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A Rickinson

Publications and source records attributed to A Rickinson.

31 records · Page 2Linked to original sources
PubMed1991

Induction of bcl-2 expression by Epstein-Barr virus latent membrane protein 1 protects infected B cells from programmed cell death.

Epstein-Barr virus (EBV) not only induces growth transformation in human B lymphocytes, but has more recently been shown to enhance B cell survival under suboptimal conditions where growth is inhibited; both effects are mediated through the coordinate action of eight virus-coded latent proteins. The effect upon cell survival is best recognized in EBV-positive Burkitt's lymphoma cell lines where activation of full virus latent gene expression protects the cells from programmed cell death (apoptosis). Here we show by DNA transfection into human B cells that protection from apoptosis is conferred through expression of a single EBV latent protein, the latent membrane protein LMP 1. Furthermore, we demonstrate that LMP 1 mediates this effect by up-regulating expression of the cellular oncogene bcl-2. The interplay between EBV infection and expression of this cellular oncogene has important implications for virus persistence and for the pathogenesis of virus-associated malignant disease.

Antigens, Viral↗
PubMed1990

Novel transcription from the Epstein-Barr virus terminal EcoRI fragment, DIJhet, in a nasopharyngeal carcinoma.

Transcription of Epstein-Barr virus (EBV) genes in epithelial tissue, one of the two principal cell types infected by EBV, is not well characterized. EBV transcription in a nasopharyngeal carcinoma established in nude mice, C15, has been analyzed by using strand-specific RNA probes and sequence analysis of a C15 cDNA library. In C15, two equally abundant mRNAs of 3.7 and 2.8 kilobases (kb) are encoded by the sequences that encode latent membrane protein (LMP). Hybridization with probes specific for the 3' end of the LMP mRNA to Northern (RNA) blots and sequence analysis of cDNAs representing the messages indicated that the 3.7- and 2.8-kb mRNAs are 3' coterminal. Sequence analysis of additional cDNAs revealed an mRNA that is spliced identically to the LMP mRNA but is initiated 5' to the promoter for LMP. A probe representing the sequences contained within the cDNA which are 5' to the LMP promoter identified the 3.7-kb mRNA in C15 and a low-abundance 3.7-kb mRNA in B95-8 RNA. These data indicate that transcription of the LMP-encoding sequences is complex and that LMP can be expressed from an additional RNA in both nasopharyngeal carcinoma and lymphoid cells. Hybridization with BamHI-A identified a predominant 4.8-kb mRNA and two less abundant larger-molecular-weight mRNAs transcribed in C15. These mRNAs are consistently expressed in all passages in nude mice of the C15 tumor. Hybridization with strand-specific probes and sequence analysis of three cDNAs revealed that these mRNAs are transcribed from left to right. Sequence analysis of cDNAs representing the 3' end of the mRNAs identified an open reading frame that could potentially encode a protein of 174 amino acids. In situ hybridization of a 35S-labeled RNA probe homologous to the BamHI-A cDNA to tissue sections revealed that the BamHI-A mRNA is not focally expressed and is transcribed in all cells within the C15 tumor. Linear forms of EBV DNA were not detected in any of the C15 tumors, and replicative viral antigens have not been detected. These data suggest that the C15 tumor represents a latently infected tumor and that the transcription from BamHI-A, which is expressed in all cells, is not associated with virus replication.

Amino Acid Sequence↗
PubMed1990

Epstein-Barr virus latent membrane protein (LMP1) and nuclear proteins 2 and 3C are effectors of phenotypic changes in B lymphocytes: EBNA-2 and LMP1 cooperatively induce CD23.

Latent Epstein-Barr virus (EBV) infection and growth transformation of B lymphocytes is characterized by EBV nuclear and membrane protein expression (EBV nuclear antigen [EBNA] and latent membrane protein [LMP], respectively). LMP1 is known to be an oncogene in rodent fibroblasts and to induce B-lymphocyte activation and cellular adhesion molecules in the EBV-negative Burkitt's lymphoma cell line Louckes. EBNA-2 is required for EBV-induced growth transformation; it lowers rodent fibroblast serum dependence and specifically induces the B-lymphocyte activation antigen CD23 in Louckes cells. These initial observations are now extended through an expanded study of EBNA- and LMP1-induced phenotypic effects in a different EBV-negative B-lymphoma cell line, BJAB. LMP1 effects were also evaluated in the EBV-negative B-lymphoma cell line BL41 and the EBV-positive Burkitt's lymphoma cell line, Daudi (Daudi is deleted for EBNA-2 and does not express LMP). Previously described EBNA-2- and LMP1-transfected Louckes cells were studied in parallel. EBNA-2, from EBV-1 strains but not EBV-2, induced CD23 and CD21 expression in transfected BJAB cells. In contrast, EBNA-3C induced CD21 but not CD23, while no changes were evident in vector control-, EBNA-1-, or EBNA-LP-transfected clones. EBNAs did not affect CD10, CD30, CD39, CD40, CD44, or cellular adhesion molecules. LMP1 expression in all cell lines induced growth in large clumps and expression of the cellular adhesion molecules ICAM-1, LFA-1, and LFA-3 in those cell lines which constitutively express low levels. LMP1 expression induced marked homotypic adhesion in the BJAB cell line, despite the fact that there was no significant increase in the high constitutive BJAB LFA-1 and ICAM-1 levels, suggesting that LMP1 also induces an associated functional change in these molecules. LMP1 induction of these cellular adhesion molecules was also associated with increased heterotypic adhesion to T lymphocytes. The Burkitt's lymphoma marker, CALLA (CD10), was uniformly down regulated by LMP1 in all cell lines. In contrast, LMP1 induced unique profiles of B-lymphocyte activation antigens in the various cell lines. LMP1 induced CD23 and CD39 in BJAB; CD23 in Louckes; CD39 and CD40 in BL41; and CD21, CD40, and CD44 in Daudi. In BJAB, CD23 surface and mRNA expression were markedly increased by EBNA-2 and LMP1 coexpression, compared with EBNA-2 or LMP1 alone. This cooperative effect was CD23 specific, since no such effect was observed on another marker, CD21.(ABSTRACT TRUNCATED AT 400 WORDS)

Antigens, CD↗
PubMed1990

Epstein-Barr virus types 1 and 2 differ in their EBNA-3A, EBNA-3B, and EBNA-3C genes.

The two Epstein-Barr virus (EBV) types, EBV-1 and EBV-2, are known to differ in their EBNA-2 genes, which are 64 and 53% identical in their nucleotide and predicted amino acid sequences, respectively. Restriction endonuclease maps and serologic analyses detect few other differences between EBV-1 and EBV-2 except in the EBNA-3 gene family. We determined the DNA sequence of the AG876 EBV-2 EBNA-3 coding region and have compared it with known B95-8 EBV-1 EBNA-3 sequences to delineate the extent of divergence between EBV-1 and EBV-2 isolates in their EBNA-3 genes. The B95-8 and AG876 EBV isolates had nucleotide and amino acid identity levels of 90 and 84%, 88 and 80%, and 81 and 72% for the EBNA-3A, -3B, and -3C genes, respectively. In contrast, nucleotide sequence identity in the noncoding DNA adjacent to the B95-8 and AG876 EBNA-3 open reading frames was 96%. We used the polymerase chain reaction to demonstrate that five additional EBV-1 isolates and six additional EBV-2 isolates have the type-specific differences in their EBNA-3 genes predicted from the B95-8 or AG876 sequences. Thus, EBV-1 and EBV-2 are two distinct wild-type EBV strains that have significantly diverged at four genetic loci and have maintained type-characteristic differences at each locus. The delineation of these sequence differences between EBV-1 and EBV-2 is essential to ongoing molecular dissection of the biologic properties of EBV and of the human immune response to EBV infection. The application of these data to the delineation of epitopes recognized in the EBV-immune T-cell response is also discussed.

Amino Acid Sequence↗
PubMed1989

Expression of Epstein-Barr virus transformation-associated genes in tissues of patients with EBV lymphoproliferative disease.

Epstein-Barr virus (EBV) has been associated with serious or fatal lymphoproliferative disease in immunocompromised patients. EBV nuclear protein 2 and latent membrane protein are characteristically expressed in B lymphocytes proliferating in vitro in response to growth transformation by EBV. These two proteins are thought to be effectors of lymphocyte growth since they increase the expression of B-lymphocyte activation (CD23) and cell-adhesion (LFA 3 and ICAM 1) molecules in vitro. Using monoclonal antibody-immune microscopy, we have demonstrated that these two EBV proteins and their associated B-lymphocyte activation or adhesion molecules are expressed in the infiltrating B lymphocytes in immunocompromised patients with EBV lymphoproliferative disease. These monoclonal antibodies should be useful in the early diagnosis of EBV lymphoproliferative disease and in distinguishing it from other B-lymphocyte cancers associated with EBV, such as Burkitt's lymphoma. The finding of EBV nuclear protein 2 and latent membrane protein and their associated activation or adhesion molecules provides a further pathophysiologic link between EBV and the proliferation of B lymphocytes in immunocompromised patients.

Adult↗
PubMed1988

Epstein-Barr virus latent infection membrane protein alters the human B-lymphocyte phenotype: deletion of the amino terminus abolishes activity.

A latent infection membrane protein (LMP) encoded by the Epstein-Barr virus (EBV) genome in latently infected, growth-transformed lymphocytes alters the phenotype of a human EBV-negative B-lymphoma cell line (Louckes) when introduced by gene transfer. These LMP-expressing cells exhibit increased homotypic adhesion due to increased expression of the adhesion molecules LFA-1 and ICAM-1. Increased homotypic adhesion could foster B-cell growth by facilitating autocrine growth factor effects. LFA-3 expression is also induced. The induction of LFA-3 and ICAM-1 results in increased heterotypic adhesion to T lymphocytes. This could result in more effective T-cell immune surveillance. Since LMP is expressed in EBV-transformed lymphocytes and has been demonstrated to transform rodent fibroblasts in vitro, a wide range of possible effects on B-lymphoma cell growth were assayed. In the Louckes B-lymphoma cell line, EBV LMP causes increased cell size, acid production, plasma membrane ruffling, and villous projections. Although cell proliferation rate was not greatly affected, the steady-state intracellular free calcium level, transforming growth factor beta responsiveness, and expression of the lymphocyte activation markers (CD23 and transferrin receptor) were increased. Thus, LMP appears to be a mediator of EBV effects on B-cell transformation. In transfected lymphoma cells, LMP localizes to patches at the cell periphery and associates with the cytoskeleton as it does in EBV-transformed B lymphocytes or in rodent fibroblasts. A partially deleted form of LMP (D1LMP) does not aggregate in patches or associate with the cytoskeleton and had little effect on B-cell growth. Thus, cytoskeletal association may be integral to LMP activity.

Antibodies, Monoclonal↗
PubMed1986

Expression of the Epstein-Barr virus nuclear protein 2 in rodent cells.

A 3.0-kilobase-pair Epstein-Barr virus (EBV) DNA segment necessary for lymphocyte immortalization encodes at least part of a nuclear protein (EBNA2) which is characteristically expressed in latently infected, immortalized cells. A 1.5-kilobase open reading frame within this DNA segment has now been inserted into a murine leukemia virus (MuLV)-derived expression vector (pZIP-NEO-SV(X)1) which provides for transcription of heterologous DNA but not for translational start. Transfection of the recombinant DNA into NIH 3T3 cells resulted in expression of a full-sized EBNA2 which localized to the cell nucleus. Significant new evidence is thereby provided that this 1.5 kilobase open reading frame includes a translational start site and encodes the entire EBNA2 protein. Transfection of the recombinant DNA into a helper cell line (psi am22b) providing amphotropic MuLV-packaging functions resulted in the release of a recombinant MuLV carrying the EBNA2 gene. This recombinant virus can infect rodent cells and convert them to stable EBNA2 expression. Rat-1 cells infected with the MuLV EBNA2 recombinant expressed EBNA2 and grew more rapidly in medium supplemented with 1 or 0.5% fetal calf serum than did Rat-1 cells infected with MuLV vector lacking EBNA2. The Rat-1 cells expressing EBNA2 remained contact inhibited, anchorage dependent, and nontumorigenic in nude mice. Different EBV isolates have one of at least two EBNA2 alleles. Despite divergence between the two alleles, a human serum recognized the prototype EBNA2 allele (EBNA2A) as well as the variant EBNA2B allele characteristic of some Burkitt tumor EBV isolates. The EBNA2B allele was also expressed from the MuLV-derived vector. The reproducible expression of EBNA2A or EBNA2B from these recombinant vectors will facilitate analysis of the EBNA2A and EBNA2B phenotypes.

Animals↗
PubMed1986

Control of human B-lymphocyte replication. II. Transforming Epstein-Barr virus exploits three distinct viral signals to undermine three separate control points in B-cell growth.

Highly purified resting (Go) B lymphocytes were monitored for their response to transforming Epstein-Barr virus (B95-8 strain), to a non-transforming mutant (P3HR-1) containing a deletion in the EBNA-2 coding region, and to inactivated virus of either type. All preparations induced an early appearance of two activation antigens, which included the CD23,p45 ("Blast-2') antigen. Thus, virus binding was sufficient for an initial activation step. Further change required an active viral genome. Infection with the P3HR-1 strain prompted the exit of cells out of Go but led to an arrest in the early G1 phase of the cycle. While initially showing sequels to activation indistinguishable from those observed with P3HR-1 virus, cells infected with B95-8 virus continued through G1 to express late activation antigens, enter S-phase and complete the replicative cycle. The addition of the phorbol ester TPA was found to compensate for the abortive cell cycle entry achieved with the P3HR-1 mutant, but could not supplement the minimal activation observed with inactivated virus. These findings demonstrate that the Epstein-Barr virus undermines three separate control points in the growth cycle of human B lymphocytes, and exploits three distinct viral signals to achieve this end.

Antigens, Viral↗
PubMed1986

Epstein Barr virus-specific immune defects in patients with persistent symptoms following infectious mononucleosis.

In a number of patients recovery from infectious mononucleosis (IM) following primary Epstein Barr virus (EBV) infection, is complicated by the persistence of symptoms for months or years. Normally recovery from infectious mononucleosis is associated with the development of EBV-specific antibodies and memory cytotoxic T-cells, which are present in the peripheral blood of all normal seropositive individuals. We studied four patients who had persistent symptoms for more than two years after infectious mononucleosis to determine if this abnormality was associated with a defect in EBV-specific or non-specific immune responses. All four patients had normal immunoglobulin concentrations, T- and B-cell numbers, T-cell proliferative responses and natural killer cell activity. However three of the four had reduced or absent antibodies to the EBV nuclear antigen (EBNA) although other EBV-specific antibody titres were normal. All four also had reduced EBV-specific cytotoxic T-cell activity as measured by the EBV regression assay. This defect was probably EBV-specific as alloreactive cytotoxic T-cell responses were normal. In addition, three of three patients tested had reduced in vitro antibody synthesis following pokeweed mitogen stimulation. These studies indicate that the syndrome of persistent symptoms following EBV mononucleosis may be associated with a defect in EBV-specific immunity, and thus suggest a possible immunological basis for the syndrome.

Adolescent↗
PubMed1983

Acquired hypogammaglobulinemia following infectious mononucleosis.

The case of a 17-year-old male who developed hypogammaglobulinemia following infectious mononucleosis is presented. The family history revealed that a male sibling had died some years earlier with encephalitis also following infectious mononucleosis. Laboratory investigations revealed adequate numbers of T and B cells and normal proportions of helper and suppressor T lymphocytes but poor in vitro responses to mitogens and an absence of hypersensitivity to skin test antigens. Serial serological examinations for Epstein-Barr virus antibodies indicated a primary immune response to this virus in a hypogammaglobulinemic individual. The patient probably represents a case of the X-linked lymphoproliferative syndrome. The unique feature of the present case is the demonstration of Epstein-Barr virus-specific T-cell memory. The significance of this finding and the variable expression of this syndrome in two members of the same family are discussed.

Adolescent↗
PubMed1980

Functional 'immaturity' of isolated B cells from patients with hypogammaglobulinaemia.

The membrane characteristics and in vitro function of the circulating B cells from patients with late-onset hypogammaglobulinaemia (CVID) and from patients with sex-linked hypogammaglobulinaemia (XLA) have been investigated using ox red blood cells coated with goat anti-human Ig. All seven patients with XLA had less than 2.0% SmIgM + ve cells and attempts to isolate these cells were unsuccessful. The majority of the 27 patients with CVID had greater than 3.0% SmIgM + ve cells. When cultured either with normal T cells and pokeweed mitogen (PWM) or with Epstein-Barr virus (EBV), isolated SIg + ve cells from eight patients with CVID produced significantly less IgM than normal cells and very little IgG or IgA. They also proliferated less well than normal cells and this correlated with IgM production. Stimulator capacity in the mixed lymphocyte reaction was normal in four out of seven cases. These properties of SIg + ve cells in the blood of patients with CVID are very similar to those of the normal 'immature' B cells found in adult bone marrow.

Agammaglobulinemia↗