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- Andersson, Kerstin, et al.
(författare)
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Yersinia pseudotuberculosis-induced calcium signaling in neutrophils is blocked by the virulence effector YopH
- 1999
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Ingår i: Infection and Immunity. - 0019-9567 .- 1098-5522. ; 67:5, s. 2567-2574
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Tidskriftsartikel (refereegranskat)abstract
- Pathogenic species of the genus Yersinia evade the bactericidal functions of phagocytes. This evasion is mediated through their virulence effectors, Yops, which act within target cells. In this study we investigated the effect of Yersinia pseudotuberculosis on Ca 2+ signaling in polymorphonuclear neutrophils. The intracellular free calcium concentration in single adherent human neutrophils was monitored during bacterial infection and, in parallel, the encounter between the bacteria and cells was observed. When a plasmid-cured strain was used for infection, adherence of a single bacterium to the cellular surface induced a β 1 integrin-dependent transient increase in the intracellular concentration of free calcium. This was, however, not seen with Yop-expressing wild-type bacteria, which adhered to the cell surface without generating any Ca 2+ signal. Importantly, the overall Ca 2+ homeostasis was not affected by the wild-type strain; the Ca 2+ signal mediated by the G-protein-coupled formyl-methionyl-leucyl- phenylalanine receptor was still functioning. Hence, the blocking effect was restricted to certain receptors and their signaling pathways. The use of different Yop mutant strains revealed that the protein tyrosine phosphatase YopH was responsible for the inhibition. This virulence determinant has previously been implicated in very rapid Yersinia-mediated effects on target cells as the key effector in the blockage of phagocytic uptake. The present finding, that Y. pseudotuberculosis, via YopH, specifically inhibits a self- induced immediate-early Ca 2+ signal in neutrophils, offers more-detailed information concerning the effectiveness of this virulence effector and implies an effect on Ca 2+-dependent, downstream signals.
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| 4. |
- Majeed, Meytham, 1960-
(författare)
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Regulation of Attachment and Early Intracellular Development of Chlamydia trachomatis in Eucaryotic Cells
- 1994
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- The obligate intracellular bacterium, Chlamydia trachomatis , a common human pathogen, causing different diseases in both males and females, as well as in infants born to infected mothers. Occasionally, these diseases involve serious complications, such as blindness and infertility. C. trachomatis has a unique biphasic life cycle: after initial inclosure in membrane~bound endosomes, the infectious elementary bodies (EBs) reorganize to the metabolically actiVe forms (RBs); the RBs divide by binary fission, and after multiple divisions, they again differentiate to form new EBs, which are subsequently released from the host cell to start a new infectious cycle. As an attempt to investigate the early events of chlamydial infection, my results show that EBs bind with high affinity to collagen type I and heparan sulfate, suggesting that this selective affinity may mediate the attachment of chlamydiae to the surface of a host cell. ER-containing endosomes avoid fusion with host celllysosomes. However, within 30 min of form ation, these endosomes form one local aggregate in the central orperinuclear region of individual cells. This aggregation is reversible and time and temperature dependent, and requires viable EBs. Clathrin and F-actin are mobilized and colocalized with EB aggregates, suggesting that the aggregation of EBs is an active process that may be biologically involved in the infectivity of chlamydiae. The aggregation and inclusion formation of EBs, and the redistribution of F-actin seem to be controlled by both extra- and intracellular Ca2+, whereas the attachment and ingestion of EBs occur independently of Ca2+ in the growth medium and at low intracellular free Ca2+ [Ca2+]i. Moreover, chlamydiae do not induce any changes in the level of [Ca2+]i, this indicates that the aggregation of EBs requires a normal homeostasis of intracellular Ca2+. By affecting F-act:in reorganization and, putatively, certain Ca2+ -binding proteins, [Ca2+]i plays a vital role in the process of chlamydiae infection. The ca2+_ and phospholipid-binding proteins, annex.ins, are selectivelytranslocated during ehlamydial infection, i.e. annexins III, IV, and V, but not annexins I and VI, translocate to the proximity of chlamydial aggregates and inclusions. Annexins differ in their ability to associate with chlamydia-containing vesicles and inclusions. This fact implies that different factors regulate the interaction of annexins I and Ill with the membrane and also suggests that there is a selective regulatorymechanism that guides endosome aggregation and that is responsible for endosome avoiding lysosome fusion during chlamydial infection. Chlamydiae also induce mobilization of intracellular Ca2+ stores. This suggests that in a chlamydia-infected cell localized [Ca2+]i changes may occur by mobilization of Ca2+ stores at the sites of ca2+ action. The physiological role of ca2+ stores redistribution during infection of the host cells with chlamydiae might be to generate subceJlular [Ca2+]i gradients needed for the intracellular itinerary of the membrane trafficking of BE-containing endosomes.
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| 5. |
- Majeed, Meytham, 1960-, et al.
(författare)
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Role of Src kinases and Syk in Fc? receptor-mediated phagocytosis and phagosome-lysosome fusion
- 2001
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Ingår i: Journal of Leukocyte Biology. - 0741-5400 .- 1938-3673. ; 70:5, s. 801-811
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Tidskriftsartikel (refereegranskat)abstract
- Phagocytosis is increased by Fc? receptors (Fc?Rs), and studies with syk-1- macrophages demonstrated that Syk kinase is required for Fc?TR phagocytosis. Similar studies with macrophages lacking the Src family kinases Hck, Fgr, and Lyn showed that these kinases are not required for phagocytosis but that they enhance the rate of particle engulfment. In this report we show that both wild-type and hck-1- fgr-1- macrophages expressed Fyn, Src, and Yes and that these kinases were activated on ingestion of immunoglobulin G (IgG)-coated particles and redistributed, together with Syk, to actin-rich phagocytic cups and the phagosomal membrane. At doses blocking IgG-dependent phagocytosis, the tyrosine kinase inhibitors PP1 and piceatannol inhibited both Src family kinase and Syk activities, as well as their redistribution to actin-rich phagocytic cups. Hck, Fgr, and Lyn were dispensable for lysosome-phagosome fusion (PLF) induced by IgG-coated particles. However, PP1 or piceatannol hampered unopsonized yeast-induced PLF despite the fact that they did not block yeast internalization.
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| 6. |
- Ydrenius, Liselotte, et al.
(författare)
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Activation of cAMP-dependent protein kinase is necessary for actin rearrangements in human neutrophils during phagocytosis
- 2000
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Ingår i: Journal of Leukocyte Biology. - 0741-5400 .- 1938-3673. ; 67:4, s. 520-528
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Tidskriftsartikel (refereegranskat)abstract
- We have investigated the role of cAMP and cAMP-dependent protein kinase (cAPK) in neutrophil phagocytosis. Inhibition of cAPK with H-89 reduced complement- and IgG-dependent phagocytosis to 83 and 46%, respectively. Fluorescence intensity measurements of phalloidin-stained actin in neutrophils showed a reduced amount of filamentous actin (F-actin) in pseudopods and around the phagosome in cells treated with H-89 or cAMP-elevating agents (forskolin and rolipram). The amount of phosphotyrosine-containing proteins was also reduced in pseudopods and around the phagosome. Taken together, the data show that cAMP/cAPK regulates F-actin reorganization during receptor-mediated phagocytosis, particularly triggered by IgG-FcR interaction. Our results support the hypothesis that active subcortical reorganization of F-actin is a prerequisite for FcR-mediated phagocytosis, but is less important during CR3-mediated ingestion.
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