on their way to the surface of the APC. Although peptides derived from some endogenous Ag can also bind to class II molecules and subsequently be recognized by class II-restricted T cells, the intracellular trafficking pathways that enable endogenous proteins to be processed for association with class II molecules remain controversial. We have analyzed the mechanism by which the envelope (env) protein of the HIV-1 is processed in infected cells for recognition by class II-restricted T cells. A large number of env-specific class II-restricted human CTL clones were shown to lyse B-lymphoblastoid cell lines expressing the env. A novel dilutional assay proved that A novel dilutional assay proved that recognition of endogenous env protein was not a consequence of release and re-uptake of the env protein and subsequent processing by the standard class II-restricted pathway. Processing of endogenous env protein required that the protein be co-translationally translocated into the endoplasmic reticulum (ER) and then exit the ER, since the class II-restricted CTL did not recognize env protein localized to the cytosol or retained in the ER of target cells. Under these conditions, however, class I-restricted recognition was readily demonstrated. Finally, class II-restricted recognition was strikingly dependent upon the steady state level of surface env protein, since extracellular reagents that removed intact env protein from the surface of target cells inhibited recognition. This inhibition operated at the Ag-processing level rather than at the level of subsequent Ag recognition. These results provide the first direct evidence that endogenously synthesized membrane proteins enter the class II-restricted Ag-processing pathway after expression on the cell surface in an intact form.

Antigen-Presenting Cells↗
PubMed1993

Qa-2 molecules are peptide receptors of higher stringency than ordinary class I molecules.

Class I molecules of the major histocompatibility complex (MHC) transport peptides to the cell surface for surveillance by T cells. Ligand specificity is stringent and differs from allele to allele. Here we report analysis of natural ligands of 'unconventional' glycophosphatidyl-anchored mouse class I molecules, Qa-2. The function of these molecules is unclear; they can serve as recognition structures for 'unrestricted' cytotoxic T cells but have not been found to present peptides to T cells, although the DNA sequence suggests a similar peptide binding groove to that of 'conventional' class I molecules, and other unconventional class I molecules can present antigens in a few cases. Pool sequencing of natural Qa-2 ligands shows that Qa-2 molecules are indeed peptide receptors, having ligand specificity similar to that of conventional class I molecules, that is, a predominant length of nine amino acids, anchor positions, and hydrophobic termination of peptides. But ligand specificity is much more stringent than with other class I molecules: of the nine positions, two are anchors and four have rather limited occupancy.

Amino Acid Sequence↗
PubMed1992

Signals delivered via the Qa-2 molecule can synergize with limiting anti-CD3-induced signals to cause T lymphocyte activation.

Qa-2 is a glycolipid anchored, MHC encoded class I molecule expressed at high levels on all murine peripheral T lymphocytes. Anti-Qa-2 antibodies have previously been found to stimulate T cells to proliferate in the presence of crosslinking antibody and PMA. We have examined the effect of anti-Qa-2 antibodies on T cells stimulated with a suboptimal concentration of immobilized anti-CD3. When anti-Qa-2 antibodies were co-immobilized with limiting anti-CD3, in the absence of PMA, a clear augmentation of T cell proliferation was seen. Interestingly, the co-stimulatory anti-Qa-2 antibodies could be directed against epitopes mapped to either the alpha 3 or the alpha 1/alpha 2 Qa-2 domains. As was the case with activation induced by soluble/crosslinked anti-Qa-2 antibodies plus PMA, CD8+ T cells were less able to be costimulated with anti-Qa-2 antibodies than CD4+ cells. Surprisingly, Ca2+ mobilization was only seen when two anti-Qa-2 antibodies reactive to separate structural domains were co-crosslinked on the surface of Indo-1 loaded T cells with a suboptimal concentration of anti-CD3. Collectively these results raise questions regarding the mechanism of Qa-2 mediated signaling and its potential role in T cell activation.

Animals↗
PubMed1992

Fetal ascitic fluid: a new source of lymphocytes for rapid chromosomal analysis.

BACKGROUND: The cells found in ascites can be processed like amniotic fluid for fetal karyotyping. We have characterized these cells and used them for a rapid cytogenetic result. CASES: Three patients presented with massive fetal ascites detected by sonography. Samples of ascitic fluid were obtained at fetal paracentesis. Cells from the fluid were cultured using standard methods for fetal blood, and were compared with fetal blood lymphocytes and amniocytes. The length of time in culture, chromosome morphology, and mitotic index of ascitic fluid cells were equivalent to those of fetal blood. In the third case, we performed immunophenotyping on the ascitic fluid cells. CONCLUSION: Ascitic fluid contains lymphocytes that permit rapid chromosomal analysis within 96 hours.

Adult↗
PubMed1991

Heat shock proteins can regulate expression of the Tla region-encoded class Ib molecule Qa-1.

T cells recognize foreign antigens in association with the highly polymorphic class I and class II molecules encoded in the major histocompatibility complex (MHC). In addition to these highly polymorphic molecules, the murine MHC also encodes, in the Qa/Tla region, several less polymorphic structures referred to as class I-like or class Ib molecules. Although no specific function has been assigned to these molecules, their overall structural similarities to the classical class I molecules and their association with beta 2-microglobulin suggest a role in antigen recognition. Recent data have suggested that the class Ib molecule Qa-1 may be involved in antigen presentation to T cells expressing gamma delta receptors. In addition, several reports have demonstrated that gamma delta T cells can respond to mycobacterial heat shock proteins. We report that transfection of a mouse fibroblast line with gene T23b leads to the surface expression of a molecule that is structurally identical to lymphocyte Qa-1b. In the transfected cells the predominant Qa-1 species was present in an immature intracellular form. The expression of mature cell surface Qa-1 was dramatically and selectively increased following heat shock. Furthermore, the addition of a tryptic digest of Mycobacterium bovis 65-kDa heat shock protein stabilized the surface expression of Qa-1b. These observations suggest that the Qa-1 molecule may be involved in the presentation of heat shock protein-derived peptides to the immune system.

Animals↗
PubMed1991

Biosynthesis of glycophospholipid bound and secreted murine class I Qa-2 polypeptides.

Murine T cells synthesize and express a cell-surface glycophospholipid anchored 40 kDa and a secreted water-soluble 39 kDa Qa-2 polypeptide. We have examined the biosynthetic pathways which lead to the production of the membrane-bound and water-soluble isoforms of the Qa-2 molecule. Using the detergent TX-114, both detergent (membrane)-bound and soluble Qa-2 polypeptides can be identified in cell lysates and can be distinguished by charge and molecular weight. Two membrane-bound forms, a 40-kDa Endo H resistant cell-surface form and a 38 kDa-Endo H sensitive form can be identified, both of which can be biosynthetically labeled with 3H-ethanolamine and can be converted to water soluble forms by digestion with a phosphatidylinositol specific phospholipase C. In addition, several water soluble polypeptides at 39, 37, 35 kDa, and a minor species at 33 kDa were identified, none of which radiolabel with 3H-ethanolamine. While the 39-kDa polypeptide was Endo H resistant, the other isoforms were sensitive to Endo H digestion. Pulse chase experiments and molecular weights of the deglycosylated core polypeptides suggest a precursor to product relationship between the intracellular water-soluble species and the mature 39-kDa secreted Qa-2 molecule. This relationship is supported by the observation that murine L cells transfected with the Qa-2 encoding class I gene Q7 fail to express membrane-bound Qa-2 molecules yet synthesize both intracellular water-soluble and secreted Qa-2 molecules. These findings argue for a pathway in which secreted soluble Qa-2 molecules are derived from intracellular precursors.

Animals↗
PubMed1991

Physical and molecular genetic analysis of Qa-2 antigen expression: multiple factors controlling cell surface levels.

We have examined the surface expression of Qa-2 on lymphocyte subpopulations and on splenocytes from inbred mouse strains by a radioactive binding assay using purified anti-Qa-2 antibodies and antibody fragments (Fab). Quantitative measurements by Scatchard analysis revealed that spleen cells from Qa-2high mice express (4-5) x 10(4) Qa-2 molecules/cell, whereas T lymphocytes have as high as (7-8) x 10(4) molecules/cell. In addition, it was determined that B lymphocytes express (5-6) x 10(3) molecules on their cell surface. The Qa-2 levels on anti-CD3-activated T cells is 1.0 x 10(5) molecules/cell. Previous experiments have shown that the quantity of Qa-2 varies in a strain-specific fashion and may be classified into three groups: Qa-2high, Qa-2medium and Qa-2low. Our results indicated that Qa-2high strains express (4-5) x 10(4) Qa-2 molecules/cell, Qa-2medium strains (B6-K2, B10.A, A/J, BALB/c and DBA/2) express (1-1.7) x 10(4) molecules/cell, and Qa-2low strains (SWR/J and DBA/1) express no more than 6 x 10(3) molecules/cell. Detection of Q7 or Q9 mRNA by the polymerase chain reaction revealed that Qa-2high strains express two functional Qa-2 enconding genes, Q7 and Q9, whereas Qa-2medium and Qa-2low strains express either Q7 or Q9. These results strongly suggest that Qa-2 gene dosage contributes in part to the variation of Qa-2 levels on the cell surface.

Animals↗
PubMed1989

Molecular basis of Qa-11 antigen and paradoxical Qa-gene expression in an H-2 recombinant.

The Qa-11 Ag expressed in certain strains with the B2-microglobulin-b allele, apparently maps into the Tla region as well as into the Qa-2 region. Moreover Qa-11 has been shown to be biochemically indistinguishable from Qa-2. Genetic complementation studies combining the right Qa and Tla regions failed to lead to Qa-11 expression. To elucidate the molecular basis of this apparent paradox, we examined the expression of Qa-11 on products of transfected Q-region class I genes. Immunochemical analysis has shown that the Qa-11 Ag is expressed on class I molecules encoded by the Q7 gene from both C57BL/10 (Q7b) and BALB/c (Q7d), but not on the protein product of the Q9 gene isolated from the C57BL/10 strain (Q9b). Inasmuch as the predicted protein products of the Q7b and Q9b genes would differ at a single amino acid, a residue critical for Qa-11 expression has been identified. Based on these results it is proposed that among the beta-2-mb strains, the Qa-11+/Qa-2+ mice are likely to express at least the Q7 gene, whereas Qa-11-/Qa-2+ mice express only Q9. In support of this model, the Qa-2+/Q-11- recombinant B6.K2, essential for the apparent mapping of Qa-11 into the Tla region, expresses only Q9 but not Q7 encoded molecules on the cell surface, and only Q9 and no processed Q7 mRNA is detected in the cytoplasm. This expression pattern in B6.K2 cannot be explained on the basis of a single crossing-over event.

Animals↗
PubMed1989

Anti-Qa-2-induced T cell activation. The parameters of activation, the definition of mitogenic and nonmitogenic antibodies, and the differential effects on CD4+ vs CD8+ T cells.

The MHC Ag Qa-2 is a glycolipid anchored class I molecule expressed at high levels on all peripheral T lymphocytes. In this study we found that anti-Qa-2 antibodies could stimulate the proliferation of murine T cells in vitro. Anti-Qa-2-induced proliferation required secondary cross-linking with anti-Ig antibody and the presence of PMA. Only Qa-2+ strains could be induced to proliferate by anti-Qa-2 antibody, but under the conditions employed, anti-CD3 could induce proliferation in Qa-2+ and Qa-2-strains. Interestingly, only anti-Qa-2 reagents directed against the alpha 3 domain of the Qa-2 class I molecule were effective in inducing proliferation. Furthermore, unlike purified CD4+ cells, purified CD8+ cells were unable to be stimulated by the anti-Qa-2 antibodies. These results lead to the inclusion of Qa-2 in a group of physiologically relevant, glycolipid-anchored, cell-surface molecules, mobilization of which can generate signals that initiate the proliferation of T cells. Such molecules may play a secondary role in cellular activation after the primary engagement of the TCR.

Animals↗
PubMed1989

Qa-2 expression in the adult murine thymus. A unique marker for a mature thymic subset.

The MHC Ag, Qa-2, is expressed on all peripheral T cells, a subset of bone marrow cells, and to a lesser extent on B cells. The Qa-2 Ag is also expressed on 5 to 6% of normal adult murine thymocytes. Through the use of flow cytometry, counterflow centrifugal elutriation and acridine orange staining, we have analyzed the cell surface phenotype, cell size, and cell cycle status of this thymic population. Our studies indicate that Qa-2+ thymocytes are large, non-mitotic, G1 cells which have the cell surface phenotype of CD5+, CD3+, J11dLO and lack receptors for peanut agglutinin. This population can be further subdivided into three categories; CD4+/CD8-, CD4-/CD8+, and CD4-/CD8-. These data indicate that Qa-2 surface expression can only be detected on thymocytes in the final stages of differentiation. The Qa-2 Ag can be used as a cell surface marker to identify a unique subset of mature thymocytes.

Animals↗
PubMed1988

Further characterization of the membrane anchor found on the tissue-specific class I molecule Qa2.

Previous studies have determined that various Qa2 serologic determinants can be removed from the surface of spleen cells by treatment with a phospholipase C. Our studies have determined that the class I molecule Qa2, expressed on the surface of spleen cells and activated T cells, behaves as an integral membrane protein based on its ability to associate with detergent micelles. Studies utilizing two purified phospholipase C have revealed that although most (90 to 95%) of the Qa2 molecules expressed on the surface of resting spleen cells are released as intact 40-kDa polypeptides associated with beta 2-microglobulin, activated T cells contain a major cell subpopulation expressing lipase-resistant Qa2 molecules. Flow cytometric analysis revealed that L3T4+-activated T cells expressed lipase-sensitive Qa2 molecules, whereas Lyt-2+ cells express lipase-resistant forms of the Qa2 molecule. The relationship between the secreted form of the Qa2 molecule and the lipase-generated soluble Qa2 molecule was investigated. Based on SDS-PAGE analysis, the secreted Qa2 molecules has a Mr of 39 kDa whereas the cell surface form released from either resting spleen or activated T cells by phosphatidylinositol-specific phospholipase C has a Mr of approximately equal to 40 kDa. Furthermore, the secreted Qa2 molecule lacks an epitope, cross-reacting determinant, often present on lipase-solubilized cell surface molecules. Thus, based on serologic and biochemical criteria, the soluble Qa2 molecules generated by an exogenous phospholipase C and the secreted Qa2 molecule are structurally distinct.

Animals↗
PubMed1988

Qa-region class I gene expression: identification of a second class I gene, Q9, encoding a Qa-2 polypeptide.

A feature of the expression of the tissue-specific class I antigen Qa-2 is the quantitative variation among mouse strains. Recently, the class I gene Q7 has been shown to encode a protein product that is biochemically indistinguishable from the lymphocyte-bound Qa-2 molecule. Utilizing gene transfection, we have identified a second Qa-2 subregion class I gene (Q9), in H-2b mice, which encodes a polypeptide biochemically similar to the Q7 and the Qa-2 polypeptides. Furthermore, we have observed that cell lines transfected with the allelic forms of the Q7 gene from C57BL/10 (Qa-2hi) or BALB/C (Qa-2low) display quantitative differences in cell-surface expression. Based on these studies, we suggest that gene dosage and allele-specific variation in cell-surface expression contribute to the strain-specific variation in the levels of Qa-2 antigen expression.

Alleles↗
PubMed1986

Qa gene expression: biosynthesis and secretion of Qa-2 molecules in activated T cells.

The biosynthesis and expression of the tissue-specific class I molecule Qa-2 have been studied in resting and activated T-cell populations. Polyclonal activation of T lymphocytes induces a 3- to 4-fold increase in the biosynthesis of Qa-2 molecules but no increase in cell-surface levels. Analysis of the biosynthetic pathway of the Qa-2 molecule in activated lymphocytes reveals that approximately equal to 70% of the newly synthesized Qa-2 molecules are secreted as soluble molecules. In resting-cell populations, Qa-2 remains entirely cell-associated. This process is unique to the Qa-2 molecule, since other class I molecules (e.g., H-2Kb and H-2Db) synthesized by activated cells remain cell-associated. The possibility that the secreted Qa-2 molecule is the product of a new Qa gene or an alternatively spliced mRNA is considered. These results indicate that the Qa-2 molecules may not just function as a cell-surface recognition structure but also may serve a role as a soluble factor synthesized by activated lymphoid cell populations.

Animals↗
PubMed1985

Molecular weight diversity among murine class I antigens: both the mature cell surface forms and the unglycosylated polypeptides vary significantly in molecular weight.

The molecular weights of the fully glycosylated cell surface form and the unglycosylated polypeptide of five murine class I antigens (H-2Kb, Db, TL, Qa-1.1, and Qa-2) were compared by SDS-PAGE. Significant molecular weight diversity was observed for both forms among these molecules. The size of the fully glycosylated forms ranged from approximately 52,000 daltons (H-2Db) to 41,000 daltons (Qa-2), whereas the unglycosylated polypeptides ranged from 43,000 daltons (H-2Kb and TL) to 33,000 daltons (Qa-2). The magnitude of the size variation observed in the unglycosylated polypeptides implies that there are differences in the gene organization, RNA processing or post-translational modifications of various class I glycoproteins.

Animals↗