Excessive degradation of intracellular protein in macrophages prevents presentation in the context of major histocompatibility complex class II molecules

The endogenous major histocompatibility complex (MHC) class II presentation pathway allows biosynthesized, intracellular antigens access for presentation to MHC class II-restricted T cells. This pathway has been well documented in B cells and fibroblasts, but may not be universally available in all antigen-presenting cell types. This study compares the ability of different antigen-presenting cells, expressing endogenous C5 protein (fifth component of mouse complement) as a result of transfection, to present their biosynthesized C5 to MHC class II-restricted T cells. B cells and fibroblasts expressing C5 were able to present several epitopes of this protein with MHC class II molecules, whereas macrophages were unable to do so, but readily presented C5 from an extracellular source. However, macrophage presentation of endogenous C5 could be achieved when they were treated with low doses of the lysosomotropic agent ammonium chloride. In the presence of an inhibitor of autophagy, presentation of endogenous C5 was abrogated, indicating that biosynthesized C5 is shuttled into lysosomal compartments for degradation before making contact with MHC class II molecules. Taken together, this suggests that proteolytic activity in lysosomes of macrophages may be excessive, compared with fibroblasts and B cells, and destroys epitopes of the C5 protein before they can gain access to MHC class II molecules. Thus, there are inherent differences in presentation pathways between antigen-presenting cell types; this could reflect their specialized functions within the immune system with macrophages focussing preferentially on internalization, degradation, and presentation of extracellular material.     

Invariant chain cleavage and peptide loading in major histocompatibility complex class II vesicles

B lymphocytes contain a novel population of endocytic vesicles involved in the transport of newly synthesized major histocompatibility complex (MHC) class II alpha beta chains and alpha beta peptide complexes to the cell surface. We now present evidence that these class II-enriched vesicles (CIIV) are also likely to be a site for the loading of immunogenic peptides onto MHC molecules. We used the serine protease inhibitor leupeptin to accumulate naturally occurring intermediates in the degradation of alpha beta-invariant chain complexes and to slow the intracellular transport of class II molecules. As expected, leupeptin caused an accumulation of Ii chain and class II molecules (I-A(d)) in endosomes and lysosomes. More importantly, however, it enhanced the selective accumulation of a 10-kD invariant chain fragment associated with sodium dodecyl sulfate (SDS)-labile (empty) alpha beta dimers in CIIV. This was followed by the dissociation of the 10-kD fragment, formation of SDS-stable (peptide-loaded) alpha beta dimers, and their subsequent appearance at the cell surface. Thus, CIIV are likely to serve as a specialized site, distinct from endosomes and lysosomes, that hosts the final steps in the dissociation of invariant chain from class II molecules and the loading of antigen-derived peptides onto newly synthesized alpha beta dimers.     

Differentiation-stage specific self-peptides bound by major histocompatibility complex class I molecules

We have tested the hypothesis that phenotypic changes of development are accompanied by expression of differentiation-stage specific peptides bound to major histocompatibility complex (MHC) class I molecules. The U937 cell line, when cultured in the presence of phorbol myristate acetate (PMA), undergoes differentiation from monoblasts to macrophage-like cells. The high-performance liquid chromatography profile of peptides eluted from purified human histocompatibility leukocyte antigen class I molecules expressed by U937 treated with PMA differs from that obtained from control, untreated U937 cells. Chemical sequencing of eluted peptides identified a peptide derived from cytomegalovirus in both treated and untreated cells. PMA-treated, but not untreated cells, displayed an additional peptide derived from interleukin 1 beta. Hence, differentiation-induction of U937 is accompanied by the presentation of at least one differentiation-stage specific peptide. Our results indicate that, similar to viral infection, cellular development and transformation is accompanied by the de novo synthesis of proteins which are processed and presented on MHC class I molecules.     

Bile regulates the expression of major histocompatibility complex class II molecules on rat intestinal epithelium

BACKGROUND & AIMS: Major histocompatibility complex (MHC) class II molecules are expressed on intestinal epithelial cells, and the intensity of this expression is regulated. The aim of this study was to test the hypothesis that bile regulates the expression of MHC class II molecules on intestinal epithelium. METHODS: Rats were deprived of intestinal bile by external drainage for 24 or 48 hours, and their intestines were collected, sectioned, and stained with the anti-MHC class II monoclonal antibodies OX4 and OX6. For one group of rats, bile flow was deviated from its usual entry point to the ileum. RESULTS: Compared with intact animals, MHC class II expression was observed to be diminished within 24 hours and totally absent after 48 hours of bile drainage. For the group in which bile flow was deviated to the ileum, staining was only observed in the region distal to the entry point. Analysis by bioassay and enzyme-linked immunosorbent assay of bile showed the presence of tumor necrosis factor and interferon gamma, respectively. CONCLUSIONS: It is concluded that the presence of bile is required for the expression of MHC class II molecules on gut epithelium and that the cytokine components of bile may be the inducing agents.     

A dominant-negative clathrin mutant differentially affects trafficking of molecules with distinct sorting motifs in the class II major histocompatibility complex (MHC) pathway

The role of clathrin in intracellular sorting was investigated by expression of a dominant-negative mutant form of clathrin, termed the hub fragment. Hub inhibition of clathrin-mediated membrane transport was established by demonstrating a block of transferrin internalization and an alteration in the intracellular distribution of the cation-independent mannose-6-phosphate receptor. Hubs had no effect on uptake of FITC-dextran, adaptor distribution, organelle integrity in the secretory pathway, or cell surface expression of constitutively secreted molecules. Hub expression blocked lysosomal delivery of chimeric molecules containing either the tyrosine-based sorting signal of H2M or the dileucine-based sorting signal of CD3gamma, confirming a role for clathrin-coated vesicles (CCVs) in recognizing these signals and sorting them to the endocytic pathway. Hub expression was then used to probe the role of CCVs in targeting native molecules bearing these sorting signals in the context of HLA-DM and the invariant chain (I chain) complexed to HLA-DR. The distribution of these molecules was differentially affected. Accumulation of hubs before expression of the DM dimer blocked DM export from the TGN, whereas hubs had no effect on direct targeting of the DR-I chain complex from the TGN to the endocytic pathway. However, concurrent expression of hubs, such that hubs were building to inhibitory concentrations during DM or DR-I chain expression, caused cell surface accumulation of both complexes. These observations suggest that both DM and DR-I chain are directly transported to the endocytic pathway from the TGN, DM in CCVs, and DR-I chain independent of CCVs. Subsequently, both complexes can appear at the cell surface from where they are both internalized by CCVs. Differential packaging in CCVs in the TGN, mediated by tyrosine- and dileucine-based sorting signals, could be a mechanism for functional segregation of DM from DR-I chain until their intended rendezvous in late endocytic compartments.     

Structural requirements for major histocompatibility complex class II invariant chain endocytosis and lysosomal targeting

The invariant chain (Ii) targets newly synthesized major histocompatibility complex class II complexes to a lysosome-like compartment. Previously, we demonstrated that both the cytoplasmic tail (CT) and transmembrane (TM) domains of Ii were sufficient for this targeting and that the CT contains two di-leucine signals, 3DQRDLI8 and 12EQLPML17 (Odorizzi, C. G., Trowbridge, I. S., Xue, L., Hopkins, C. R., Davis, C. D., and Collawn, J. F. (1994) J. Cell Biol. 126, 317-330). In the present study, we examined the relationship between signals required for endocytosis and those required for lysosomal targeting by analyzing Ii-transferrin receptor chimeras in quantitative transport assays. Analysis of the Ii CT signals indicates that although 3DQRDLI8 is necessary and sufficient for endocytosis, either di-leucine signal is sufficient for lysosomal targeting. Deletions between the two signals reduced endocytosis without affecting lysosomal targeting. Transplantation of the DQRDLI sequence in place of the EQLPML signal produced a chimera that trafficked normally, suggesting that this di-leucine sequence coded for an independent structural motif. Structure-function analysis of the Ii TM region showed that when Ii TM residues 11-19 and 20-29 were individually substituted for the corresponding regions in the wild-type transferrin receptor, lysosomal targeting was dramatically enhanced, whereas endocytosis remained unchanged. Our results therefore demonstrate that the structural requirements for Ii endocytosis and lysosomal targeting are different.     

Minimum structural requirements for peptide presentation by major histocompatibility complex class II molecules: implications in induction of autoimmunity

The precise mechanisms of failure of immunological tolerance to self proteins are not known. Major histocompatibility complex (MHC) susceptibility alleles, the target peptides, and T cells with anti-self reactivity must be present to cause autoimmune diseases. Experimental autoimmune encephalomyelitis (EAE) is a murine model of a human autoimmune disease, multiple sclerosis. In EAE, residues 1-11 of myelin basic protein (MBP) are the dominant disease-inducing determinants in PL/J and (PL/J x SJL/J)F1 mice. Here we report that a six-residue peptide (five of them native) of MBP can induce EAE. Using peptide analogues of the MBP-(1-11) peptide, we demonstrate that only four native MBP residues are required to stimulate MBP-specific T cells. Therefore, this study demonstrates lower minimum structural requirements for effective antigen presentation by MHC class II molecules. Many viral and bacterial proteins share short runs of amino acid similarity with host self proteins, a phenomenon known as molecular mimicry. Since a six-residue peptide can sensitize MBP-specific T cells to cause EAE, these results define a minimum sequence identity for molecular mimicry in autoimmunity.     

Assembly of an abundant endogenous major histocompatibility complex class II/peptide complex in class II compartments

To identify the intracellular site(s) of formation of an endogenous class II/peptide complex in a human B cell line, we employed kinetic pulse-chase labeling experiments followed by subcellular fractionation by Percoll density gradient centrifugation and immunogold labeling on ultrathin cryosections. For direct demonstration of assembly of such complexes, we used the monoclonal antibody YAe, which detects an endogenous complex of the mouse class II molecule I-Ab with a 17-amino acid peptide derived from the alpha chain of HLA-DR (DR alpha52-68). We show that in human B lymphocytes, these class II/peptide complexes assemble and transiently accumulate in major histocompatibility complex class II-enriched compartments before reaching the cell surface.     

Expression of major histocompatibility complex class II antigens in the canine intestine

Immunohistochemical techniques were used to assess major histocompatibility complex (MHC) class II expression by enterocytes and lamina propria cells in the canine intestinal tract. Duodenal enterocyte class II expression was faint and limited to the lower crypt region whereas jejunal and ileal enterocyte expression was stronger, being present in both crypt and villus areas. Enterocyte staining was of greatest intensity in crypts adjacent to Peyer's patches and intense membrane staining of most Peyer's patch lymphocytes was also seen. Enterocyte MHC class II expression in the colon was largely limited to the lower crypt region. Within the lamina propria, of all intestinal sites examined, a heterogeneous population of cells were MHC class II positive and these had morphological features of macrophages and dendritic cells. Lymphocytes, plasma cells, fibroblasts and vascular endothelium were not stained. Definition of constitutive expression of MHC class II within the canine intestine may be important in identifying upregulation of this molecule in inflammatory bowel diseases.     

Two distinct gamma interferon-inducible promoters of the major histocompatibility complex class II transactivator gene are differentially regulated by STAT1, interferon regulatory factor 1, and transforming growth factor beta

The speed of the myofilament lattice spacing response to rapid changes in load or length of single, intact muscle fibres of the frog, was investigated during isometric tetani. During ramp releases at close to Vmax and during step length changes (completed within 250 microseconds), lattice spacing was calculated from the equatorial X-ray diffraction pattern (sampled at 250 microseconds time resolution using synchrotron radiation). Ramp releases (total shortening=1.39 %) caused a spacing increase, described with an exponential function (alpha=271 s-1, amplitude=1.15 nm) plus an elastic component having the time course of discharge of axial tension (amplitude 0.28 nm). For a step release (amplitude=0.87%), lattice expansion could be described with an exponential (alpha =1005 s-1, amplitude=0.56 nm) plus an elastic component of 0.25 nm amplitude. Lattice compression was associated with a step stretch (amplitude=0.62 %), and was also quasi-exponential (alpha=367 s-1, amplitude=0.74 nm), with an elastic component of 0.28 nm. The spacing change time course for length steps resembled that of the accompanying quick recovery of axial tension and the associated change in the meridional 14.5 nm reflection intensity, which are both believed to be determined by the kinetics of the molecular power stroke. Therefore, this shows that lattice spacing changes, arising from radial forces exerted by attached crossbridges, are fast enough to occur during the power stroke event.     

Major histocompatibility complex class II molecules, hemopoiesis and the marrow microenvironment

Currently available data indicate that the earliest identifiable hemopoietic progenitor in normal marrow is CD34+ MHC class II-; subsequent expression of MHC class II antigens is maturation and lineage dependent. Studies on embryonal cells suggest that CD34+DR- cells are actually the common precursors for stromal and hemopoietic elements, with the earliest hemopoietic precursor being CD34+DR+. DQ antigens are apparently not expressed in cells of hemopoietic potential and the expression of DQ appears to be regulated differentially from DR and DP. MHC class II antigens are also expressed on some stromal cells, especially those with endothelial and macrophage features. MHC class II molecules are involved in hemopoietic cell/stroma interaction. The presence of anti-MHC class II monoclonal antibodies (MABs) at early stages of stem cell proliferation/differentiation, at least under conditions of marrow stress, induces signals which may result in final, especially granulocytic, differentiation of later precursors. These may interfere with the survival of those cells which are required for long-term hemopoietic reconstitution. Observations in allogeneic marrow transplant recipients support a role of MHC molecules as expected in allogeneic interactions. Results in autologous models point towards a role of MHC class II molecules other than that of a histocompatibility marker insofar as these molecules or signals transmitted by them appear to be involved in the regulation of hemopoiesis.     

Molecular docking of superantigens with class II major histocompatibility complex proteins

The molecular recognition of two superantigens with class II major histocompatibility complex molecules was simulated by using protein-protein docking. Superantigens studied were staphylococcal enterotoxin B (SEB) and toxic shock syndrome toxin-1 (TSST-1) in their crystallographic assemblies with HLA-DR1. Rigid-body docking was performed sampling configurational space of the interfacial surfaces by employing a strategy of partitioning the contact regions on HLA-DR1 into separate molecular recognition units. Scoring of docked conformations was based on an electrostatic continuum model evaluated with the finite-difference Poisson-Boltzmann method. Estimates of nonpolar contributions were derived from the buried molecular surface areas. We found for both superantigens that docking the HLA-DR1 surface complementary with the SEB and TSST-1 contact regions containing a homologous hydrophobic surface loop provided sufficient recognition for the reconstitution of native-like conformers exhibiting the highest-scoring free energies. For the SEB complex, the calculations were successful in reproducing the total association free energy. A comparison of the free-energy determinants of the conserved hydrophobic contact residue indicates functional similarity between the two proteins for this interface. Though both superantigens share a common global association mode, differences in binding topology distinguish the conformational specificities underlying recognition.     

Engagement of the B lymphocyte antigen receptor induces presentation of intrinsic immunoglobulin peptides on major histocompatibility complex class II molecules

By means of the clonotypic variable region, the immunoglobulin (Ig) is a tumor-specific antigen on B cell neoplasms. We report that engagement of the B cell antigen receptor (BcR) promotes presentation of peptides derived from the B cell's intrinsic Ig to major histocompatibility complex (MHC) class II-restricted T cells. Thus, anti-Ig endowed normal, ex vivo B lymphocytes from H-2d, Ig constant heavy chain allotype b (IgCHb) mice with the capacity to stimulate an I-Ad-restricted T cell clone which recognizes the gamma 2ab 435-451 allopeptide. The corresponding self gamma 2aa peptide is cryptic and 6000-fold less antigenic than the gamma 2ab allopeptide. Even so, the syngeneic B cell lymphoma A20 which expresses surface(s) IgG2aa, was also recognized by the T cells after BcR ligation. Thus, anti-Ig triggered the disclosure of a cryptic tumor antigen determinant. We propose that autoantigens, by engaging the BcR of self-reactive B cells, induce presentation of intrinsic Ig peptides to which the T helper cell (Th) repertoire is not tolerant. In this way, B cells with anti-self potential may be activated without Th recognition of nominal autoantigen.     

Peptide epitopes from noncytosolic Listeria monocytogenes can be presented by major histocompatibility complex class I molecules

Listeria monocytogenes is an intracellular pathogen which escapes the phagosome and resides in the cytosol of the host cell. Using Listeria innocua and a mutant strain of L. monocytogenes (listeriolysin O negative), which do not enter the cytosol of the host cell, we demonstrate class I presentation of an epitope of p60, a protein secreted by L. monocytogenes, to a class I-restricted CD8+ cytotoxic T lymphocyte clone.     

Anti-adhesive signals are mediated via major histocompatibility complex class II molecules in normal and neoplastic human B cells: correlation with B cell differentiation stage

We show that major histocompatibility complex (MHC) class II molecules on B cells transit signals which regulate adhesion in a negative manner. Engagement of MHC class II molecules with antibodies results in detachment of B cells previously bound to interferon-gamma-activated human umbilical cord venous endothelial cells. This process depends on metabolic energy, active signaling and protein tyrosine kinase activity. The adhesion pathway influenced by this signaling event is neuraminidase sensitive. The anti-adhesive signaling program is activated in B cell lines with a mature phenotype, e.g. normal B cells from spleen and tonsil. In contrast, cell lines with a pre-B cell phenotype and normal B cells from peripheral blood are refractory to MHC class II-mediated regulation of adhesion. These results extend to neoplastic cells from patients with lymphoproliferative diseases representing different stages of B cell maturation. These results suggest that MHC class II-mediated signals regulate B cell adhesion in a developmentally programmed fashion; this might have implications for clinical behavior of B cell malignancies.     

Helicobacter pylori infection in immunized mice lacking major histocompatibility complex class I and class II functions

The role of major histocompatibility complex (MHC) class I- and class II-restricted functions in Helicobacter pylori infection and immunity upon oral immunization was examined in vivo. Experimental challenge with H. pylori SS1 resulted in significantly greater (P 相似文献   

Interferon-gamma differentially regulates antigen-processing functions in distinct endocytic compartments of macrophages with constitutive expression of class II major histocompatibility complex molecules

Treatment of pregnant female Sprague-Dawley rats on Gestational Day 15 with a single oral dose of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) (0.5, 1.0, or 2.0 micrograms/kg) or indole-3-carbinol (I3C, 1.0 or 100 mg/kg), an aryl hydrocarbon (Ah) receptor agonist which is found in cruciferous vegetables, resulted in reproductive abnormalities in the male offspring (three to five litters in each treatment group). Anogenital distance and crown to rump length were altered by both compounds; however, the timing of the effects (Day 1 or 5) was variable and the responses were not necessarily dose-dependent. In 62-day-old offspring, seminal vesicle (24 to 26%), prostate (32 to 44%), testicular parenchymal (14%), and epididymal weight (19%) were decreased by one or more doses of TCDD. In contrast, I3C at one or more doses decreased daily sperm production/g testicular parenchyma (13 to 20%) and daily sperm production/testis (22%). Total number of sperum in the epididymis was significantly decreased (30 to 33%) in rats perinatally exposed to TCDD and this was due to a decreased (49 to 51%) number of sperm in the tail of the epididymis. Perinatal exposure to I3C did not affect any of these parameters. TCDD did not affect epididymal transit time of sperm through the complete epididymis at any of the doses (0.5 to 2.0 micrograms/kg). However, at the two highest doses (1.0 and 2.0 micrograms/kg), TCDD increased epididymal transit rate of sperm through the tail of the epididymis by 33 and 37%, respectively. In contrast, primarily due to decreased transit rate (39%) of sperm through the head plus body of the epididymis. I3C (1 mg/kg) significantly increased total epididymal transit time by 31%. In conclusion, perinatal exposure of pregnant rats to I3C, an Ah receptor agonist similar to TCDD, causes reproductive abnormalities in male rat offspring; however, I3C and TCDD elicited both common and different responses.