Altered T cell surface glycosylation in HIV-1 infection results in increased susceptibility to galectin-1-induced cell death.
(73/292)
The massive T cell death that occurs in HIV type 1 (HIV-1) infection contributes profoundly to the pathophysiology associated with AIDS. The mechanisms controlling cell death of both infected and uninfected T cells ("bystander" death) are not completely understood. We have shown that HIV-1 infection of T cells results in altered glycosylation of cell surface glycoproteins; specifically, it decreased sialylation and increased expression of core 2 O-glycans. Galectin-1 is an endogenous human lectin that recognizes these types of glycosylation changes and induces cell death of activated lymphocytes. Therefore we studied the possible contribution of galectin-1 in the pathophysiology of AIDS. O-glycan modifications were investigated on peripheral lymphocytes from AIDS patients. Oligosaccharides from CD43 and CD45 of CEM cells latently infected with HIV-1 were chemically analyzed. Consistent with our previous results, we show that HIV-1 infection results in accumulation of exposed lactosamine residues, oligosaccharides recognized by galectin-1 on cell surface glycoproteins. Both latently HIV-1-infected T cell lines and peripheral CD4 and CD8 T cells from AIDS patients exhibited exposed lactosamine residues and demonstrated marked susceptibility to galectin-1-induced cell death, in contrast to control cultures or cells from uninfected donors. The fraction of cells that died in response to galectin-1 exceeded the fraction of infected cells, indicating that death of uninfected cells occurred. Altered cell surface glycosylation of T cells during HIV-1 infection increases the susceptibility to galectin-1-induced cell death, and this death pathway can contribute to loss of both infected and uninfected T cells in AIDS. (+info)
CpG DNA induces a class II transactivator-independent increase in class II MHC by stabilizing class II MHC mRNA in B lymphocytes.
(74/292)
Microbial products, such as CpG DNA and LPS, enhance class II MHC (MHC-II) expression and Ag presentation by dendritic cells, but this effect does not occur with macrophages and is largely unexplored in B cells. Although MHC-II expression is influenced by transcriptional regulation, which is governed by class II transactivator (CIITA) in all cells, microbial products enhance MHC-II expression by dendritic cells in part by increasing MHC-II protein stability. In this study, we show that the CpG-induced increase in MHC-II expression by B lymphocytes is not due to protein stabilization or changes in CIITA expression or activity, but instead is due to increased stability of MHC-II mRNA. This CIITA-independent mechanism adds a new layer of complexity to regulation of MHC-II and may increase T cell help for B cell Ab responses to microbial or vaccine Ags. (+info)
Modeling TCR signaling complex formation in positive selection.
(75/292)
T cell receptor signaling in the thymus can result in positive selection, and hence progressive maturation to the CD4(+)8(-) or CD4(-)8(+) stage, or induction of apoptosis by negative selection. Although it is poorly understood how TCR ligation at the CD4(+)8(+) stage can lead to such different cell fates, it is thought that the strength of signal may play a role in determining the outcome of TCR signaling. In this study, we have characterized the formation of an active signaling complex in thymocytes undergoing positive selection as a result of interaction with thymic epithelial cells. Although this signaling complex involves redistribution of cell surface and intracellular molecules, reminiscent of that observed in T cell activation, accumulation of GM1-containing lipid rafts was not observed. However, enforced expression of the costimulatory molecule CD80 on thymic epithelium induced GM1 polarization in thymocytes, and was accompanied by reduced positive selection and increased apoptosis. We suggest that the presence or absence of CD80 costimulation influences the outcome of TCR signaling in CD4(+)8(+) thymocytes through differential lipid raft recruitment, thus determining overall signal strength and influencing developmental cell fate. (+info)
T cell aggregation induced through CD43: intracellular signals and inhibition by the immunomodulatory drug leflunomide.
(76/292)
The CD43 coreceptor molecule has been shown to participate in lymphocyte adhesion and activation. Leukocyte homotypic aggregation results from a cascade of intracellular signals delivered to the cells upon engagement of different cell-surface molecules with their natural ligands. This phenomenon requires an active metabolism, reorganization of the cytoskeleton, and relocalization of cell-surface molecules. The aim of this study was to identify some of the key members of the signaling cascade leading to T lymphocyte homotypic aggregation following CD43 engagement. CD43-mediated homotypic aggregation of T lymphocytes required the participation of Src kinases, phospholipase C-gamma2, protein kinase C, phosphatidylinositol-3 kinase, as well as extracellular-regulated kinase 1/2 and p38. Data shown here suggest that these signaling molecules play a central role in regulating actin cytoskeleton remodeling after CD43 ligation. We also evaluated the ability of immunomodulatory drugs such as leflunomide to block the CD43-mediated homotypic aggregation. Leflunomide blocked the recruitment of targets of the Src family kinases as well as actin polymerization, diminishing the ability of T lymphocytes to aggregate in response to CD43-specific signals, suggesting that this drug might control the migration and recruitment of lymphoid cells to inflamed tissues. (+info)
Expression cloning of a cDNA encoding UDP-GlcNAc:Gal beta 1-3-GalNAc-R (GlcNAc to GalNAc) beta 1-6GlcNAc transferase by gene transfer into CHO cells expressing polyoma large tumor antigen.
(77/292)
A cDNA encoding UDP-GlcNAc:Gal beta 1-3GalNAc-R (GlcNAc to GalNAc) beta 1-6GlcNAc transferase (EC 2.4.1.102), which forms critical branches in O-glycans, has been isolated by an expression cloning approach using Chinese hamster ovary (CHO) cells. Increased activity of this enzyme and the concomitant occurrence of the O-glycan core 2 structure [Gal beta 1-3(GlcNAc beta 1-6)GalNAc] has been observed in a variety of biological processes, such as T-cell activation and immunodeficiency due to the Wiskott-Aldrich syndrome and AIDS. Since CHO cells do not express this enzyme, CHO cell lines were established to stably express polyoma large tumor (T) antigen, which enables transient expression cloning. Because the antibody used was found to detect most efficiently the oligosaccharide products attached to leukosialin, the CHO cells were also stably transfected with leukosialin cDNA. By using this particular CHO cell line, a cDNA that encodes a protein determining the formation of the core 2 structure was isolated from an HL-60 cDNA library. The cDNA sequence predicts a protein with type II membrane topology, as has been found for all other mammalian glycosyltransferases cloned to date. The expression of the presumed catalytic domain as a fusion protein with the IgG binding domain of protein A enabled us to demonstrate unequivocally that the cDNA encodes the core 2 beta-1,6-N-acetylglucosaminyltransferase, the enzyme responsible for the formation of Gal beta 1-3(GlcNAc beta 1-6)GalNAc structures. No activity with this enzyme was detected toward the acceptors for other beta 1-6GlcNAc transferases. (+info)
Detection and localization of Epstein-Barr viral genomes in angioimmunoblastic lymphadenopathy and angioimmunoblastic lymphadenopathy-like lymphoma.
(78/292)
We studied 23 cases of angioimmunoblastic lymphadenopathy (AILD) and AILD-like lymphoma for evidence of Epstein-Barr virus (EBV) using the polymerase chain reaction (PCR) and in situ hybridization studies. EBV nucleic acid sequences were found by either PCR or in situ hybridization in 96% of the cases. There was a wide range in the number of EBV-positive cells among the different cases as detected by in situ hybridization. The EBV-positive cells most often possessed nuclei of intermediate to large size. Double-labeling immunohistochemistry/in situ hybridization studies demonstrated that most of the EBV-positive cells expressed the B-lineage antigen CD20 (as detected by L26), with a minority of the EBV-positive cells stained for the T-lineage associated antigen, CD43 (as detected by Leu 22). The abnormally high amounts of EBV found in AILD and AILD-like lymphoma may be a reflection of decreased immunocompetence in these patients. The presence of EBV-positive B cells may explain the presence of B-cell clones found by others as well as the paradoxical occurrence of B-cell lymphoma in a primary T-cell lymphoproliferative disorder. (+info)
Poly-N-acetyllactosaminyl O-glycans attached to leukosialin. The presence of sialyl Le(x) structures in O-glycans.
(79/292)
Poly-N-acetyllactosamine extension has been found in O-glycans in addition to N-glycans and glycosphingolipids. Attempts were made in HL-60 and K562 cells to determine the amount of poly-N-acetyllactosaminyl O-glycans in the major sialoglycoprotein, leukosialin. Leukosialin was immunoprecipitated from [3H]glucosamine-labeled HL-60 and K562 cells. Glycopeptides were prepared by Pronase digestion, and O-glycan-containing glycopeptides were isolated by affinity chromatography using Jacalin-agarose. The glycopeptides bound to Jacalin-agarose and those unbound were treated with alkaline borohydride, and the released O-glycans were fractionated by Bio-Gel P-4 filtration. Sequential glycosidase digestion of the O-glycans, with or without pretreatment by fucosidase or neuraminidase, revealed the following conclusions. 1) Leukosialin from HL-60 cells contains about 1-2 poly-N-acetyllactosaminyl O-glycan chains/molecule. 2) About 50% of these poly-N-acetyllactosaminyl O-glycans contain sialyl Le(x) termini, NeuNAc alpha 2-->3Gal beta 1-->4 (Fuc alpha 1-->3)GlcNAc beta 1-->R. The amount of sialyl Le(x) structure in leukosialin is roughly equivalent to that on cell surfaces of HL-60 cells. 3) Leukosialin from K562 cells, on the other hand, contains no detectable amount of poly-N-acetyllactosaminyl O-glycans. 4) The presence of poly-N-acetyllactosamine in O-glycans is dependent on the core 2 beta 1,6-N-acetylglucosaminyl transferase. 5) Jacalin-agarose binds to sialylated small oligosaccharides such as NeuNAc alpha 2-->3Gal beta 1-->3(NeuNAc alpha 2-->6) GalNAc but not the hexasaccharide NeuNAc alpha 2-->3Gal beta 1-->3(NeuNAc alpha 2-->3Gal beta 1-->4GlcNAc beta 1-->6) GalNAc. These results indicate that the formation of polylactosaminyl O-glycans and sialyl Le(x) structure in O-glycans is dependent on the core 2 formation. (+info)
Differential expression of ezrin/radixin/moesin (ERM) and ERM-associated adhesion molecules in the blastocyst and uterus suggests their functions during implantation.
(80/292)
Development of the blastocyst to implantation competency, differentiation of the uterus to the receptive state, and a cross talk between the implantation-competent blastocyst and the uterine luminal epithelium are all essential to the process of implantation. In the present investigation, we examined the possibility for a potential cross talk between the blastocyst and uterus involving the ezrin/radixin/moesin (ERM) proteins and ERM-associated cytoskeletal cross-linker proteins CD43, CD44, ICAM-1, and ICAM-2. In normal Day 4 blastocysts and after rendering dormant blastocysts to implantation-competent by estrogen in vivo (activated), the outer surface of mural trophectoderm cells showed much higher levels of radixin as compared to those in the polar trophectoderm cells, inner cell mass (ICM), and primitive endoderm. In contrast, ezrin was present on both the mural and the polar trophectoderm cell surfaces of normal Day 4 and activated blastocysts at higher intensity than dormant blastocysts. A distinct localization was noted in the primitive endoderm of dormant blastocysts that was not apparent in activated or normal Day 4 blastocysts. The expression of moesin was modestly higher at the mural trophectoderm of implantation-competent blastocysts, while the localization appeared to be present primarily on the polar trophectoderm cell surface of Day 4 blastocysts. The localization of ERM-associated adhesion molecules CD43, CD44, and ICAM-2 was more intense in the implantation-competent blastocysts compared with the dormant blastocysts. However, while CD44 was present both in the trophectoderm and in ICM, CD43 and ICAM-2 were localized primarily to the trophectoderm. The signal for ICAM-1 was very intense in the ICM but was modest in the trophectoderm. No significant changes in fluorescence intensity were noted between activated and dormant blastocysts. In the receptive uterus on Day 4 of pregnancy, ERM proteins were localized to the uterine epithelium, while on Day 5 the localization, especially of radixin and moesin, extended to the stroma surrounding the implantation chamber. With respect to ERM-associated adhesion molecules, while CD44 and ICAM-1 were exclusively localized in the stroma on Day 4, CD43 and ICAM-2 were localized to the epithelium. On Day 5, the localization of CD44 and ICAM-1 became highly concentrated in the antimesometrial stroma of the implantation chamber. The localization of CD43 and ICAM-2 remained mostly epithelial, although some stromal localization of CD43 was noted on Day 5. These results suggest that differential expression and distribution of ERM proteins and ERM-associated adhesion molecules are involved in the construction of the cellular architecture necessary for blastocyst activation and uterine receptivity leading to successful implantation. (+info)