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J H Weis

Publications and source records attributed to J H Weis.

At least 37 records · Page 2Linked to original sources
PubMed1997

Expression of the murine CD21 gene is regulated by promoter and intronic sequences.

Murine CD21 gene products are expressed primarily on the surface of B lymphocytes and follicular dendritic cells. To identify the genetic elements that control the tissue-specific expression of the CD21 gene, we analyzed, via transient transfections, the 5' proximal promoter region of the CD21 gene (1272 bp 5' of the initiating ATG). This region possessed strong promoter activity, but it was not tissue specific, in that T cell expression was equivalent to that of B cells. These data suggested that the anticipated tissue-specific control element(s) lies 3' of the initiating ATG. Analysis of a novel minigene construct that possessed both the 5' promoter region and a large region (9 kb) of the CD21 gene 3' of the initiating ATG demonstrated the expected tissue-specific expression. Further analysis using the luciferase reporter system indicated that such control elements reside in the first intron (5.5 kb in size), which separates the exons encoding the signal sequence and the first extracellular short consensus repeat domain of the mature protein. Further dissection of intron 1 demonstrated that the sequences controlling the tissue-specific expression of the murine CD21 gene are contained in the 5' 1.6-kb region of this intron. This 1.6-kb fragment was fractionated into an 800-bp sequence at the 5' end that showed very significant inhibitory activity in both B and T cells and a 3' 800-bp sequence that demonstrated moderate repression in T cells, but enhancer activity in B cells. These data suggest this region of the CD21 gene possesses a number of functionally distinct sites that positively and negatively regulate CD21 gene transcription.

Animals↗
PubMed1997

Borrelia burgdorferi outer surface protein A (OspA) activates and primes human neutrophils.

Lyme disease is caused by infection with the spirochete Borrelia burgdorferi and is characterized by bacterial persistence and inflammation of many host tissues. B. burgdorferi express outer surface lipoproteins, including OspA, with inflammatory properties that could contribute to the localized tissue inflammation. Neutrophils are the predominant infiltrate into the inflamed arthritic joints, and are crucial for controlling the spirochete infection. They may also contribute to the joint pathology associated with Lyme arthritis. This study examines the effect of OspA on the activities of the neutrophil. Picomolar concentrations of OspA induce surface markers associated with neutrophil activation: increased CD10 and CD11b expression; decreased CD62-L expression; and an increased adherence to extracellular matrix. These events were similar in kinetics and magnitude to those induced by the strong activators LPS and FMLP. Like LPS, OspA could prime neutrophils for FMLP-induced release of lysosomal granules and production of superoxide. Thus, models of Lyme arthritis should include the possible contribution of direct activation of neutrophils to both defense and disease.

Antigens, Surface↗
PubMed1996

Modulation of expression of the anti-inflammatory cytokines interleukin-13 and interleukin-10 by interleukin-3.

Interleukin (IL)-4, IL-10 and IL-13 are cytokines with potent anti-inflammatory activities. Prevention of pathological inflammation at mucosal surfaces appears to be due, in part, to the presence of these cytokines. One potential source for these cytokines is the mast cell which resides at mucosal surfaces. Demonstrated in this report are the findings that bone marrow-derived mucosal-like mast cells constitutively expressed IL-13 whereas bone marrow-derived connective tissue-like mast cells demonstrated IL-13 transcription only after Fc epsilon RI-mediated activation or the addition of exogenous IL-3. A similar pattern of expression of IL-10 by these mast cell types was also evident and matches that of IL-4 previously reported. Intracellular cytokine staining indicated that IL-10 protein is constitutively expressed by the bone marrow-derived mucosal-like mast cells but is only evident in the bone marrow-derived connective tissue-like mast cells after induction with IL-3. The increase of IL-13 and IL-10 transcripts in the connective tissue-like mast cells following IL-3 treatment is not mast cell specific, in that splenic and bone marrow cells also demonstrated the same phenomenon. These data suggest that mucosal mast cells may have a constitutive repertoire of Th2 cytokines with potential anti-inflammatory activity, while connective tissue mast cells may not. However, production of such cytokines can be induced in the connective tissue mast cell and other cell types of the immune response by the addition of IL-3.

Animals↗
PubMed1996

Mucosal T cells and mast cells share common adhesion receptors.

Mast cells and T cells share common homing/adhesion receptors, which suggests that they have evolved to use similar trafficking pathways. Furthermore, these cells have been shown to co-localize in the gastrointestinal mucosa, and it is possible that they act in concert to achieve an immune response. Here, Tracey Smith and John Weis discuss the implications of this potential collaboration.

Animals↗
PubMed1994

Preferential expression of interleukin-12 or interleukin-4 by murine bone marrow mast cells derived in mast cell growth factor or interleukin-3.

In this report we demonstrate that murine bone marrow cells cultured in either interleukin (IL)-3 or mast cell growth factor (MGF, also known as c-kit ligand and stem cell factor) differentially express cytokine genes. Bone marrow cells cultured in IL-3 differentiate and proliferate, taking on a mucosal mast cell-like phenotype. These cells express the IL-4 gene. Bone marrow cells cultured in MGF take on a connective tissue mast cell-like phenotype and possess transcripts for both of the subunits of the IL-12 cytokine. Bone marrow cells cultured in both IL-3 and MGF express the IL-4 gene at lower levels than that seen for the IL-3 culture alone, but do not possess IL-12 gene transcripts. The level of IL-12 subunit transcripts derived from the MGF-derived bone marrow cells was compared to that found in splenocytes and activated macrophages, the only cells in which IL-12 production has been previously documented. Both of the IL-12 subunit transcripts were found, compared to a beta-actin control, to be present within MGF-derived cells in the same if not higher quantities than the splenocyte or macrophage cultures. Mucosal mast cells have been previously implicated in the development of the T helper type 2 (TH2) T cell phenotype via their expression of IL-4. The finding that the MGF-derived connective tissue-like mast cells possess IL-12 transcripts suggests that the development of the TH1 T cell pathway may be positively influenced by this type of mast cell.

Animals↗
PubMed1994

Murine M290 integrin expression modulated by mast cell activation.

Intraepithelial T lymphocytes possess a (TGF beta)-inducible integrin complex that consists of the beta 7 integrin chain in association with a novel alpha chain, murine alpha M290. Using antibodies directed against the murine alpha M290 chain, IL-3-dependent murine bone marrow-derived mast cells demonstrated positive staining after induction with TGF beta. Sequence information from the human intraepithelial T lymphocyte-alpha E gene sequence was used to clone the murine homolog from a TGF beta-induced T cell library. The murine alpha E homolog gene encodes a protein recognized by antibodies to alpha M290. Expression of this gene by bone marrow-derived mast cells was found to be induced by TGF beta or IgE-mediated cross-linking of Fc epsilon R1. Furthermore, exposure of murine TK-1 cells to supernatants from activated mast cells induces the expression of the alpha M290 gene, indicating that a mast cell mediator(s) can act in a paracrine manner to induce expression of this integrin complex by other cells.

Amino Acid Sequence↗
PubMed1994

Heritable susceptibility to severe Borrelia burgdorferi-induced arthritis is dominant and is associated with persistence of large numbers of spirochetes in tissues.

In human Lyme disease, symptoms with widely varying levels of severity have been observed. A mouse model of Lyme disease has been developed which allows analysis of mice with mild, moderate, and severe pathologies after inoculation with the spirochete Borrelia burgdorferi. To determine whether the differences in symptoms reflect differences in the number of spirochetes persisting in affected tissues, a sensitive PCR technique was developed to detect B. burgdorferi DNA in virtually any tissue of an infected mouse. This analysis, which detects DNA from as few as three spirochetes, revealed the presence of B. burgdorferi DNA in many tissues from severely arthritic C3H/HeJ mice as early as 1 week postinfection. The heart, ear, and ankle were particularly heavily infected, although B. burgdorferi DNA was also detected in spleen, liver, brain, kidney, bladder, uterus, and lymph nodes. In contrast, much lower levels of spirochete DNA were detected in tissues of infected BALB/c mice, which develop less severe arthritis when infected with B. burgdorferi than do C3H/HeJ mice. This difference was evident throughout the 5-week analysis. A competitive PCR method allowed determination of the absolute number of spirochete gene sequences in infected tissues. Ankles and hearts from C3H/HeJ mice were found to harbor 10(7) copies of the B. burgdorferi ospA gene, while these tissues from BALB/c mice contained 5- and 10-fold less B. burgdorferi DNA, respectively. The genetic regulation of severe pathology was analyzed by infecting the offspring of a cross between C3H/HeJ and BALB/c mice. The F1 mice developed severe arthritis and contained high levels of Borrelia DNA in the heart and ankle, similar to the C3H/HeJ parent. These findings indicate that susceptibility to severe arthritis is a dominant trait and suggest that it may correlate with high levels of persisting spirochetes. Models of pathology in Lyme disease should take into consideration the fact that severity of pathology may be directly related to the number of organisms in infected tissues.

Animals↗
PubMed1993

Enhanced expression of the complement regulatory protein, membrane inhibitor of reactive lysis (CD59), is regulated at the level of transcription.

The membrane inhibitor of reactive lysis (MIRL) is an 18-Kd glycosyl phosphatidylinositol anchored membrane glycoprotein that inhibits the cytolytic activity of complement. MIRL is expressed by all hematopoietic elements and by a wide variety of nonhematopoietic tissues. A deficiency of MIRL is primarily responsible for the greater sensitivity of the erythrocytes of paroxysmal nocturnal hemoglobinuria to complement mediated lysis. Because of its critical role in protecting host cells from injury by complement, we hypothesized that mechanisms exist that allow MIRL expression to be regulated. To investigate this hypothesis, both MIRL RNA and MIRL protein expression were analyzed following exposure of K562 erythroleukemia cells to a variety of potential stimulants. Incubation with dexamethasone, calcium ionophore, lipopolysaccharide, interleukin 1, tumor necrosis factor, hemin, and cyclic AMP had no effect on MIRL expression. However, incubation with phorbol 12-myristate 13 acetate (PMA), induced a marked increase in MIRL RNA as determined by Northern blot analysis. This enhanced expression of MIRL RNA was associated with an increase in MIRL protein expression as determined by immunoprecipitation of metabolically labeled proteins, Western blot analysis, and immunobinding assay. Enhanced MIRL RNA expression was first detected after 8 hours and increased through 24 hours of observation. Inhibitors of either protein synthesis or transcription abrogated the PMA-induced enhancement of MIRL RNA expression. Together, these results are consistent with a model in which PMA induces synthesis of a trans acting protein that enhances transcription of the MIRL gene.

Antigens, CD↗
PubMed1993

Functional identification of transcription control sequences of the mouse Crry gene.

The mouse C receptor-related gene Crry is expressed by a wide variety of cells. Those sequences required for the transcriptional control of this gene were identified by deletion analysis of regions 5' of the initiating ATG. Fusion of Crry promoter sequences to the reporter gene CAT identified a region approximately 1,500 bp upstream of the transcriptional start site that enhanced transcription of this gene construct. Gel shift and methylation interference assays were performed, and a specific protein-DNA complex was identified within this region. These assays defined a 16-bp sequence 1,642 bp 5' of the initiating ATG that bound a protein in nuclear extracts prepared from all murine cell lines and tissues examined. The methylation interference assay indicated that the core region of the DNA sequence recognized by the protein was GGAA, the common core binding site for the ets family of proto-oncogenes. Oligonucleotides prepared from this sequence with the GGAA sequence did inhibit the DNA/protein complex formation, whereas those with a mutant GGAA sites did not. The minimal site identified by methylation interference was able to up-regulate transcription when placed downstream of a heterologous promoter, whereas the same sequence with an altered GGAA site could not. Thus, this site functions as an enhancer.

Animals↗
PubMed1993

Murine macrophages lack expression of the Cr2-145 (CR2) and Cr2-190 (CR1) gene products.

Murine macrophages have been described as possessing complement receptors for C3b and iC3b. These binding activities have been assumed to be due to the presence of the CR1 and CR3 proteins, respectively. The mouse Cr2 gene produces two distinct gene products of approximately 190,000 relative molecular mass (M(r)) (Cr2-190) and 145,000 M(r) (Cr2-145). Because of the similarity in size to human complement receptors, the Cr2-190 protein has been dubbed murine CR1 while the murine Cr2-145 product has been termed murine CR2. In order to define the complement receptor genes expressed by murine macrophages, we investigated the expression patterns of Cr2-190, Cr2-145 and another mouse complement receptor, Crry, in three different mouse macrophage populations: bone marrow-derived macrophages, thioglycollate-elicited peritoneal macrophages and the macrophage cell line, J774. Neither of the Cr2 gene transcripts encoding the Cr2-145 and Cr2-190 proteins could be detected in these populations by RT-RPCR analysis although Crry transcripts were evident. Cr2-145 and Cr2s-190 proteins could not be detected on the surface of thioglycollate-elicited macrophages using a monoclonal antibody that recognizes both proteins. Thus, contrary to previously published data, murine macrophages do not possess the Cr2 gene products.

Animals↗
PubMed1993

Murine complement receptor gene expression: Cr2 gene transcripts are depressed during a high dose microbial challenge.

The murine Cr2 gene encodes mRNA that produce two protein products predicted to be approximately 145,000 M(r) (Cr2-145) and 190,000 M(r) (Cr2-190). All cells examined which express the Cr2 gene produce transcripts encoding both the Cr2-145 and Cr2-190 proteins: both transcripts are constitutively expressed by mature B cells. To determine if Cr2 expression could be altered by activating splenic B cells, splenic cultures were incubated with lipopolysaccharide (LPS) and cell surface Ig chains were cross-linked with anti-mu. In the presence of LPS and anti-mu both Cr2 and Oct-2 transcripts were diminished while the control beta-actin transcript levels remained unchanged. However, when LPS alone was added, only the Cr2 transcript levels were diminished. To test if these findings could be reproduced in vivo, animals were provided with a peritoneal injection of either Escherichia coli or Listeria monocytogenes and transcript levels analysed. The quantities of both Cr2 transcripts, as well as those encoding Oct-2, were substantially reduced in splenocytes and peripheral lymphatic tissues obtained from these infected mice while those encoding the mouse Crry protein, the B-cell marker CD19 and beta-actin remained unchanged. These data suggest that when confronted with a major bacterial infection, murine B cells respond by shutting down synthesis of transcripts encoding the Cr2 and Oct-2 gene products.

Animals↗
PubMed1992

Expression of murine beta 7, alpha 4, and beta 1 integrin genes by rodent mast cells.

The screening of a rat mast cell cDNA library with a probe selected to recognize those genes preferentially or exclusively expressed by mast cells identified a rat gene sequence, RF-17, that shared homology with the beta-integrins. This integrin was expressed in rat tissues enriched for mast cells and T cells. The rat RF-17 sequence was used to isolate the murine homologue from a spleen cDNA library. The murine gene encodes a protein of 806 amino acids that is the probable homologue to the human beta 7 chain. Transcripts specific for the murine gene are found in the thymus, spleen, and lung. To attempt to identify the gene product for this new integrin chain, we examined the murine T cell line TK-1, which expresses a novel integrin heterodimer, lymphocyte Peyer's patch high endothelial venule adhesion molecule (LPAM-1), of a known alpha 4 chain and an unknown beta P chain, for expression of murine beta 7 (RF-17). This cell line expresses high levels of RF-17 transcripts, suggesting that beta P is encoded by the beta 7 gene. Bone marrow cells induced to differentiate into mast cells via IL-3 express the beta 7 gene as well as the genes encoding the murine integrin alpha 4 and beta 1 proteins. Surface staining analysis indicates that these cells express an alpha 4-containing integrin complex throughout the differentiation process. These data suggest that the Peyer's patch homing LPAM-1 receptor expressed by a subset of T cells consists of the beta 7 gene product and the alpha 4 chain, and that this integrin chain complex is also found on the surface of maturing mast cells. The presence of beta 1 transcripts also suggests that these maturing mast cells possess the LPAM-2 integrin complex (alpha 4/beta 1) as well. The experimental strategy described in this manuscript has, thus, identified a novel murine beta-integrin chain that is expressed by rodent T cells and mast cells.

Amino Acid Sequence↗
PubMed1992

Identification of sites for distinct DNA binding proteins including Oct-1 and Oct-2 in the Cr2 gene.

The murine Cr2 gene produces two distinct products in a variety of murine cell types. Both of these transcripts appear to initiate from the same position within the gene but vary from one another via an alternative splicing event within the coding exons. An analysis of those gene sequences that might control the cell specific expression of the Cr2 gene has identified a region of Cr2 5' of the transcription start site that is conserved in both the murine Cr2 and human CR2 genes. When this region was examined using the gel shift assay with nuclear extracts from cells expressing Cr2 (B cells) and those that do not (T cells and fibroblasts), at least four distinct proteins were identified that bound to at least three distinct sites. The DNA sequence recognized by two of these proteins is the octamer sequence recognized by a family of transcriptional regulators including the B cell specific Oct-2 protein. During an acute bacterial infection, the levels of Oct-2 and Cr2 mRNA are both depressed. This suggests that the Oct-2 protein directly controls the transcriptional activity of the Cr2 gene and that during such an infection, the levels of Ag receptors on B cells (Ig and complement receptors) are diminished.

Animals↗
PubMed1992

Modulation of integrin expression during mast cell differentiation.

Previously we have reported that rodent mast cells synthesize the mRNA encoding the alpha and beta integrin chains (alpha 4, beta 1 and beta 7) of the lymphocyte Peyer's patch adhesion molecule (LPAM)-1 and LPAM-2 lymphocyte homing receptors, and that they possess an alpha 4-containing integrin complex on their cell surface. In this report, we have examined the expression of these integrin chain genes by mature connective tissue mast cells (CTMC) and by bone marrow-derived mast cells (BMMC) differentiated from bone marrow precursor cells in the presence of interleukin (IL)-3 and/or the c-kit ligand (also known as mast cell growth factor and stem cell growth factor). High levels of both the beta 7 and beta 1 transcripts were present in mature CTMC while those encoding the alpha 4 chain were absent. Similarly, when BMMC were grown in IL-3 for 28 days and analyzed for integrin chain transcripts, those specific for the alpha 4 chain were also diminished compared to beta 7 and beta 1 transcripts. To compare the expression of these integrin genes during mucosal mast cell and CTMC development, BMMC were derived in the presence of IL-3 alone, c-kit alone, or IL-3/c-kit together. These experiments indicated that c-kit inhibited the transcription of the beta 7 and Fc epsilon RI genes while enhancing alpha 4 transcript levels. The enhancement of alpha 4 levels, however, was abrogated with the addition of IL-3. Similarly, the c-kit-induced depression of beta 7 and Fc epsilon RI transcript levels was overcome by the addition of IL-3. These data suggest that the integrin complexes synthesized by the mast cells may differ depending upon their path of differentiation and that another alpha chain integrin may be synthesized to complex with the beta 7 and/or beta 1 chains.

Animals↗
PubMed1992

Development of a sensitive reverse transcriptase PCR assay, RT-RPCR, utilizing rapid cycle times.

We describe a procedure to analyze rare gene transcripts quantitatively using a reverse transcriptase-polymerase chain (RT-PCR) reaction. RNA purified from cells and tissues is reverse-transcribed using random hexamer primers and is amplified using very short cycle times. The products are trace-labeled with [32P[dCTP, which allows for the quantitation of the products by gel electrophoresis and excision of bands. The quantity of the PCR product is directly proportional to the quantity of cDNA added and is reproducible from a single cDNA preparation or from samples derived from separate preparations. Because cDNA synthesis is primed with random oligonucleotides, the same cDNA sample preparation can be examined for many different gene products.

Animals↗
PubMed1990

Loss of human CR1- and murine Crry-like exons in human CR2 transcripts due to CR2 gene mutations.

Analysis of the coding sequences of the murine Cr2 gene indicated that it contains two distinct regions of homology to the murine Crry, and human CR1 and CR2 genes. The last 15 short consensus repeats (SCR) of Cr2 are very similar to the 15 reported SCR of human CR2. These 15 SCR of Cr2, plus the transmembrane and cytoplasmic domains, make up a protein very similar in organization and sequence to the human CR2 gene product. Another Cr2 transcript contains the sequences encoding the previously described 15 SCR plus those encoding another six amino terminal SCR. These amino terminal SCR are very similar to those of Crry and the first six SCR seen within the CR1 long homologous repeats. Amino acid sequence similarity analysis, however, indicated that the sequences encoding the six amino terminal SCR of Cr2 evolved from a separate lineage of SCR than Crry and CR1, suggesting that the human counterparts to these additional Cr2 SCR had not been identified. Using the cDNA sequences specific for the amino terminal SCR of murine Cr2, the human counterparts were isolated and localized within the human CR2 gene between those nucleotides encoding the signal sequence and those encoding the first SCR of the mature human CR2 protein. Unlike the murine Cr2 gene products, these Crry/CR1-like sequences of the CR2 gene are not maintained in mature CR2 mRNA, and thus represent pseudoexons. DNA sequence analysis of the pseudoexon homologous to that encoding the first SCR of the murine Cr2 gene indicated that a stop codon has been introduced within the human coding sequence but that the 5' and 3' splice recognition sequences appear to be functional. Although the stop codon would block the translation of a transcript with this exon, it would not inhibit the incorporation of the exon within the mature CR2 transcript. Therefore, another mutation must have been introduced within the human CR2 gene such that both this exon and the other CR1/Crry-like exons are removed from mature CR2 transcripts.

Amino Acid Sequence↗