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PubMed1992

T-cell receptor-gamma delta association with lymphocyte populations in sheep intestinal mucosa.

T cells expressing T-cell receptor (TcR)-gamma delta and CD8 represent a significant population in mouse and chicken intra-epithelial lymphocytes (IEL) but represent a minor population in human IEL. We examined the TcR-gamma delta usage and co-expression of CD5, CD4, CD8 and major histocompatibility complex (MHC) class II on isolated sheep IEL and lamina propria lymphocytes (LPL), and compared them with the TcR-gamma delta + cells in peripheral blood, intestinal lymph and jejunal Peyer's patches (PP). There were a number of notable differences. TcR-gamma delta + cells comprised 18% of IEL and 10% of LPL. Among the population of TcR-gamma delta + IEL, 24% were CD8+ and 54% were CD5+, which contrasts with the TcR-gamma delta + cells in blood and intestinal lymph that were universally CD5+ CD4- CD8-. A notable feature of the IEL was the presence of distinct CD8+ and TcR-gamma delta + populations that lacked CD5. Also a high percentage of IEL and LPL were CD2+ and MHC class II+. Analysis of the expression of MHC class II on T-cell subsets, as an indicator of activation, showed that 60-95% of the various IEL and LPL subsets were MHC class II+ compared with only 5-40% in jejunal PP, lymph nodes, spleen and blood. Therefore, it is possible that the circulating TcR-gamma delta + and CD8+ cells that localize in the gut epithelium might become activated and stop the expression of CD5 under the influence of the local microenvironment. These cells appear not to emigrate while still expressing the TcR-gamma delta + (CD8+) CD5- MHC class II+ phenotype. Our data, together with those from other studies, show that there is much heterogeneity in the use of TcR-gamma delta and accessory T-cell molecules by IEL.

Animals↗
PubMed1992

Differential regulatory effects of intercellular adhesion molecule-1 on costimulation by the CD28 counter-receptor B7.

Although ligation of the CD3/TCR complex initiates an activation signal in T cells, additional costimulatory signals generated during cell-to-cell interactions with APC transduced via ligation of CD11a/CD18 and CD28 by their specific counter-receptor intercellular adhesion molecule (ICAM)-1 and B7, respectively, are required for optimal T cell proliferation and cytokine synthesis. Using soluble IgC gamma 1 fusion proteins of these costimulatory counter-receptors, we have recently shown that unactivated resting CD4+ T cells and Ag-primed CD4+ T cells differ in their response to the costimulation by ICAM-1 and B7. Preferential proliferative responses of resting T and Ag-primed T cells to ICAM-1 and B7, respectively, prompted us to speculate that ICAM-1-induced signals may regulate coupling of the CD28 signaling pathway. Furthermore, both B7 and ICAM-1 are co-expressed on APC and thus, may co-regulate activation-driven maturation of T cells. In this study, we have examined regulatory effects of IgC gamma 1 fusion proteins of B7, ICAM-1, and ICAM-2 (a homologue of ICAM-1) on each other's costimulation. We first demonstrate that TCR-directed costimulation of resting CD4+ T cells with ICAM-1 (ICAM-1 priming) but not ICAM-2 induces increased responsiveness to B7. Priming of CD4+ T cells with ICAM-1 induced higher expression of both CD18 and CD28 than that with either B7 or ICAM-2. Cross-linking of CD28 induced faster and significantly higher cytoplasmic free calcium mobilization response in ICAM-1-primed CD4+ T cells than in resting, B7-primed, or ICAM-2-primed CD4+ T cells. B7 synergized with ICAM-1 but not ICAM-2 to augment proliferative responses of not only resting CD4+ T cells but also those that had been primed with either ICAM. Unlike resting or ICAM-2-primed CD4+ T cells, ICAM-1-primed CD4+ T cells efficiently proliferated in response to the synergistic costimulation of B7 and ICAM-2. In contrast, both ICAM-1 and ICAM-2 inhibit B7-driven proliferation of Ag-primed CD4+ T cells. Thus, B7 and ICAM-1 exert contrasting regulatory effects on the proliferation of CD4+ T cells depending on their state of activation-induced maturation.

Antigens, CD↗
PubMed1992

Patterns of membrane CD45 isoform expression by leukaemic blasts and normal mature myeloid cells.

The membrane expression of CD45R isoforms by leukaemic blasts from 92 cases of acute myeloid (AML) and 12 cases of lymphoblastic leukaemia was analysed by single and two-colour flow cytometry. Compared to normal mature granulocytes, which invariably expressed UCHL1 (CD45RO) but not 2H4 (CD45RA), leukaemic myeloid blasts showed 2H4+ UCHL1- and 2H4- ++UCHL1- composite patterns of expression with the first of these phenotypes being associated in particular with the most immature AML subtype (M0). Similar analyses of blast cell fractions from monocytic AML variants, revealed wide variation in CD45R expression in cases of M4, whereas M5 leukaemias were typically 2H4+ ++UCHL1+ with minor 2H4- UCHL1- components. As most normal mature monocytes were also 2H4+ ++UCHL1+, this suggested that monocytic myeloid differentiation was primarily associated with the "acquisition" of UCHL1 and the development of 2H4/UCHL1 co-expressing cells. The expression of membrane CD45RA by leukaemic blasts is an abnormal characteristic and may be related to the maintenance of an undifferentiated state by malignant myeloid precursors.

Adult↗
PubMed1992

Pattern of IL-1 receptor gene expression suggests role in noninflammatory processes.

The cytokine IL-1 can be elaborated by most nucleated cell types and exerts its pleiotropic effects on a variety of tissues through binding to its cognate receptor. Two forms of IL-1R, designated types I and II, have been identified. These proteins share limited amino acid identity but both bind IL-1 alpha and IL-1 beta. A large number of cell types have been shown to possess IL-1R in vitro, but few studies have addressed the question of in vivo expression. Using in situ hybridization in normal adult BALB/c mice, we have examined the tissue distributions of both IL-1R types. The results demonstrate that although lymphoid tissues in healthy animals express low or nondetectable levels of receptor transcript, nonlymphoid tissues constitutively express readily detectable levels of IL-1R mRNA. Thymus and spleen samples were largely negative for both IL-1R types I and II, whereas half of the lymph nodes examined expressed IL-1R type I. In nonlymphoid tissues, IL-1R type I transcript was detected in the dentate gyrus of the brain, endocrine pancreas, cardiac endothelium, epidermis, dermis, hair follicle epithelium, uterine serosa, vaginal stroma, vaginal squamous epithelium, developing oocytes, and granulosa cells of ruptured follicles. In contrast, the IL-1R type II probe hybridized to epidermis, dermis, and vaginal basal epithelium. The two types of IL-1R were rarely, if ever, co-expressed by the same cell. The distributional segregation of the receptor types and the expression of IL-1R in normal nonlymphoid tissues has broad implications for the role of IL-1 in homeostatic physiology.

Animals↗
PubMed1992

CDC43 and RAM2 encode the polypeptide subunits of a yeast type I protein geranylgeranyltransferase.

The question regarding the identity of the alpha and beta subunits of the yeast type I protein geranylgeranyltransferase was explored using prokaryotic expression of candidate genes. The Saccharomyces cerevisiae CDC43 and RAM2 genes were expressed in Escherichia coli and cell extracts examined for the ability to transfer [3H]geranylgeranyl diphosphate to an appropriate CaaX protein substrate. Individual expression of each gene yielded no activity; however, co-expression of the two genes resulted in high levels of [3H] geranylgeranyl incorporation into the substrate protein Ras-Cys-Val-Val-Leu. The activity was partially purified yielding approximately 12,600 units/liter. The partially purified enzyme geranylgeranylated the Ras-Cys-Val-Val-Leu, Ras-Cys-Ala-Ile-Leu, Ras-Cys-Ile-Ile-Leu, and Ras-Cys-Thr-Ile-Leu substrates but not the Ras-Cys-Val-Leu-Ser or Ras-Ser-Val-Leu-Ser substrates. The protein geranylgeranyltransferase was highly specific for geranylgeranyl diphosphate and poorly transferred farnesyl. The recombinant enzyme was indistinguishable from the native type I geranylgeranyltransferase in yeast extracts. As has been reported for the protein farnesyltransferase, the yeast type I protein geranylgeranyltransferase is also a magnesium-requiring, zinc metalloenzyme. Interestingly, the recombinant enzyme functioned with calcium as the only divalent cation, although addition of zinc increased calcium-dependent activity 2-fold.

Alkyl and Aryl Transferases↗
PubMed1992

Identification of B beta chain domains involved in human fibrinogen assembly.

Fibrinogen chains are assembled in a stepwise manner in the rough endoplasmic reticulum prior to secretion of the final six-chain dimeric molecule. Previous studies indicated that the synthesis of B beta may be a rate-limiting factor in the assembly of human fibrinogen. To determine the domains of B beta which interact with the other two component chains of fibrinogen, deletion mutants of B beta were transiently co-expressed, together with A alpha and gamma chains, in COS cells, and fibrinogen assembly and secretion were measured. Deletion of the COOH-terminal half of the B beta chain (amino acids 208-461) did not affect assembly and secretion. Assembly of A alpha, gamma, and B beta also occurred when the first NH2-terminal 72 amino acids of B beta were deleted, but not when 93 amino acids were deleted. This indicates that the B beta domain between amino acids 73 and 93 is necessary for the assembly of the three fibrinogen chains. This domain marks the start of the alpha-helical "coiled-coil" region of fibrinogen.

Amino Acid Sequence↗
PubMed1992

Alternative exon usage and processing of the major histocompatibility complex-encoded proteasome subunits.

The finding that two subunits of the proteasome, LMP2 and LMP7, are encoded in the major histocompatibility complex (MHC) has linked the proteasome which represents a major extralysosomal proteolytic system to the processing of intracellular antigens. Here we describe a second form of the human LMP7 cDNA, LMP7-E2, which has been identified during the characterization of novel genes in the MHC. The analysis of the genome organization of LMP7 revealed that LMP7-E1 and LMP7-E2 arise by alternative exon usage. Using specific antibodies against LMP2 and LMP7, we show that they are co-expressed with class I MHC molecules as well as a putative peptide transporter. The polypeptides encoded by LMP7 and LMP2 undergo proteolytic processing when incorporated into proteasomes, and the LMP7 precursor is derived mainly from LMP7-E2. Furthermore, our data suggest that LMP7 and LMP2 are mutually dependent for their incorporation into the proteasomal complex.

Amino Acid Sequence↗
PubMed1992

Studies of the catalytic activities and substrate specificities of Saccharomyces cerevisiae myristoyl-coenzyme A: protein N-myristoyltransferase deletion mutants and human/yeast Nmt chimeras in Escherichia coli and S. cerevisiae.

Saccharomyces cerevisiae myristoyl-CoA:protein N-myristoyltransferase (Nmt1p) is an essential, 455-residue, monomeric enzyme. Amino- and carboxyl-terminal deletion mutants of Nmt1p were genetically engineered to determine the minimal domain necessary to maintain catalytic activity. Enzyme activity was assessed by (i) sequentially inducing Nmt1p or its mutant derivatives and one of two eukaryotic substrates for the wild type enzyme (S. cerevisiae Gpa1p and rat Go alpha) in Escherichia coli, a bacterium with no endogenous myristoyltransferase activity, and monitoring Nmt-dependent incorporation of exogenous [3H]myristate into the G protein alpha subunits or (ii) an in vitro enzyme assay using lysates prepared from bacteria producing wild type or mutant Nmts. The data indicate that the minimal catalytic domain of Nmt1p is located between Ile59-->Phe96 and Gly451-->Leu455. Analyses of the ability of mutant nmtps to rescue the lethal phenotype of an nmt1 null allele in a haploid strain of yeast grown on rich media, with or without blockade of cellular fatty acid synthetase, suggest that the amino-terminal 59 residues of Nmt1p may play an important noncatalytic role, functioning as a targeting signal so this cytosolic enzyme can access cellular myristoyl-CoA pools generated from activation of exogenous C14:0 by acyl-CoA synthetase(s). Moreover, there appear to be differences in the location or accessibility of myristoyl-CoA pools derived from fatty acid synthetase and acyl-CoA synthetases. The E. coli co-expression system was used to map structural elements that determine differences in the peptide substrate specificities of Nmt1p and the orthologous human Nmt. Rat Go alpha is a substrate for both enzymes, whereas human Gz alpha is a substrate only for human NMT. Studies of a series of chimeric enzymes composed of elements from the amino- or carboxyl-terminal portions of human and yeast Nmts indicate that (i) recognition/utilization of Gz alpha involves elements distributed from the amino-terminal half through the region defined by Leu352-->Lys410 of the 416 residue human enzyme and (ii) formation of a fully functional peptide binding site and a fully functional myristoyl-CoA binding site in either of these enzymes requires contributions from both their amino-terminal and carboxyl-terminal halves.

Acyltransferases↗
PubMed1992

Unidirectional, heterologous desensitization of the pertussis toxin receptor by the CD3/TCR complex.

Prolonged exposure of many types of receptors to their cognate agonists can lead to a progressive lack of responsiveness. When this occurs after stimulation by the primary agonist for a given receptor it is termed homologous desensitization, and heterologous desensitization when to an agonist binding to a different type of receptor. Pertussis toxin (PTx) is a potent mitogen for human T lymphocytes. We have previously identified the human T cell PTx receptor (PTx-R) as a 43-kDa plasma membrane protein that, when stimulated, leads to the production of the intracellular second messengers, inositol-1,4,5-triphosphate, 1,2-sn-diacylglycerol, and elevated cytosolic calcium. The PTx-R appears to require the co-expression of the CD3/TCR complex because mutant cells that lack the AgR, but express the PTx-R, fail to respond to PTx. In this report, we have investigated the relationship between these two receptor systems. Activation of the PTx-R with submaximal concentrations of PTx did not affect the ability of an anti-CD3 antibody combined with rabbit anti-mIg to stimulate increases in intracellular free calcium concentration [Ca2+]i or diacylglycerol in human peripheral blood T cells. However, treatment with soluble anti-CD3 mAb, which lead to only a modest increase in [Ca2+]i, completely inhibited the effect of PTx. The cells were not refractory to further stimulation of the AgR because cross-linking with rabbit anti-mIg resulted in the standard maximal stimulation. This effect could be observed within 1 min of treatment with anti-CD3 mAb, and persisted for at least 1 h. The effect was not caused by production of either diacylglycerol (leading to activation of PK-C) or an increase in [Ca2+]i by anti-CD3 mAb because the effect could not be mimicked by either phorbol esters or a calcium ionophore. Pretreatment of either resting T lymphoblasts or PBL with anti-CD3 mAb also prevented enhanced [3H]TdR incorporation stimulated by PTx. These observations suggest a model in which T cells can regulate amplification of a non-AgR stimulatory pathway by heterologous desensitization.

Calcium↗
PubMed1992

Induction of Fc epsilon RII/CD23 on PHA-activated human peripheral blood T lymphocytes and association of fyn tyrosine kinase with Fc epsilon RII/CD23.

Despite the evidence for the expression of Fc epsilon RII/CD23, a glycoprotein that is a low-affinity Fc receptor for IgE, obtained on T cell lines and some pathological T cells, that of Fc epsilon RII/CD23 on normal human T cells is still unclear. We studied the emergence of T cells bearing Fc epsilon RII/CD23 in short-term culture of normal human peripheral blood mononuclear cells stimulated with 15 microliters/ml phytohemagglutinin (PHA). Using two-dimension flow cytometry, more than 10% of Fc epsilon RII/CD23(+) cells were shown to co-express CD3 antigen. Both CD4(+) and CD8(+) T cells expressed Fc epsilon RII/CD23. The expression of mRNA for Fc epsilon RII/CD23 on PHA and IL-4 stimulated PBMC was demonstrated by northern blotting and in-situ hybridization. The mechanism of signal transduction through Fc epsilon RII/CD23 was dissected by transfection of cDNA coding for Fc epsilon RII to the human natural killer-like cell line YT, activation of which was easily detected by the induction of interleukin-2 receptor/p55 (Tac). Cross-linking of Fc epsilon RII/CD23 with H107 anti-Fc epsilon RII monoclonal antibody enhanced IL-2R/p55 expression on YT cells transfected with Fc epsilon RII cDNA (YTSER). A possible involvement of protein-tyrosine kinase in the Fc epsilon RII-mediated signal transduction was studied using YTSER. Fc epsilon RII was physically associated with an src-family tyrosine kinase p59fyn and not with p56lck, which was also found in YT cells. Recently it was reported that p59fyn was associated with T-cell antigen receptor. Our results collectively suggest the multiple function of p59fyn which may be implicated in the Fc epsilon RII-mediated activation signal in YT cells.

Blotting, Northern↗
PubMed1992

Purification and ligand binding of a soluble class I major histocompatibility complex molecule consisting of the first three domains of H-2Kd fused to beta 2-microglobulin expressed in the baculovirus-insect cell system.

A recombinant baculovirus encoding a single-chain murine major histocompatibility complex class I molecule in which the first three domains of H-2Kd are fused to beta 2-microglobulin (beta 2-m) via a 15-amino acid linker has been isolated and used to infect lepidopteran cells. A soluble, 391-amino acid single-chain H-2Kd (SC-Kd) molecule of 48 kDa was synthesized and glycosylated in insect cells and could be purified in the absence of detergents by affinity chromatography using the anti-H-2Kd monoclonal antibody SF1.1.1.1. We tested the ability of SC-Kd to bind antigenic peptides using a direct binding assay based on photoaffinity labeling. The photoreactive derivative was prepared from the H-2Kd-restricted Plasmodium berghei circumsporozoite protein (P.b. CS) peptide 253-260 (YIPSAEKI), a probe that we had previously shown to be unable to bind to the H-2Kd heavy chain in infected cells in the absence of co-expressed beta 2-microglobulin. SC-Kd expressed in insect cells did not require additional mouse beta 2-m to bind the photoprobe, indicating that the covalently attached beta 2-m could substitute for the free molecule. Similarly, binding of the P.b. CS photoaffinity probe to the purified SC-Kd molecule was unaffected by the addition of exogenous beta 2-m. This is in contrast to H-2KdQ10, a soluble H-2Kd molecule in which beta 2-m is noncovalently bound to the soluble heavy chain, whose ability to bind the photoaffinity probe is greatly enhanced in the presence of an excess of exogenous beta 2-m. The binding of the probe to SC-Kd was allele-specific, since labeling was selectively inhibited only by antigenic peptides known to be presented by the H-2Kd molecule.

Adenovirus E1A Proteins↗
PubMed1992

Biphenotypic hybrid acute leukaemia detected by two colour flow cytometry.

Immunophenotypic studies using immunofluorescent flow cytometry were performed on the blast cells of 36 patients with acute leukaemia using a panel of eight monoclonal antibodies. Six patients had blasts which co-expressed markers for lymphoid and myeloid differentiation, and which were therefore defined as biphenotypic hybrid acute leukaemia. Of the six, three patients were in the paediatric age group (below 12 years old) while the other three were more than 12 years old. Peripheral blood counts were variable; however, bone marrow infiltration was extensive (blasts > or = 75% in all). At the time of study, remission was achieved in only two patients. The authors' data show that biphenotypic hybrid acute leukaemia is not rare in Malaysia. This represents a subgroup of acute leukaemia identifiable by immunophenotyping but not by the French-American-British classification based on morphological and basic cytochemical studies alone. The recognition of this subgroup is important for both practical and theoretical reasons. There are implications for treatment of the individual patient because treatment directed at a single lineage may not be effective. The two colour flow cytometry proved to be a useful tool for diagnosis and classification of acute leukaemia.

Adolescent↗
PubMed1992

Class II major histocompatibility complex (Ia) antigen-bearing dendritic cells within the iris and ciliary body of the rat eye: distribution, phenotype and relation to retinal microglia.

The density, distribution and surface phenotype of dendritic cells (DC) and macrophage populations within the ciliary body and iris of Wistar Furth rats were studied by a combination of flat mounting, tangential sectioning, pre-embedding fixation, with a single and double immunohistochemical techniques. Monoclonal antibodies included anti-Ia (OX6) and other dendritic cell/macrophage (ED1 and ED8) or mature tissue macrophage markers (ED2). Single and double staining revealed a network (approximately 400 cells/mm2) of Ia+ cells within the epithelium of the ciliary processes with the morphological and surface phenotypic characteristics of DC populations in other tissues. A minor proportion of DC co-expressed ED1 and ED8, but not ED2. In contrast the immunopositive cells in the lamina propria displayed a more generalized phenotype, including ED2 expression, and pleiomorphic morphology suggesting a preponderance of cells of macrophage lineage. Flat mounts of iris revealed a remarkably regular network of Ia+ DC at a density of 450 cells/mm2. The network of DC in the ciliary epithelium terminated at the cilioretinal junction where they formed a continuous syncytium with retinal microglia which did not display Ia staining. The demonstration of networks of cells with relevant morphological and phenotypical properties of professional antigen-presenting cells at strategic locations within the eye has important implications in relation to ocular immune regulation and on the theories of the mechanism of anterior chamber-associated immune deviation (ACAID). Namely, until now it has been assumed that 'immune privilege' in the anterior chamber of the eye is partly a consequence of there being a paucity of class II+ cells in the surrounding tissues. Dendritic cells in the eye may function as antigen-presenting cells, sampling endogenous and exogenous intraocular antigens and possibly migrating from the eye to draining lymphoid organs (the spleen) where they may generate systemic immune responses. Equally dendritic cells could potentially regulate local immune responses for example in various forms of autoimmune uveoretinal inflammatory disease.

Animals↗
PubMed1992

Phenotypic modulation of perisinusoidal cells following acute liver injury: a quantitative analysis.

Expression of the alpha-(smooth muscle) isoform of actin (alpha-SMA) by non-parenchymal cells in rat liver was studied following induction of acute liver injury using a single sublethal dose of carbon tetrachloride (CCl4). In normal liver, alpha-SMA immunoreactivity was identified in the smooth muscle cells of hepatic arteries and in the walls of portal and hepatic vein branches. Occasional alpha-SMA-containing stellate shaped cells were found in acinar zone 3 but most perisinusoidal cells (PSCs) did not express this protein. In CCl4-treated animals, there was an increase in the number of immunoreactive cells in perivenular zones, reaching a peak at day 3 following exposure to the toxin. These cells were morphologically identical to desmin-positive PSCs and the kinetics of the responses of alpha-SMA-positive and desmin-positive cells were similar. In en face labelling experiments, evidence of co-expression of alpha-SMA and desmin by non-parenchymal cells was obtained, although some desmin-positive PSCs did not appear to express alpha-SMA. These results suggest that PSCs rapidly undergo phenotypic modulation in response to acute liver injury with acquisition of alpha-SMA expression. It is proposed that these phenotypic changes coincide with functional alterations, such activated 'myofibroblast-like' cells being responsible for the enhanced matrix protein synthesis necessary for tissue repair.

Actins↗
PubMed1992

The capsular polysaccharide of Bacteroides fragilis comprises two ionically linked polysaccharides.

Recently, we have shown that the capsular polysaccharide of Bacteroides fragilis NCTC 9343 is composed of an aggregate of two discrete large molecular weight polysaccharides (designated polysaccharides A and B). Following disaggregation of this capsular complex by very mild acid treatment, high resolution NMR spectroscopy demonstrated that polysaccharides A and B consist of highly charged repeating unit structures with unusual substituent groups (Baumann, H., Tzianabos, A. O., Brisson, J.-R., Kasper, D.L., and Jennings, H.J. (1992) Biochemistry 31, 4081-4089). Presently, we report that the capsular polysaccharide of B. fragilis represents a complex structure that is formed as a result of ionic interactions between polysaccharides A and B. Electron microscopy of immunogold-labeled organisms (with monoclonal antibodies specific for polysaccharides A and B) demonstrated that the two polysaccharides are co-expressed on the cell surface of B. fragilis. We have shown that the purified capsule complex is made up exclusively of polysaccharide A and polysaccharide B (no other macromolecular structure was detected) in a 1:3.3 ratio and that disaggregation of this complex into the native forms of the constituent polysaccharides could be accomplished by preparative isoelectric focusing. Structural analyses of the native polysaccharides A and B showed that they possessed the same repeating unit structures as the respective acid-derived polysaccharides. The ionic nature of the linkage between polysaccharides A and B was demonstrated by reassociation of the native polysaccharides to form an aggregated polymer comparable to the original complex. The distinctive composition of this macromolecule may provide a rationale for the unusual biologic properties associated with the B. fragilis capsular polysaccharide.

Bacteroides fragilis↗
PubMed1992

A conditional secretory mutant in an Ig L chain is caused by replacement of tyrosine/phenylalanine 87 with histidine.

The kappa-chain of the myeloma MOPC 21 is an unusual L chain, in that it is not secreted unless complexed with a H chain. This nonsecreted kappa-chain seems to be retained in the endoplasmic reticulum in association with the protein BiP/GRP78, both in myeloma cells and when expressed in COS-1 fibroblasts. By assaying the fate of the MOPC 21 kappa-chain and its mutated derivatives in COS-1 cells, we show that the cause of the nonsecreted phenotype is the presence of histidine in position 87 of the variable domain. When this amino acid is changed back to the tyrosine that usually occupies position 87, secretion of the unassembled kappa-chain is restored. As in B lymphoid cells, co-expression of gamma H chains in COS-1 cells complements the mutation in the L chain, and rescues secretion of the arrested kappa. Thus, the presence of histidine at position 87 creates a conditional L chain secretory mutant: it is not compatible with normal transport of free L chain, but can be rescued in the presence of H chain.

Base Sequence↗
PubMed1992

Coexpression of type I and type II human macrophage scavenger receptors in macrophages of various organs and foam cells in atherosclerotic lesions.

Macrophage scavenger receptors are trimeric membrane glycoproteins implicated in the pathologic deposition of cholesterol in arterial walls during atherogenesis. Two types of cDNAs for functional human receptors have been cloned, but their physiologic roles remain obscure. To study the expression of these receptors, the authors generated antibodies against scavenger receptor type-specific synthetic peptide. Immunohistochemical examination using these antibodies and other anti-human receptor antibodies shows that type I and type II receptor proteins can be detected in foam cells in various stages of atherosclerosis, most evidently in fatty streaks. Co-expression of the two types of receptor protein was also detected in macrophages of various organs. Both types of the protein were detected on the surface and the membrane of endosomes in macrophages. These results indicate that both type I and type II scavenger receptors are expressed and functionally active in physiologic and pathologic conditions.

Arteriosclerosis↗
PubMed1992

Recombinant pollen allergens from Dactylis glomerata: preliminary evidence that human IgE cross-reactivity between Dac g II and Lol p I/II is increased following grass pollen immunotherapy.

We previously described the isolation of three identical complementary DNA (cDNA) clones, constructed from Orchard/Cocksfoot grass (Dactylis glomerata) anther messenger RNA (mRNA), expressing a 140,000 MW beta-galactosidase fusion protein recognized by IgE antibodies in atopic sera. Partial nucleotide sequencing and inferred amino acid sequence showed greater than 90% homology with the group II allergen from Lolium perenne (Lol II) indicating they encode the group II equivalent, Dac g II. Western blot immunoprobing of recombinant lysates with rabbit polyclonal, mouse monoclonal and human polyclonal antisera demonstrates immunological identity between recombinant Dac g II, Lol p I and Lol p II. Similar cross-identity is observed with pollen extracts from three other grass species: Festuca rubra, Phleum pratense and Anthoxanthum odoratum. Recombinant Dac g II was recognized by species- and group-cross-reactive human IgE antibodies in 33% (4/12) of sera randomly selected from grass-sensitive individuals and in 67% (14/21) of sera from patients receiving grass pollen immunotherapy, whilst 0/4 sera from patients receiving venom immunotherapy alone contained Dac g II cross-reactive IgE. Cross-reactive IgG4 antibodies were detectable in 95% of sera from grass pollen immunotherapy patients. These preliminary data suggest that conventional grass pollen allergoid desensitization immunotherapy may induce IgE responses to a cross-reactive epitope(s) co-expressed by grass pollen groups I and II (and possibly group III) allergens.

Allergens↗