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A W Harris

Publications and source records attributed to A W Harris.

At least 109 records · Page 6Linked to original sources
PubMed1984

Differential expression of the invariant chain in mouse tumor cells: relationship to B lymphoid development.

We have examined 25 cultured lines of mouse tumor cells for synthesis of the Ii, an Ia-associated polypeptide, by using an anti-Ii monoclonal antibody. Six of the T lymphomas tested did not produce detectable levels of Ii or of surface Ia antigens. Three B lymphomas and two plasmacytomas that express surface Ia antigens were found to synthesize the Ii. In addition, Ii was immunoprecipitated from two of five Ia- pre-B lymphomas, two of four Ia- plasmacytomas, two Ia- myeloid tumors, and two fibroblast cell lines including LM(TK-). Because Ia antigens have so far been found only on cells that also synthesize Ii, we suggest that the Ii is a marker of those cells that in certain states of development or activation express Ia antigens.

Animals↗
PubMed1983

Production of RNA for secreted immunoglobulin mu chains does not require transcriptional termination 5' to the microM exons.

The mouse immunoglobulin mu gene encodes both secreted and surface-bound mu heavy chains produced by cells of the B lymphoid series. Transcripts of the mu gene are processed into mu mRNA species which differ at their 3' termini, bearing either 'microsecond' or 'microM' segments, distinguishing secreted and cell-membrane-bound mu polypeptides. During maturation of surface IgM-bearing B cells to IgM-secreting plasma cells, both the total amount of mu mRNA and the ratio of microsecond- to microM-terminated mRNA increase greatly. Two possible mechanisms for the developmental regulation of 3' RNA processing cannot yet be distinguished. One mechanism would yield the microsecond terminus by specific cleavage of a common presursor transcript encompassing both microsecond and the microM exons (Fig. 1), the other by regulated termination of transcription upstream from the microM exons. While the first mechanism would produce, as a by-product, RNA fragments containing microM exons, the second would not. We report here the detection of such microM fragments in cells producing predominantly microsecond-terminated RNA species.

Base Sequence↗
PubMed1983

Interferon-gamma induces enhanced expression of Ia and H-2 antigens on B lymphoid, macrophage, and myeloid cell lines.

The levels of class II major histocompatibility complex (MHC) antigens (la antigens) on cells of a cultured B lymphoma line (WEHI-279) were significantly increased after 24 hr incubation with medium conditioned by concanavalin A-stimulated mouse or rat spleen cells, or by an azobenzenearsonate- (ABA) specific T cell clone that had been stimulated with ABA-coupled spleen cells or concanavalin A. The levels and properties of the la-inducing activity correlated with those of interferon-gamma (IFN-gamma) measured by inhibition of virus plaque formation. Both the la-inducing activity and the IFN-gamma from the T cell clone had an apparent m.w. of 40,000 determined by gel filtration, were sensitive to treatment with trypsin or exposure to pH 2, but were stable to heat (56 degrees C, 1 hr). The induction of la antigens on WEHI-279 cells was dose-dependent, and the maximum response occurred at a concentration corresponding to 1 to 2 U/ml of antiviral activity. This T cell-derived IFN-gamma-like molecule also increased the expression of cell surface la antigens on another B cell line (WEHI-231), and cell lines of macrophage (J774) and myeloid (WEHI-3B and WEHI-265) origin. Furthermore, in all cases the levels of class I MHC (H-2K or H-2D) antigens were also increased. Similar patterns of induction of Ia and H-2 antigens were obtained with supernatants containing IFN-gamma produced by a monkey cell line (COS) that had been transfected with a plasmid bearing the cloned murine IFN-gamma gene. This activity was sensitive to pH 2 and was not present in the supernatant from COS cells that were not transfected with the murine IFN-gamma gene. These results established that IFN-gamma is the T cell-derived molecule that induces the enhanced expression of Ia and H-2 antigens on B cells and macrophages. A major physiologic role of IFN-gamma may be to regulate immune function through the enhanced expression of MHC antigens.

Animals↗
PubMed1981

Subunit structure of cell surface proteins: disulfide bonding in antigen receptors, Ly-2/3 antigens, and transferrin receptors of murine T and B lymphocytes.

The surface proteins of lymphocytes from spleen and thymus and several cultured lymphoid tumor lines were radioiodinated in situ, solubilized with Triton X-100, and examined for the presence of disulfide-bonded subunits by two-dimensional (intact, reduced) NaDodSO4/polyacrylamide gel electrophoresis. [Hynes, R. O. & Destree, A. (1977) Proc. Natl. Acad. Sci. USA 74, 2855-2859]. Few lymphocyte surface proteins were found to consist of disulfide-bonded subunits, and the most prominent of these could be identified. In normal B lymphocytes and B-lymphoma cells, IgD or IgM (or both) were the major disulfide-bonded proteins, and these were easily detectable, even without immunoprecipitation. In contrast, analysis of thymocytes and T-lymphoma cells did not reveal any protein resembling immunoglobulin in its chain structure. The major labeled thymocyte membrane protein consisting of disulfide-bonded subunits was identified as the Ly-2/3 antigen. It appeared to contain disulfide-bonded homodimers of Mr 35,000 (alpha 2) noncovalently associated with a second pair of homodimers of Mr 30,000 (beta 2). Peptide mapping showed these polypeptides to be homologous. A third disulfide-bonded homodimer, which was heterogeneous in apparent Mr, appeared to be part of the Ly-2/3 complex. All cultured T- and B-lymphoma lines examined were found to possess a major surface protein that appeared to be a disulfide-bonded homodimer of a polypeptide of Mr 95,000. This protein was identified as the receptor for transferrin. It is suggested that the presence of two or more subunits in cell surface receptors renders their ligand functionally bivalent, making ligand-induced receptor aggregation possible.

Animals↗
PubMed1981

Alkaline phosphatase in hematopoietic tumor cell lines of the mouse: high activity in cells of the B lymphoid lineage.

Alkaline phosphatase (EC 3.1.3.1) was assayed in a large number of cultured mouse tumor cell line using p-nitrophenylphosphate as the substrate. Of 19 lines of the B lymphoid lineage, including Abelson pre-B, B lymphoma, and plasma cell tumor lines, all but 1 had substantial activity averaging 407 nmol/min/mg protein (with a range from 5 to 900). Nine T lymphoid and 9 nonlymphoid hematopoietic lines examined had low activity of 0.7 to 4.2 nmol/min/mg protein. The enzyme was markedly enriched in plasma membrane preparations from the B lymphoid cells, but not in those from most T lymphoma cells. The activity of another plasma-membrane-bound enzyme, gamma-glutamyl transferase, did not vary systematically with the type of cell line but was exceptionally high in 1 T lymphoma line. Investigation of pH dependence and susceptibility to inhibition by L-phenylalanine and L-homoarginine indicated similarity of the alkaline phosphatase from B cell lines to the enzyme recoverable from normal mouse kidney, placenta, bone marrow, and lymphoid organs. The enzyme seems to provide a useful marker for tumor lines of the B lymphoid lineage and for their plasma membranes.

Alkaline Phosphatase↗
PubMed1981

mu Polypeptide phenotypes of lymphoid and myeloid cell lines containing RNA transcripts of the immunoglobulin C mu gene.

We have examined whether polyadenylated-RNA molecules encoded by the immunoglobulin C mu gene (C mu RNA) that have recently been shown to occur in mouse T lymphoma, myeloid tumor, and pre-B lymphoma cell lines are translated. Three T lymphoma, 1 myeloid tumor, and 4 Abelson lymphoma cell lines, all known to contain C mu RNA molecules, were examined for evidence of synthesis of mu polypeptides using radioimmunoassay, biosynthetic peptides were detected in any of the T lymphoma or myeloid cell lines, although mu-chains were readily detected in 2 B lymphoma lines known to contain small amounts of mu mRNA. Detection failure is unlikely to be the result of assay insensitivity and was not caused by lack of an appropriate antiserum reactivity or a requirement for mu-chains to be associated with immunoglobulin light chains. Proteolytic activity that might degrade newly synthesized mu polypeptides in these cell lines was not detected. In 3 of the 4 Abelson lymphoma cell lines containing C mu RNA, corresponding polypeptides were absent, although in the 4th they were easily detected, indicating that transcription of the C mu gene is not the sole determinant of whether a cell will synthesize the corresponding polypeptide. The parallel between the C mu RNA+, mu polypeptide- Abelson cell lines and the T lymphoma cell lines suggests that the onset of C mu transcription may occur in both T and B lineages before translational competence is acquired.

Animals↗
PubMed1980

Immunoglobulin Cmu RNA in T lymphoma cells is not translated.

It is widely believed that immunoglobulin genes might encode at least part of the receptor for antigen on the T lymphocyte. Evidence supporting this comes from the effects of anti-immunoglobulin idiotype antibodies on cellular immune networks and from the presence of idiotypes on immunologically active factors from T cells. Detailed molecular characterization of the receptors, however, has been seriously hampered by the lack of a suitable cellular source from which it might be isolated. The recent demonstration of Kemp et al. that thymocytes and certain cultured lines of mouse T lymphoma cells contain polyadenylated RNA molecules encoded by the immunoglobulin Cmu gene (Cmu RNA) prompted us to identify the corresponding protein molecules in those cells. As the haploid mouse genome contains a single Cmu gene, any polypeptide encoded by this gene should react with at least some of the antibodies present in rabbit anti-mouse IgM antiserum. In this letter we report that a number of T lymphoma lines, regardless of whether they contain Cmu RNA, synthesize no detectable mu polypeptides.

Animals↗
PubMed1980

The immunoglobulin mu constant region gene is expressed in mouse thymocytes.

It has been a matter of controversy whether the functional capacity of T cells to discriminate between antigens is mediated via immunoglobulin, an immunoglobulin-like molecule, or by the product(s) of unrelated genes. The progenitors of immunoglobulin-secreting cells, B cells, express membrane-bound immunoglobulin as the antigen-specific receptor on their surface. For T cells, although products of immunoglobulin heavy chain variable region genes are implicated as receptor components, there has been no compelling immunochemical evidence for participation of either immunoglobulin light chains or heavy chain constant regions (see refs 2-6 for the disparate views). Recently, using cloned immunoglobulin DNA sequences as hybridization probes, we have demonstrated that the immunoglobulin Cmu gene, but not the Cmu gene, is expressed as polyadenylated RNA in some T cell tumour (T lymphoma) cell lines. Individual T lymphoma lines yielded up to three discrete mu RNA species of different sizes (1.9, 2.2 and 3.0 kilobases), each species being different in size from the major mu RNA species present in B lymphoma cells (2.4 and 2.7 kilobases). We show here that cells from the normal mouse thymus contain mu RNA species, indistinguishable in size from those in T lymphoma cells, but contain little if any kappa RNA.

Animals↗
PubMed1980

Transcripts of the immunoglobulin C mu gene vary in structure and splicing during lymphoid development.

Two classes of transcripts of the mouse immunoglobulin mu constant region (C mu) gene can be distinguished. Whereas B cells contain mu mRNAs of 2.4 and 2.7 kilobases (kb), many T lymphoma, Abelson pre-B lymphoma, and myeloid tumor cell lines contain polyadenylylated RNA species bearing C mu sequences (C mu RNAs) of 1.9, 2.1, 2.3, and 3.0 kb. Production of C mu RNAs, unlike mu mRNAs, does not require recombination with the joining region (JH) locus. To define the structure of C mu RNAs, RNA from representative cell lines was fractionated by gel electrophoresis, transferred to diazobenzyloxymethyl paper, and tested for hybridization with 23 DNA fragments that collectively span a chromosomal mu gene, cloned from a plasmacytoma. All the C mu RNAs bear sequences derived from each of the four C mu domains, but none contain the intervening sequences separating domains; thus each represents a spliced RNA species. The 1.9-kb C mu RNA contains the 3' sequence characteristic of secreted mu chain, whereas the longer species bear that of membrane-bound mu chin. Hence C mu RNAs and mu mRNAs are equivalent throughout the C mu and 3' terminal regions. They differ markedly, however, in their 5' regions, because the 3.0-and 2.3-kb C mu RNAs bear sequences from within the conventional JH-C mu intervening sequence. Because these sequences are several kb from C mu, this region must contain at least one hitherto unsuspected splice site. C mu RNAs may not express immunological diversity, because no evidence was found that they bear variable regions. T and pre-B lymphoid and myeloid cells contain equivalent C mu RNA species, which coexist with mu mRNAs in some pre-B and B lymphoid lines. C mu RNA expression appears to reflect an activated state of the C mu gene common to cells at early stages of T, B, and myeloid development.

Animals↗
PubMed1980

Expression of the immunoglobulin C mu gene in mouse T and B lymphoid and myeloid cell lines.

We have used cloned nucleotide sequences as probes to search for immunoglobulin RNA in T and B lymphoid and nonlymphoid tumor cell lines. Polyadenylated RNA from the cells was fractionated electrophoretically by size and fixed to diazobenzyloxymethyl-paper, and then immunoglobulin RNA species were revealed by hybridization with 32P-labeled cloned sequences for mu heavy chain and kappa light chain. kappa mRNA was detected only in B lymphoid lines known to synthesize kappa chains. No mu RNA was detectable in erythroleukemia, mastocytoma, or sarcoma cells. RNA bearing mu constant region (C mu) sequences, however, was detected in four of nine T lymphoma and four of five myeloid tumor cell lines tested, as well as in B lymphoma and "pre-B" Abelson lymphoma cells. In contrast to the single mu RNA species of 2.6 kilobases (kb) in a plasmacytoma, individual T lymphoma and myeloid lines yielded up to three discrete mu RNA species of apparent size 1.9, 2.2, and 3.0 kb, each different from the two mu RNA species in a B lymphoma line (2.4 and 2.7 kb) or the single species in "pre-B" lymphoma cells (2.9 kb). Both chromosomal complements of the C mu gene in STRij-4 T cells were found to be rearranged from their embryonic (germ line) context. The results suggest that the C mu gene functions not only in T cells, wherein the ability to recognize antigen is long established, but also in myeloid cells.

Animals↗
PubMed1979

Deoxyribonucleoside triphosphate pools and differential thymidine sensitivities of cultured mouse lymphoma and myeloma cells.

The effects of various concentrations of thymidine on DNA synthesis and deoxyribonucleoside triphosphate contents of a highly thymidine-sensitive cultured mouse lymphoma cell line (WEHI-7) and a relatively resistant mouse myeloma cell line (HPC-108) have been studied by 32P-labelling techniques. DNA synthesis in the myeloma cells was inhibited by thymidine at concentrations of 10(-3) M or greater, while DNA synthesis in the lymphoma cells was inhibited by concentrations 30-fold lower, consistent with the 25-fold difference between the two cell lines in sensitivity to growth inhibition by thymidine. Thymidine caused marked elevation of the dTTP and dGTP pools, slight elevation or no change in the dATP pool and a marked decrease in the dCTP pool in cells of both lines. The greater resistance of HPC-108 cells to thymidine inhibition was related to the finding that they normally contained a much higher concentration of dCTP than did the WEHI-7 cells. Pool size measurements on thymidine-treated (10(-4) M) cells of an additional seven sensitive lymphoma and six relatively resistant myeloma cell lines indicated that in all 15 lines studied, with one exception, a critical concentration of dCTP of about 32 nmol per ml of cell volume was required for the maintenance of normal rates of DNA synthesis. The dCTP content found normally in the lymphoma cells was only a little above this concentration. Amongst the myeloma lines, three contained similarly low levels of dCTP, but were more resistant to thymidine inhibition probably because of their inefficient production of dTTP from thymidine. Cells of the other four myeloma lines (including HPC-108) normally contained much higher dCTP concentrations. The mechanism of thymidine action was explained by reference to the known allosteric properties of ribonucleotide reductase.

Animals↗
PubMed1979

Variations in cellular sensitivity to glucocorticoids: observations and mechanisms.

The spectrum of physiological, pathological, and genetic variations in sensitivity to glucocorticoids is reviewed. The receptor for these hormones is common to most mammalian tissues, and yet the responses are widely divergent. Although there may be differences in the receptors to account for some of this diversity, it is likely that it is largely due to cellular programming not involving the receptors. In addition to the intertissue differences in sensitivity, it is also clear that intra-tissue differences occur. The greatest amount of information has been accumulated with lymphoid cell systems and there are sensitivity differences to specific responses such as cell killing or effects on immunological functions. In these systems, there can be major variations in either the extent of the response (e. g., from mild growth inhibition to cellular killing) or whether any effect is observed. Further, dose requirements for certain responses can vary by several orders of magnitude. Within a given tissue there may be developmental changes in sensitivity that are not due to obvious changes in the receptor, and decreased sensitivity with aging that in some cases has been associated with changes in receptor binding activity. Finally, the cellular sensitivity can either be influenced by hormones and other factors that affect the ability of the glucocorticoid to elicit a particular response (in a synergistic or antagonistic manner), or the same function regulated by the glucocorticoid can be inducible by the steroid, appearing some time after administration of the steroid and disappearing after steroid removal. Genetic variations in sensitivity to glucocorticoids also occur. In humans these may be generalized, affecting glucocorticoid action in all responsive tissues, and could be important in the pathogenesis of certain diseases. Perhaps the most striking genetic alterations, however, are observed in cultured lymphoid and fibroblastic cells and in acute lymphoblastic leukaemia cells ordinarily growth inhibited or killed by the glucocorticoid. Mutant cell lines arise that are highly resistant and most of these have abnormalities in the glucocorticoid receptor. In some cases binding activity is totally lost, easily expalining the resistance. In other cases, there is a more modest reduction in binding or a change in receptor properties that give it increased or decreased nuclear and DNA binding activity. An analysis of these cell lines suggests that many of the defects are in some receptor property presently not understood that makes the receptor ineffective rather than the defect being due to the quantitative changes in receptor levels detected. The frequency of emergence of steroid-resistant cells can vary widely from about 10(-5) in S49 cells to less than 10(-8) in certain thymic cell lines...

Animals↗