PubMed Health⌕ Search

Biomedical subjects

S W Granger

Publications and source records attributed to S W Granger.

8 recordsLinked to original sources
PubMed2001

Constitutive expression of LIGHT on T cells leads to lymphocyte activation, inflammation, and tissue destruction.

LIGHT, a member of the TNF family of cytokines (homologous to lymphotoxin, exhibits inducible expression and competes with HSV glycoprotein D for herpesvirus entry mediator, a receptor expressed on T cells), is induced on activated T cells and mediates costimulatory and antitumor activity in vitro. Relatively little information is available on the in vivo effects of LIGHT expression, particularly within the T cell compartment. In this work, we describe transgenic mice that express human LIGHT under the control of the CD2 promoter, resulting in constitutive transgene expression in cells of the T lymphocyte lineage. LIGHT-transgenic animals exhibit abnormalities in both lymphoid tissue architecture and the distribution of lymphocyte subsets. They also show signs of inflammation that are most severe in the intestine, along with tissue destruction of the reproductive organs. These LIGHT-mediated effects were recapitulated when immune-deficient mice were reconstituted with bone marrow from LIGHT-transgenic donor mice. T cells in the LIGHT-transgenic mice have an activated phenotype and mucosal T cells exhibit enhanced Th1 cytokine activity. The results indicate that LIGHT may function as an important regulator of T cell activation, and implicate LIGHT signaling pathways in inflammation focused on mucosal tissues.

Animals↗
PubMed2001

Genomic characterization of LIGHT reveals linkage to an immune response locus on chromosome 19p13.3 and distinct isoforms generated by alternate splicing or proteolysis.

LIGHT is a member of the TNF cytokine superfamily that signals through the lymphotoxin (LT)beta receptor and the herpesvirus entry mediator. LIGHT may function as a costimulatory factor for the activation of lymphoid cells and as a deterrent to infection by herpesvirus, which may provide significant selective pressure shaping the evolution of LIGHT. Here, we define the molecular genetics of the human LIGHT locus, revealing its close linkage to the TNF superfamily members CD27 ligand and 4-1BB ligand, and the third complement protein (C3), which positions LIGHT within the MHC paralog on chromosome 19p13.3. An alternately spliced isoform of LIGHT mRNA that encodes a transmembrane-deleted form is detected in activated T cells and gives rise to a nonglycosylated protein that resides in the cytosol. Furthermore, membrane LIGHT is shed from the cell surface of human 293 T cells. These studies reveal new mechanisms involved in regulating the physical forms and cellular compartmentalization of LIGHT that may contribute to the regulation and biological function of this cytokine.

4-1BB Ligand↗
PubMed2001

The helper virus envelope glycoprotein affects the disease specificity of a recombinant murine leukemia virus carrying a v-myc oncogene.

Many retroviruses that carry oncogenes (acute transforming viruses) are generally replication-defective and therefore require co-infection with a replication competent 'helper' retrovirus for infectivity. The helper virus provides the retroviral proteins necessary for particle production and infection. These include the envelope glycoproteins that specifically bind to cell surface receptors and mediate viral adsorption and entry. Thus, a particular helper virus may influence the nature of disease induced by an oncogene-containing retrovirus due to tissue tropism of the helper. In a previous study, a replication-defective recombinant Moloney murine leukemia virus containing the v-myc oncogene was generated (M-MuLV(myc); Brightman B.K., Pattengale P.K., and Fan H., J Virol 60: 68-81, 1986). When M-MuLV(myc) was inoculated into mice using the non-pathogenic amphotropic murine leukemia virus (Am-MuLV 4070) as a helper, T- and B-lymphoblastic lymphomas resulted with the following two surface phenotypes, namely, (1) Thy 1.2+, B220- and (2) Thy 1.2-, B220+. Thy 1.2 surface antigen is characteristic of cells of the lymphoid lineage, whereas B220 surface antigen is characteristic of cells of the B-lymphoid lineage. In these experiments, to assess the influence of the helper virus on the disease specificity of M-MuLV(myc), two weakly pathogenic ecotropic helper MuLVs that interact with different cell surface receptors than Am-MuLV (Mo+PyF101 and AKV MuLV) were used to pseudotype M-MuLV(myc). In both cases, when inoculated into mice, these pseudotypes induced only T-lymphoblastic lymphoma. These results indicate that for M-MuLV(myc) the types of the tumors induced are influenced by the helper virus utilized, and they suggest that different lymphoid cells may express different levels of retroviral receptors.

Animals↗
PubMed1999

Tandemization of a subregion of the enhancer sequences from SRS 19-6 murine leukemia virus associated with T-lymphoid but not other leukemias.

Most simple retroviruses induce tumors of a single cell type when infected into susceptible hosts. The SRS 19-6 murine leukemia virus (MuLV), which originated in mainland China, induces leukemias of multiple cellular origins. Indeed, infected mice often harbor more than one tumor type. Since the enhancers of many MuLVs are major determinants of tumor specificity, we tested the role of the SRS 19-6 MuLV enhancers in its broad disease specificity. The enhancer elements of the Moloney MuLV (M-MuLV) were replaced by the 170-bp enhancers of SRS 19-6 MuLV, yielding the recombinants DeltaMo+SRS(+) and DeltaMo+SRS(-) M-MuLV. M-MuLV normally induces T-lymphoid tumors in all infected mice. Surprisingly, when neonatal mice were inoculated with DeltaMo+SRS(+) or DeltaMo+SRS(-) M-MuLV, all tumors were of T-lymphoid origin, typical of M-MuLV rather than SRS 19-6 MuLV. Thus, the SRS 19-6 MuLV enhancers did not confer the broad disease specificity of SRS 19-6 MuLV to M-MuLV. However, all tumors contained DeltaMo+SRS M-MuLV proviruses with common enhancer alterations. These alterations consisted of tandem multimerization of a subregion of the SRS 19-6 enhancers, encompassing the conserved LVb and core sites and adjacent sequences. Moreover, when tumors induced by the parental SRS 19-6 MuLV were analyzed, most of the T-lymphoid tumors had similar enhancer alterations in the same region whereas tumors of other lineages retained the parental SRS 19-6 MuLV enhancers. These results emphasize the importance of a subregion of the SRS 19-6 MuLV enhancer in induction of T-cell lymphoma. The relevant sequences were consistent with crucial sequences for T-cell lymphomagenesis identified for other MuLVs such as M-MuLV and SL3-3 MuLV. These results also suggest that other regions of the SRS 19-6 MuLV genome contribute to its broad leukemogenic spectrum.

3T3 Cells↗
PubMed1998

In vivo footprinting of the enhancer sequences in the upstream long terminal repeat of Moloney murine leukemia virus: differential binding of nuclear factors in different cell types.

The enhancer sequences in the Moloney murine leukemia virus (M-MuLV) long terminal repeat (LTR) are of considerable interest since they are crucial for virus replication and the ability of the virus to induce T lymphomas. While extensive studies have identified numerous nuclear factors that can potentially bind to M-MuLV enhancer DNA in vitro, it has not been made clear which of these factors are bound in vivo. To address this problem, we carried out in vivo footprinting of the M-MuLV enhancer in infected cells by in vivo treatment with dimethyl sulfate (DMS) followed by visualization through ligation-mediated PCR (LMPCR) and gel electrophoresis. In vivo DMS-LMPCR footprinting of the upstream LTR revealed evidence for factor binding at several previously characterized motifs. In particular, protection of guanines in the central LVb/Ets and Core sites within the 75-bp repeats was detected in infected NIH 3T3 fibroblasts, Ti-6 lymphoid cells, and thymic tumor cells. In contrast, factor binding at the NF-1 sites was found in infected fibroblasts but not in T-lymphoid cells. These results are consistent with the results of previous experiments indicating the importance of the LVb/Ets and Core sequences for many retroviruses and the biological importance especially of the NF-1 sites in fibroblasts and T-lymphoid cells. No evidence for factor binding to the glucocorticoid responsive element and LVa sites was found. Additional sites of protein binding included a region in the GC-rich sequences downstream of the 75-bp repeats (only in fibroblasts), a hypersensitive guanine on the minus strand in the LVc site (only in T-lymphoid cells), and a region upstream of the 75-bp repeats. These experiments provide concrete evidence for the differential in vivo binding of nuclear factors to the M-MuLV enhancers in different cell types.

3T3 Cells↗
PubMed1998

Tumorigenic potential of a recombinant retrovirus containing sequences from Moloney murine leukemia virus and feline leukemia virus.

A recombinant retrovirus, termed MoFe2-MuLV, was constructed in which the U3 region of T-lymphomagenic Moloney murine leukemia virus (Mo-MuLV) was replaced by that of FeLV-945, a provirus of unique long terminal repeat (LTR) structure identified only in non-T-cell, non-B-cell lymphomas of the domestic cat. The LTR of FeLV-945 is unusual in that it contains only a single copy of the transcriptional enhancer followed 25 bp downstream by a 21-bp sequence in triplicate in tandem. Infectivity of MoFe2-MuLV was demonstrated in vitro in SC-1 cells and in vivo in neonatal NIH-Swiss mice. Tumors occurred in MoFe2-MuLV-infected animals following a latency period of 4 to 10 months (average, 6 months). The results of Southern blot analysis of the T-cell receptor beta locus demonstrated that all tumors were lymphomas of T-cell origin. MoFe2-MuLV LTRs were amplified by PCR from tumor DNA and were characterized by nucleotide sequence analysis. LTRs from the tumors that occurred with relatively shorter latency predominantly retained the original MoFe2-MuLV sequence intact and unaltered. Tumors that occurred with relatively longer latency contained LTRs that also retained the 21-bp sequence triplication characteristic of the original virus but had acquired various duplications of enhancer sequences. The repeated identification of enhancer duplications in late-appearing tumors suggests that the duplication affords a selective advantage, although apparently not in the efficient induction of T-cell lymphoma. Proto-oncogenes known to be targets of insertional mutagenesis in the majority of Mo-MuLV-induced tumors or in feline non-T-cell, non-B-cell lymphomas were shown not to be rearranged in any tumor examined. Mink cell focus-inducing (MCF) proviral DNA was readily detectable in some, but not all, tumors. The presence or absence of MCF did not correlate with the kinetics of tumor induction. These studies indicate that the single-enhancer, triplication-containing FeLV LTR, typical of non-T-cell, non-B-cell lymphomas in cats, is competent in the induction of T-cell lymphoma in mice. The findings suggest that the mechanism of MoFe2-MuLV-mediated lymphomagenesis may differ from that of Mo-MuLV-mediated disease, considering the possible involvement of novel oncogenes and the variable presence of MCF recombinants.

Animals↗
PubMedDate not supplied

Purification of Moloney murine leukemia virus chromatin from infected cells by an affinity method.

Our goal was to develop a system to study proteins that associate in vivo with the Moloney murine leukemia virus (M-MuLV) enhancer elements by the isolation of intact proviral chromatin. The M-MuLV long terminal repeats (LTRs) contain tandemly repeated transcriptional enhancer sequences consisting of smaller motifs that bind cellular DNA-binding proteins implicated in transcriptional regulation. The M-MuLV enhancers are also important for disease specificity and latency of disease induction. To enrich for proviral chromatin containing M-MuLV LTR sequences, an affinity purification scheme was employed that relies on the affinity of bacterial Lac repressor protein for Lac operator (LacO) DNA sequences. An infectious M-MuLV recombinant was constructed that contains bacterial LacO sequences inserted into a nonessential region downstream from the 5' LTR of the virus (M-MuLV-LacO). Nuclei from M-MuLV-LacO-infected cells were digested with PvuII (which will liberate an LTR fragment containing LacO sequences), and digested chromatin was leached from the nuclei in hypotonic buffer. M-MuLV-LacO chromatin was then recovered by binding to an affinity matrix consisting of a beta-galactosidase-Lac repressor fusion protein anchored to acrylamide beads by an anti-beta-galactosidase monoclonal antibody [7]. Specifically bound chromatin was eluted under physiological conditions by incubation with the galactose analog isopropyl-beta-D-thiogalactopyranoside. Southern blot analysis confirmed the specific enrichment of M-MuLV proviral chromatin by this method.

Animals↗