L-plastin: Structure, Regulation, and Roles in Cancer Invasion and in Macrophages

The cytoskeleton consists of 3 filamentous components: intermediate filaments, microtubules, and actin filaments. Actin filaments continuously assemble and disassemble far out of equilibrium to adapt cells in response to external stimuli. Actin filaments organization and dynamic are controlled by a multitude of actin-binding proteins including actin-bundling proteins. L-plastin, expressed abundantly in lymphocytes and monocytes, is an actin-bundling protein that roles in immune defense and in metastatic invasion of cancer cells. The actin-bundling activity of L-plastin is regulated not only by intracellular calcium concentration, but by phosphorylation of Ser5. The actin-bundling activity of L-pastin decreases by increased calcium concentration but is promoted by phosphorylation of Ser5. The morphology changes and motility of cells requires continuous remodeling of actin filaments which demands the sensitive nature of L-plastin to Ca2+-signal, phosphorylation of Ser5, and probably additional regulation. This review briefly describes the structure and regulation of L-plastin, and roles for L-plastin in cancer invasion and in macrophages.

Enzymatic characterization of Mycobacterium smegmatis ADP-ribosyltransferase

ADP-ribosyltransferase (ADPRT) catalyzes the reaction in which the ADP-ribose moiety of beta-NAD+ is transferred to specific amino acid residues in target proteins. The ADPRT of Mycobacterium smegmatis has been known to inactivate rifampin through ADP-ribosylation. However, the enzymatic characteristics and functions of the enzyme have not been elucidated yet. In this study, the ADPRT-glutathione S-transferase (GST) fusion protein was expressed in Escherichia coli and enzymatic characteristics of the fusion protein were investigated. ADPRT-GST fusion protein was an ADPribosyltransferase that had no NAD glycohydrolase activity. ADPRT-GST fusion protein showed no self-inactivation phenomenon that is a universal nature for all NAD glycohydrolases and is important in regulating its activity. ADPRT activity of the enzyme was decreased by novobiocin and isonicotinic acid hydrazide. These results suggest that Mycobacterium smegmatis ADPRT could be regulated by a different way from other NADases and involved in bacterial physiological process through a post-translational modification of cytosolic proteins.

Involvement of the CXC chemokines Mig and IP-10 in response to M. bovis BCG in mice

The non-ELR-containing CXC chemokines Mig and IP-10 have been shown to function as chemotactic cytokines for activated T lymphocytes. In this study, we examined the potential involvement of Mig and IP-10 in antimycobacterial response of mice immunized or infected with M. bovis BCG. The accumulation of Mig and IP-10 mRNA in resident peritoneal monocytes (RPMPHI) was slightly reduced by stimulation with vBCG, and the degree was greater for 24 hr culture even though IFN-gamma was added. Expression of Mig, IP-10, and IFN-gamma in 24 hr delayed-type hypersensitivity (DTH) response was stronger in vBCG-immune mice than in the non-immune. The increase of DTH measured by foot-pad thickness appears to be clearly related to the levels of chemokines Mig and IP10 messages and those of IFN-gamma and IL-12. Stimulation with vBCG for 2 days decreased or completely dropped the levels of Mig message in non-immune or immune splenocytes, respectively, whereas IP-10 message was slightly decreased in 2 days culture. Moreover, messages for IL-12 (p40) showed similar kinetics for Mig. The levels of Mig and IP-10 mRNA during the course of infection with BCG were not readily changed in lungs, livers, and spleens from BCG-infected mice. Although there was no obvious changes of Mig and IP-10 messages in the target organs during infection process, we found that the infection progressed over the first 3 wk before being contained by the emerging immune response suggested from detectable amount of IFN-gamma mRNA around this time. In view of selectivity of chemokines Mig and IP-10 for activated T cells, these data suggest that chemokine Mig and IP-10, especially in collaboration with IL-12 and IFN-gamma, may play a role as T cell recruiters in immune response against mycobacterial infection.

IFN -r Enhances Induction of Chemokines Mig and IP10 mRNA from THP - 1 Cells Stimulated with Lipoarabinomannan / 대한면역학회지

Lipoarabinomannans (LAM) is believed as a potential virulence factor of Mycobacterium tuberculosis. LAM exhibits marked differences in biological activities depending on the types, arabinofuranosyl-terminated LAM (AraLAM) derived from a rapidly growing Mycobacterium sp. and heavily mannosylated LAM (ManLAM) derived from the Erdman strain. Collaboration between macrophages and T cells, especially macrophage activation by gamma interferon (IFN-r) and chemoattraction of T cells at the very inflammatory foci would be essential in defence against M. tubercu/osis. Chemokines Mig and IP-10 are inducible by IFN-r from macrophages and have been shown to act in vitro as T cell chemoattractants. However, little is known of LAMs capacity to induce chemokines Mig and IP-10 in macrophages. In this experiment, Mig and IP10 mRNA was expressed in the delayed-type hypersensitivity (DTH) against BCG in BCG-immune mice. In some experiments, both Mig and IP-10 mRNA was evidently induced with different time courses in THP-1 cells stimulated with whole live M. tubercu/osis H37Rv (Erdman). To investigate whether Mig and IP-10 genes are differentially induced depending on the type of LAM, PCR amplification was used to detect mRNA of Mig and IP-10 from the THP-1 human monocytic cells stimulated with LAM. AraLAM, but not ManLAM, induced weakly Mig and IP-10 mRNA in the THP-1 cells. The induction of Mig and IP-10 was dependent upon the dose of AraLAM and exhibited different time courses. The mRNA for Mig and IP-10 was induced within 2 hr and 4 hr from the initiation of treatrnent and has disappeared by 8 hr and 24 hr under the experimental conditions used in this study, respectively. IFN-y at 100 U/ml, but not at 10 U/ml, was itself a good stimulus of both Mig and IP- 10 expression, and synergized with either AraLAM or ManLAM for induction of both Mig and IP-10. The expression patterns of MCP-3 were somewhat similar to those of Mig and IP10 in all of the experiments. These data indicate that IFN-r may contribute to effective macrophage function if macrophages are not fully affected by ManLAM, and chemokines Mig and IP-10 may a role in recruitment of T cells at inflammatory foci of tuberculosis.

Genomic Organization of ht eGene for Human Mig Chemokine / 대한면역학회지

"Mig is a gamma interferon-inducible T cell chemoattractant that is a member of the chemokine family of cytokines. In order to gain a better understanding of the molecular mechanisms that regulate expression of the Mig gene, we have characterized the Mig gene and compared its structure and regulatory sequences with that of its ciosest IP10 gene. The genomic organization of the Mig gene reveals three introns that interrupt the transcribed sequence into four functional domains with a single ""CAT""- and ""TATA""-like structure. Primer extension analysis was used to identify the transcriptional initiation site that is located 50 bp upstream to the methionine codon that begins the long open reading frame. Comparison of the intron-exon structure of this gene to the gene for IP10 establishes that both genes are interrupted in precisely the same positions within homologous codons. The similarity of the intron-exon structure of the Mig and IP10 genes further support the hypothesis that Mig and IP10 genes have evolved from a common ancestral gene by gene duplication. The 5'-flanking region of Mig gene shows no overall sequence similarity with that from its closest IP10 gene whose production is also affected by gamma interferon. However, there are regions including a sequence with similarity to the NFxB binding site, AP-1 binding site, and ISRE. The r-RF-1 binding site is well conserved from -204 to -194 from the transcription start site in the Mig gene. Given the importance of IFN-r for effective immunity in tuberculosis and induction of Mig and IP10 genes in macrophages by IFN-r, we demonstrated induction of the genes Mig and IP10 with different message levels in the THP-1 human monocytic cell lines stimulated with whole M. tuberculosis. Despite the very similarity in genomic organization and the overlap in biological activities between MIG and IP10, our data described herein further support the suggestion that these chemokines rnay role nonredundantly in vivo. Moreover, our studies done on the Mig gene should provide the structural framework for future studies and begin to dissect cis-acting DNA sequences that are critical for gene regulation mediated by cell surface receptors."

Anti-IL-4 antibody inhibits antigen specific IgE response but fails to prevent chicken gamma globulin-induced active systemic anaphylaxis: evidence for the involvement of IgG antibodies

It has recently been reported that interleukin-4 (IL-4) is required for the production of IgE, and anti-IL-4 monoclonal antibody (mAb) inhibits in vivo IgE responses. These suggest that blocking of IL-4 activity may be useful for the prevention or treatment of immediate hypersensitivity disorders. In this study we investigated whether anti-IL-4 has a regulatory role in chicken-gamma globulin (CGG)-induced active systemic anaphylaxis. Multiple injections of anti-IL-4 (up to 40 mg/mouse) failed to protect the mice from fatal anaphylaxis. Anti-IL-4 strongly suppressed CGG-specific IgE response (>90%) without any suppressive effect on CGG-specific IgG (IgG1, IgG2a, IgG2b, and IgG3) responses. Because these data suggest the possibility that fatal anaphylaxis could be induced by IgG antibodies, we examined the possibility using anti-CGG polyclonal and the subclasses of IgG monoclonal antibodies. Passive sensitization of mice with polyclonal antibodies elicited severe and fatal anaphylactic shock; about 50% of the mice died. The activity of antibodies was not diminished by heat treatment (56 degrees C, 2h), suggesting that the anaphylaxis was not mediated by IgE. Shock was also elicited by each subclass of IgG mAb; of these, IgG1 was the most effective. Combination of the IgG subclasses elicited more exaggerated shock; about 30% of mice died. These data indicate that IgG antibodies are themselves sufficient to induce systemic anaphylaxis. Therefore, the failure of anti-IL-4 to prevent active anaphylaxis is probably due to the inability of anti-IL-4 to suppress the production of IgG antibodies.

HgCl2 Dysregulates the Immune Response of Balb/c Mice / 예방의학회지

The studies reported here were undertaken to investigate the effects of mercury chloride on immune system of Balb/c mouse employing a flexible tier of in vitro and in vivo assays. Mercury chloride inhibited the proliferative responses of spleen cells to lipopolysaccharide, pokeweed mitogen, and phytohemagglutinin as a dose-dependent manner. This inhibitory effect was observed not only when HgCl2 was added 2nd or 3rd day of 3 days culture period but also when spleen cells was pretreated with HgC12 for 2 hours. Mercury chloride, however, potentiated the production of IgM and IgG from spleen cells. During the HgCl2 administration by drinking for 3 weeks, the weight gain of mice was significantly blunted than that of control group mice, while no overt signs related to mercury toxicity were noted in any mice of experimental group. There was no change in thymus and spleen weights, and in histological findings of kidney, bone marrow of femur, thymus, spleen, and politeful lymph node after 3 weeks of mercury exposure. However, HgC12 induced a significant increase of total serum IgM, IgG including IgG1, IgG2a and IgG2b, and IgE in Balb/c mice. Treatment in vivo with anti-IL-4 monoclonal antibody significantly abrogated the HgCl2-induced increase in total serum IgG1 and IgE. Whereas HgCl2 potentiated total serum IgM and IgG, there was, there was no difference in total serum hemagglutinin to SRBC(Sheep Red Blood cell) between experimental and control group mice when these mice were immunized with SRBC. All these findings observed in Balb/c mice suggest that mercury perturbates well-orchestrated regulation of immune responses before developing histopathological changes in lymphoid tissues.

Effects of monoclonal anti-IL-4 antbody(11B11) and interferons onIgE production in vivo and hypersensitivity reactions I. inductionof systemic anaphylaxis in mice by antigen-specific IgG and IgG subclasses monoclonal antibodies / 대한면역학회지

No abstract available.

Production and partial charaterization of monoclonal antibodiesreactive with human class II antigens / 대한면역학회지

No abstract available.