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‣ Pathways for self-tolerance and the treatment of autoimmune diseases

Goodnow, Christopher
Fonte: Lancet Publishing Group Publicador: Lancet Publishing Group
Tipo: Artigo de Revista Científica
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Antigen delivers both immunogenic and tolerogenic signals to lymphocytes. The outcome of antigen exposure represents a complex integration of the timing of antigen binding with signals from many other immunogenic and tolerogenic costimulatory pathways. A

‣ Structural and genetic characterization of the Shigella boydii type 18 O antigen

Feng, Lu; Senchenkova, Sof'ya N; Wang, Wei; Shashkov, Alexander S; Liu, Bin; Shevelev, Sergei; Liu, Dan; Knirel, Yuriy A; Wang, Lei
Fonte: Elsevier Publicador: Elsevier
Tipo: Artigo de Revista Científica
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Shigella strains are important human pathogens and are normally identified by their O antigens. O antigen is an essential part of the lipopolysaccharide present in the outer membrane of Gram-negative bacteria and plays a role in pathogenicity. Structural

‣ Molecular analysis of Shigella boydii O1 O-antigen gene cluster and its PCR typing

Tao, Jiang; Wang, Lei; Liu, Dan; Bastin, David A; Genge, Yunqi; Feng, Lu
Fonte: NRC Research Press Publicador: NRC Research Press
Tipo: Artigo de Revista Científica
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Shigella is an important human pathogen and is closely related to Escherichia coli. O-antigen is the most variable part of the lipopolysaccharide on the cell surface of Gram-negative bacteria and plays an important role in pathogenicity. The O-antigen gen

‣ 3-[123I]iodo-alpha-methyl-L-tyrosine transport and 4F2 antigen expression in human glioma cells

Langen, Karl-Josef; Bonnie, Rosemarie; Muhlensiepen, Heinz; Jansen, Paul; Broer, Stefan; Holschbach, Marcus; Coenen, Heinz
Fonte: Elsevier Publicador: Elsevier
Tipo: Artigo de Revista Científica
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3[123I]iodo-α-methyl-L-tyrosine is a tracer of amino acid transport in brain tumors using single-photon emission-computed tomography and predominantly transported by amino acid transport system L. The 4F2 antigen has been identified to be linked to syste

‣ Bystander B cells rapidly acquire antigen receptors from activated B cells by membrane transfer

Quah, Ben; Barlow, Vaughan; McPhun, Virginia; Matthaei, Klaus; Hulett, Mark; Parish, Christopher
Fonte: National Academy of Sciences (USA) Publicador: National Academy of Sciences (USA)
Tipo: Artigo de Revista Científica
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The B cell antigen receptor (BCR) efficiently facilitates the capture and processing of a specific antigen for presentation on MHC class II molecules to antigen-specific CD4+ T cells (1). Despite this, the majority of B cells are thought to play only a li

‣ Transport of endosomal early antigen 1 in the rat sciatic nerve and location in cultured neurons

Bartlett, Selena; Reynolds, Anna; Weible, Michael; Noakes, Peter G; Hendry, Ian
Fonte: Lippincott Williams & Wilkins Publicador: Lippincott Williams & Wilkins
Tipo: Artigo de Revista Científica
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Early endosomal antigen I (EEAI) is known to be a marker of early endosomes and in cultured hippocampal neurons it preferentially localizes to the dendritic but not the axonal compartment. We show in cultured dorsal root ganglia and superior cervical gang

‣ Role of Syk in B-cell development and antigen-receptor signaling

Cornall, Richard J; Cheng, Alex; Pawson, Tony; Goodnow, Christopher
Fonte: National Academy of Sciences (USA) Publicador: National Academy of Sciences (USA)
Tipo: Artigo de Revista Científica
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Antigen receptors (BCRs) on developing B lymphocytes play two opposing roles-promoting survival of cells that may later bind a foreign antigen and inhibiting survival of cells that bind too strongly to self-antigens. It is not known how these opposing outcomes are signaled by BCRs on immature B cells. Here we analyze the effect of a null mutation in the Syk tyrosine kinase on maturing B cells displaying a transgene-encoded BCR that binds hen egg lysozyme (HEL). In the absence of HEL antigen, HEL-specific BCRs are expressed normally on the surface: of Syk-deficient immature B-lineage cells, but this fails to promote maturation beyond the earliest stages of B-lineage commitment. Binding of HEL antigen, nevertheless, triggers phosphorylation of CD79α/β BCR subunits and modulation of receptors from the surface in Syk- deficient cells, but it cannot induce an intracellular calcium response. Continuous binding of low- or high-avidity forms of HEL, expressed as self- antigens, fails to restore the signal needed for maturation. Compared with the effects in the same system of null mutations in other BCR signaling elements, such as CD45 and Lyn kinase, these results indicate that Syk is essential for transmitting a signal that initiates the program of B- lymphocyte maturation.

‣ Comparison of whole gene and whole virus scrambled antigen approaches for DNA prime and fowlpox virus boost HIV type 1 vaccine regimens in macaques

Pamungkas, Joko; De Rose, Robert; Iskandriati, Diah; Noviana, Rachmitasari; Paramastri, Yasmina; Dale, C Jane; Shoobridge, Maryanne; Medveczky, C; Ramshaw, Ian; Thomson, Scott; Kent, Stephen J
Fonte: Mary Ann Liebert Inc. Publicador: Mary Ann Liebert Inc.
Tipo: Artigo de Revista Científica
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T cell immunity plays a critical role in controlling HIV-1 viremia, and encoding a limited set of HIV-1 genes within DNA and poxvirus vectors can, when used sequentially, induce high levels of T cell immunity in primates. However, a limited breadth of T cell immunity exposes the host to potential infection with either genetically diverse HIV-1 strains or T cell escape variants of HIV-1. In an attempt to induce maximally broad immunity, we examined DNA and recombinant fowlpox virus (rFPV) vaccines encoding all HIV-1 genes derived from a global HIV-1 consensus sequence, but expressed as multiple overlapping scrambled 30-amino acid segments (scrambled antigen vaccines, or SAVINEs). Three groups of seven pigtail macaques were immunized with sets of DNA and rFPV expressing Gag/Pol antigens only, the whole genome SAVINE antigens, or no HIV-1 antigens and T cell immunity was monitored by ELISpot and intracellular cytokine staining. High levels of cross-subtype HIV-specific T cell immunity to Gag were consistently induced in the seven macaques primed with DNA and rFPV vaccines expressing Gag/Pol as intact proteins. It was, however, difficult to repeatedly boost immunity with further rFPV immunizations, presumably reflecting high levels of anti-FPV immunity. Unfortunately...

‣ Targeting dendritic cells with antigen-containing liposomes: A highly effective procedure for induction of antitumor immunity and for tumor immunotherapy

Van Broekhoven, Christina; Parish, Christopher; Demangel, Caroline; Britton, Warwick; Altin, Joseph
Fonte: American Association for Cancer Research Publicador: American Association for Cancer Research
Tipo: Artigo de Revista Científica
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Dendritic cells (DCs) are potent stimulators of immunity, and DCs pulsed with tumor antigen ex vivo have applications in tumor immunotherapy. However, DCs are a small population of cells, and their isolation and pulsing with antigen can be impractical. Here we show that a crude preparation of plasma membrane vesicles (PMV) from the highly metastatic murine melanoma (B16-OVA) and a surrogate tumor antigen (OVA) can be targeted directly to DCs in vivo to elicit functional effects. A novel metal-chelating lipid, 3(nitrilotriacetic acid)-ditetradecylamine, was incorporated into B16-OVA-derived PMV, allowing recombinant hexahistidine-tagged forms of single chain antibody fragments to the DC surface molecules CD11c and DEC-205, to be conveniently " engrafted" onto the vesicle surface by metal-chelating linkage. The modified PMV, or similarly engrafted synthetic stealth liposomes containing OVA or OVA peptide antigen, were found to target DCs in vitro and in vivo, in experiments using flow cytometry and fluorescence confocal microscopy. When used as vaccines in syngeneic mice, the preparations stimulated strong B16-OVA-specific CTL responses in splenic T cells and a marked protection against tumor growth. Protection was dependent on the simultaneous delivery of both antigen and a DC maturation or "danger signal" signal (IFN-γ or lipopolysaccharide). Administration of the DC-targeting vaccine to mice challenged with B16-OVA cells induced a dramatic immunotherapeutic effect and prolonged disease-free survival. The results show that the targeting of antigen to DCs in this way is highly effective at inducing immunity and protection against the tumor...

‣ Scaffolding of antigen receptors for immunogenic versus tolerogenic signaling

Jun, Jesse; Goodnow, Christopher
Fonte: Nature Publishing Group Publicador: Nature Publishing Group
Tipo: Artigo de Revista Científica
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Lymphocyte antigen receptors are responsible for inducing the opposite responses of immunity or tolerance. How the correct polarity of antigen receptor signaling is encoded has been an enduring enigma. Here we summarize recent advances defining key scaffolding molecules, CARMA1 (also known as CARD11) and the Cbl family of ubiquitin ligases, required for either immunogenic or tolerogenic signaling by antigen receptors. These scaffolding proteins may determine the polarity of response to antigen by promoting assembly around antigen receptors of competing multiprotein signal complexes: immunosomes versus tolerosomes. Each of the factors that influence immunogenicity or tolerogenicity-stage of lymphocyte differentiation, concurrent engagement of inhibitory or costimulatory receptors, extent of receptor crosslinking, and prior antigen experience-may be integrated in lymphocytes through their capacity to influence the probability of assembling immunosomes versus tolerosomes.

‣ Antigen-specific systemic and reproductive tract antibodies in foxes immunized with salmonella typhimurium expressing bacterial and sperm proteins

Bird, Phillip I; Verma, Naresh; Bradley, Mark P
Fonte: CSLI Publications Publicador: CSLI Publications
Tipo: Artigo de Revista Científica
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Attenuated Salmonella typhimurium strains are potential 'safe' delivery vectors of an oral immunocontraceptive vaccine for the European red fox (Vulpes vulpes). In the present study, model bacterial (Escherichia coli heat-labile enterotoxin B subunit, LTB) and fox sperm (fSP10) antigens were expressed in S. typhimurium SL3261 (ΔaroA) under the control of the trc promoter. Adult female foxes were given three oral immunizations with SL3261 containing either LTB (SL3261/pLTB), fSP10 (SL3261/pFSP10) or a control plasmid (pKK233-2 or pTrc99A). All foxes raised serum (IgG) and vaginal (IgG and IgA) antibodies against S. typhimurium lipopolysaccharide (LPS). Each fox that received SL3261/pLTB raised high titre LTB-specific serum and vaginal IgG antibodies. However, only one of four foxes immunized with SL3261/pFSP10 raised an anti-fSP10 immune response, in the form of low titre serum and vaginal IgG antibodies. No vaginal IgA antibodies were raised against either LTB or fSP10 in these experiments. The immune responses against recombinant LTB and fSP10 resulted chiefly from the initial dose of antigen in the inocula and were minimally influenced by continued in vivo antigen expression. This study demonstrates for the first time in the female red fox that oral Salmonella can elicit specific systemic and reproductive tract antibodies against heterologous...

‣ Role of flanking sequences and phosphorylation in the recognition of the simian-virus-40 large T-antigen nuclear localization sequences by importin-a

Fontes, Marcos; Teh, Trazel; Toth, Gabor; John, Anna; Pavo, Imre; Jans, David A; Kobe, Bostjan
Fonte: Portland Press Publicador: Portland Press
Tipo: Artigo de Revista Científica
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The nuclear import of simian-virus-40 large T-antigen (tumour antigen) is enhanced via phosphorylation by the protein kinase CK2 at Ser112 in the vicinity of the NLS (nuclear localization sequence). To determine the structural basis of the effect of the sequences flanking the basic cluster KKKRK, and the effect of phosphorylation on the recognition of the NLS by the nuclear import factor importin-α (Impa), we co-crystallized non-autoinhibited Impa with peptides corresponding to the phosphorylated and non-phosphorylated forms of the NLS, and determined the crystal structures of the complexes. The structures show that the amino acids N-terminally flanking the basic cluster make specific contacts with the receptor that are distinct from the interactions between bipartite NLSs and Impα. We confirm the important role of flanking sequences using binding assays. Unexpectedly, the regions of the peptides containing the phosphorylation site do not make specific contacts with the receptor. Binding assays confirm that phosphorylation does not increase the affinity of the T-antigen NLS to Impα. We conclude that the sequences flanking the basic clusters in NLSs play a crucial role in nuclear import by modulating the recognition of the NLS by Impα...

‣ The Novel Chelator lipid 3(nitrilotriacetic acid)-ditetradecylamine (NTA3-DTDA) promotes stable binding of His-tagged proteins to liposomal membranes: Potent anti-tumor responses induced by simultaneously targeting antigen, cytokine and costimulatory signals to T cells.

Van Broekhoven, Christina; Altin, Joseph
Fonte: Elsevier Publicador: Elsevier
Tipo: Artigo de Revista Científica
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Recent studies indicate that the chelator lipid nitrilotriacetic acid ditetradecylamine (NTA-DTDA) can be used to engraft T cell costimulatory molecules onto tumor cell membranes, potentially circumventing the need for genetic manipulation of the cells for development of cell- or membrane-based tumor vaccines. Here, we show that a related lipid 3(nitrilotriacetic acid)-ditetradecylamine (NTA3-DTDA, which has three NTA moieties in its headgroup instead of one) is several-fold more effective than NTA-DTDA at promoting stable His-tagged protein engraftment. IAsys biosensor studies show that binding of His-tagged B7.1 (B7.1-6H) to NTA3-DTDA-containing membranes, exhibit a faster on-rate and a slower off-rate, compared to membranes containing NTA-DTDA. Also, NTA3-DTDA-containing liposomes and plasma membrane vesicles (PMV) engrafted with B7.1-6H and CD40-6H exhibit greater binding to T cells, in vitro and in vivo. Engrafted NTA3-DTDA- containing PMV encapsulated cytokines such as IL-2, IL-12, GM-CSF and IFN-γ, allowing targeted delivery of both antigen and cytokine to T cells, and stimulation of antigen-specific T cell proliferation and cytotoxicity. Importantly, use of B7.1-CD40-engrafted PMV containing IL-2 and IL-12 as a vaccine in DBA/2J mice induced protection against challenge with syngeneic tumor cells (P815 mammary mastocytoma)...

‣ Resistance to CpG DNA-induced autoimmunity through tolerogenic B cell antigen receptor ERK signaling

Rui, Lixin; Garcia De Vinuesa, Maria Carola; Blasioli, Julie; Goodnow, Christopher
Fonte: Nature Publishing Group Publicador: Nature Publishing Group
Tipo: Artigo de Revista Científica
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CpG sequences in self-DNA are an important potential trigger for autoantibody secretion in systemic lupus and other systemic autoimmune disorders. It is not known how this ubiquitous threat may be controlled by active mechanisms for maintaining self tolerance. Here we show that two distinct mechanisms oppose autoantibody secretion induced by CpG DNA in anergic B cells that are constantly binding self-antigen. Uncoupling of the antigen receptor (BCR) from a calcineurin-dependent pathway prevents signals that synergize with CpG DNA for proliferation. The BCR does not become desensitized by activating the extracellular response kinase (ERK) MAP kinase pathway, however, and continuous self-antigen signaling to ERK inhibits CpG DNA-induced plasma cell differentiation. These two mechanisms seem to act as a general control against autoantibody production elicited by Toll-like receptors, and their regulation of T cell-independent responses to Toll-like receptor 9 (TLR9) is probably crucial for resistance to systemic autoimmunity.

‣ Intestinal exposure to a parasite antigen in utero depresses cellular and cytokine responses of the mucosal immune system

Paalangara, R; McClure, Susan J; McCullagh, Peter
Fonte: Elsevier Publicador: Elsevier
Tipo: Artigo de Revista Científica
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The response of the mucosal immune system of 4-6-week old lambs to viable Trichostrongylus colubriformis larvae was compared in two groups of animals, one exposed to T. colubriformis antigen and the other to saline while in utero. Exposure to larval antigen two-thirds of the way through gestation resulted in significant reduction in the frequency of jejunal goblet cells and of ileal eosinophils, CD1b+ antigen-presenting cells and CD4+, CD5+ and CD8+ cells. However, it resulted in a significant increase in the jejunal CD8+ response to postnatal challenge. The expression of the cytokines TNF-α and IL-1β in the ileum, and of jejunal NSE, was significantly reduced by in utero exposure, whereas those of jejunal TNF-α and ileal TGF-β were increased. The observed changes in cellular and cytokine responses to challenge with viable larvae, in those lambs previously exposed in utero, indicated that the intestinal mucosal immune system remains susceptible to down-regulation until considerably later in foetal development than is the case for other components of the immune system.

‣ Functional inhibitory human leucocyte antigen class I receptors on natural killer (NK) cells in patients with chronic NK lymphocytosis

Warren, Hilary; Christiansen, Frank; Witt, Campbell
Fonte: Blackwell Publishing Ltd Publicador: Blackwell Publishing Ltd
Tipo: Artigo de Revista Científica
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Chronic natural killer (NK) lymphocytosis is a rare disorder characterized by an indolent clinical course. Despite high NK cell numbers, many patients present with only mild clinical symptoms, and are often asymptomatic. NK cells are equipped with a range of receptors that bind human leucocyte antigen (HLA)-class I molecules. The killer immunoglobulin-like receptors (KIR, CD158) bind groups of HLA alleles, the CD94/NKG2 receptors bind HLA-E, and the CD85j (ILT2, LIR-1) receptor binds to the relatively non-polymorphic α3 domain of HLA molecules. Inhibitory HLA class I receptors silence NK cells against cells expressing normal levels of HLA class I. Analysis of NK cells in six patients with chronic NK lymphocytosis revealed a high level of the inhibitory CD94/NKG2A receptor on all NK cells. In four patients, KIR were absent, in one patient a single KIR was expressed in the absence of self-ligand, and in one patient CD85j and multiple KIR were expressed. Cytotoxicity assays demonstrated that all HLA class I receptors were functional. The ability of monoclonal antibodies to block the receptors and allow killing of autologous target cells established that both receptor and ligand expression were adequate for inhibitory function. We propose that the silent behaviour of NK cells in patients with chronic NK lymphocytosis is due to effective inhibitory HLA class I receptors.

‣ Dendritic cells infected with a vaccinia virus interleukin-2 vector secrete high levels of IL-2 and can become efficient antigen presenting cells that secrete high levels of the immunostimulatory cytokine IL-12

Mukherjee, Sutapa; Upham, John; Ramshaw, Ian; Bundell, Christine; van Bruggen, Ivonne; Robinson, Bruce; Nelson, Delia
Fonte: Nature Publishing Group Publicador: Nature Publishing Group
Tipo: Artigo de Revista Científica
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Dendritic cell (DC) therapies using DC presenting tumor antigen/s can induce CD8+ CTL that mediate tumor eradication, nonetheless many patients remain unresponsive. Thus, cytokine gene vectors applied to DC may amplify these responses. Herein, we examined the responses that monocyte-derived DC (at different maturational stages) make when infected with a vaccinia virus-interleukin-2 (VV-IL-2) vector in vitro. VV-IL-2-infected DC secreted significant levels of bioactive IL-2 and maintained their antigen presentation function. However, we show that DC are exquisitely sensitive to their local antigenic microenvironment, and that responses generated by one antigen can be altered by another. VV-IL-2 infection of immature DC led to DC activation (upregulation of CD80, CD86 and class II surface molecules) when the virus was propagated through xenogeneic, but not syngeneic, mammalian cells; these DC secreted IL-10 and tumor necrosis factor-α (TNF-α), but not IL-12. In contrast, after VV-IL-2 infection (regardless of their mammalian cellular context), IFNγ/LPS-matured DC inevitably downregulated their antigen presenting machinery. In conclusion, immunostimulatory DC can be generated by VV-IL-2, but this depends upon (i) infecting immature DC only...

‣ Functional analysis of the O antigen glucosylation gene cluster of Shegella Flexneri bacteriophage SfX

Guan, S; Bastin, David A; Verma, Naresh
Fonte: Consultants Bureau Publicador: Consultants Bureau
Tipo: Artigo de Revista Científica
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Previous studies have shown that Shigella flexneri bacteriophage X (SfX) encodes a glucosyltransferase (GtrX, formerly Gtr), which is involved in O antigen modification (serotype Y to serotype X). However, GtrX alone can only mediate a partial conversion. More recently, a three-gene cluster has been identified next to the attachment site in the genome of two other S. flexneri bacteriophages (i.e. SfV and SfII). This gene cluster was postulated to be responsible for a full O antigen conversion. Here it is reported that besides the gtrX gene, the other two genes in the gtr locus of SfX were also involved in the O antigen modification process. The first gene in the cluster (gtrA) encodes a small highly hydrophobic protein which appears to be involved in the translocation of lipid-linked glucose across the cytoplasmic membrane. The second gene in the cluster (gtrB) encodes an enzyme catalysing the transfer of the glucose residue from UDP-glucose to a lipid carrier. The third gene (gtrX) encodes a bacteriophage-specific glucosyltransferase which is largely responsible for the final step, i.e. attaching the glucosyl molecules onto the correct sugar residue of the O antigen repeating unit. A three-step model for the glucosylation of bacterial O antigen has been proposed.

‣ Antigen presentation in Syrian hamster cells: substrate selectivity of TAP controlled by polymorphic residues in TAP1 and differential requirements for loading of H2 class l molecules

Lobigs, Mario; Mullbacher, Arno; Blanden, Robert; Hammerling, G; Momburg, Frank
Fonte: Springer Publicador: Springer
Tipo: Artigo de Revista Científica
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Expression of mouse major histocompatibility complex (MHC) class I molecules in different cell lines derived from Syrian hamsters has revealed antigen presentation deficiencies of some H2 allelic products in two cell lines (BHK and NIL-2) which were overcome by transient expression of the rat transporter associated with antigen processing. Here we show that in both cell lines the endogenous MHC class I cell surface expression was completely down-regulated. Lymphokine treatment induced endogenous and recombinant mouse MHC class I cell surface expression to levels similar to that in other Syrian hamster cell lines competent for antigen presentation through transduced H2 molecules. Accordingly, constitutive downregulation of expression of accessory molecules of the MHC class I pathway can reveal differences between H2 class I alleles in antigen presentation not encountered when the expression levels are augmented. In addition to the differential expression of MHC class I pathtway genes, two cell lines representing competent (FF) and defective (BHK) antigen presentation phenotypes for mouse class I MHC restriction elements demonstrated substantial sequence polymorphism in Tap1 but not Tap2. Cytokine-treated FF or BHK cells and human TAP-deficient T2 cells transfected with FF or BHK TAP1 in combination with FF TAP2 differed in their preference for C-terminal peptide residues...

‣ Modulation of transporter associated with antigen processing (TAP)-mediated peptide import into the endoplasmic reticulum by flavivirus infection

Momburg, Frank; Mullbacher, Arno; Lobigs, Mario
Fonte: American Society for Microbiology Publicador: American Society for Microbiology
Tipo: Artigo de Revista Científica
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In contrast to many other viruses that escape the cellular immune response by downregulating major histocompatibility complex (MHC) class I molecules, flavivirus infection can upregulate their cell surface expression. Previously we have presented evidence that during flavivirus infection, peptide supply to the endoplasmic reticulum is increased (A. Müllbacher and M. Lobigs, Immunity 3:207-214, 1995). Here we show that during the early phase of infection with different flaviviruses, the transport activity of the peptide transporter associated with antigen processing (TAP) is augmented by up to 50%. TAP expression is unaltered during infection, and viral but not host macromolecular synthesis is required for enhanced peptide transport. This study is the first demonstration of transient enhancement of TAP-dependent peptide import into the lumen of the endoplasmic reticulum as a consequence of a viral infection. We suggest that the increased supply of peptides for assembly with MHC class I molecules in flavivirus-infected cells accounts for the upregulation of MHC class I cell surface expression with the biological consequence of viral evasion of natural killer cell recognition.