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LFA-1-mediated adhesion is regulated by cytoskeletal restraint and by a Ca2+-dependent protease, calpain.

Stewart MP, McDowall A, Hogg N - J. Cell Biol. (1998)

Bottom Line: The activity of integrins on leukocytes is kept under tight control to avoid inappropriate adhesion while these cells are circulating in blood or migrating through tissues.We report here that after exposure of T cells to these agonists, integrin is released from cytoskeletal control by the Ca2+-induced activation of a calpain-like enzyme, and adhesive contact between cells is strengthened by means of the clustering of mobilized LFA-1 on the membrane.We propose that methods of leukocyte stimulation that cause Ca2+ fluxes induce LFA-1 adhesion by regulation of calpain activity.

View Article: PubMed Central - PubMed

Affiliation: Leukocyte Adhesion Laboratory, Imperial Cancer Research Fund, Lincoln's Inn Fields, London WC2A 3PX, United Kingdom.

ABSTRACT
The activity of integrins on leukocytes is kept under tight control to avoid inappropriate adhesion while these cells are circulating in blood or migrating through tissues. Using lymphocyte function-associated antigen-1 (LFA-1) on T cells as a model, we have investigated adhesion to ligand intercellular adhesion molecule-1 induced by the Ca2+ mobilizers, ionomycin, 2, 5-di-t-butylhydroquinone, and thapsigargin, and the well studied stimulators such as phorbol ester and cross-linking of the antigen-specific T cell receptor (TCR)-CD3 complex. We report here that after exposure of T cells to these agonists, integrin is released from cytoskeletal control by the Ca2+-induced activation of a calpain-like enzyme, and adhesive contact between cells is strengthened by means of the clustering of mobilized LFA-1 on the membrane. We propose that methods of leukocyte stimulation that cause Ca2+ fluxes induce LFA-1 adhesion by regulation of calpain activity. These findings suggest a mechanism whereby engagement of the TCR could promote adhesion strengthening at an early stage of interaction with an antigen-presenting cell.

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LFA-1 clustering induced by thapsigargin and anti-TCR–CD3 triggering is inhibited by calpeptin. Clustering of  LFA-1 induced by 5 μM thapsigargin (a) and 10 μg/ml anti-CD3  mAb (b) was prevented by preincubation with 100 μg/ml calpeptin, (c and d, respectively), for 30 min at 37°C. Bar, 10 μm.
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Figure 7: LFA-1 clustering induced by thapsigargin and anti-TCR–CD3 triggering is inhibited by calpeptin. Clustering of LFA-1 induced by 5 μM thapsigargin (a) and 10 μg/ml anti-CD3 mAb (b) was prevented by preincubation with 100 μg/ml calpeptin, (c and d, respectively), for 30 min at 37°C. Bar, 10 μm.

Mentions: It was observed that some of the brightest LFA-1 fluorescence was found where cells were in contact with each other, and we therefore quantified the level of fluorescence in these regions (Table I, Contact zone) to determine whether this high fluorescence reflected a redistribution of LFA-1 to cell–cell contacts or was merely the additive value of the two cell membrane measurements. As the ratio of the average fluorescence intensity at cell contact areas to that of free membrane was ∼2 (Table I, Contact/Free), we concluded that there is no large scale redistribution of LFA-1 to points of cell contact upon thapsigargin treatment. In summary, the confocal results revealed that upon thapsigargin stimulation LFA-1 becomes clustered but does not change its distribution to regions of the cell membrane in contact with other cells. Similar results were obtained with the other Ca2+-mobilizing agents as well as PdBu (data not shown), and by cross-linking the TCR/CD3 (see Fig. 7) suggesting that local LFA-1 clustering may be a general feature of several T cell adhesion- activating protocols.


LFA-1-mediated adhesion is regulated by cytoskeletal restraint and by a Ca2+-dependent protease, calpain.

Stewart MP, McDowall A, Hogg N - J. Cell Biol. (1998)

LFA-1 clustering induced by thapsigargin and anti-TCR–CD3 triggering is inhibited by calpeptin. Clustering of  LFA-1 induced by 5 μM thapsigargin (a) and 10 μg/ml anti-CD3  mAb (b) was prevented by preincubation with 100 μg/ml calpeptin, (c and d, respectively), for 30 min at 37°C. Bar, 10 μm.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2140165&req=5

Figure 7: LFA-1 clustering induced by thapsigargin and anti-TCR–CD3 triggering is inhibited by calpeptin. Clustering of LFA-1 induced by 5 μM thapsigargin (a) and 10 μg/ml anti-CD3 mAb (b) was prevented by preincubation with 100 μg/ml calpeptin, (c and d, respectively), for 30 min at 37°C. Bar, 10 μm.
Mentions: It was observed that some of the brightest LFA-1 fluorescence was found where cells were in contact with each other, and we therefore quantified the level of fluorescence in these regions (Table I, Contact zone) to determine whether this high fluorescence reflected a redistribution of LFA-1 to cell–cell contacts or was merely the additive value of the two cell membrane measurements. As the ratio of the average fluorescence intensity at cell contact areas to that of free membrane was ∼2 (Table I, Contact/Free), we concluded that there is no large scale redistribution of LFA-1 to points of cell contact upon thapsigargin treatment. In summary, the confocal results revealed that upon thapsigargin stimulation LFA-1 becomes clustered but does not change its distribution to regions of the cell membrane in contact with other cells. Similar results were obtained with the other Ca2+-mobilizing agents as well as PdBu (data not shown), and by cross-linking the TCR/CD3 (see Fig. 7) suggesting that local LFA-1 clustering may be a general feature of several T cell adhesion- activating protocols.

Bottom Line: The activity of integrins on leukocytes is kept under tight control to avoid inappropriate adhesion while these cells are circulating in blood or migrating through tissues.We report here that after exposure of T cells to these agonists, integrin is released from cytoskeletal control by the Ca2+-induced activation of a calpain-like enzyme, and adhesive contact between cells is strengthened by means of the clustering of mobilized LFA-1 on the membrane.We propose that methods of leukocyte stimulation that cause Ca2+ fluxes induce LFA-1 adhesion by regulation of calpain activity.

View Article: PubMed Central - PubMed

Affiliation: Leukocyte Adhesion Laboratory, Imperial Cancer Research Fund, Lincoln's Inn Fields, London WC2A 3PX, United Kingdom.

ABSTRACT
The activity of integrins on leukocytes is kept under tight control to avoid inappropriate adhesion while these cells are circulating in blood or migrating through tissues. Using lymphocyte function-associated antigen-1 (LFA-1) on T cells as a model, we have investigated adhesion to ligand intercellular adhesion molecule-1 induced by the Ca2+ mobilizers, ionomycin, 2, 5-di-t-butylhydroquinone, and thapsigargin, and the well studied stimulators such as phorbol ester and cross-linking of the antigen-specific T cell receptor (TCR)-CD3 complex. We report here that after exposure of T cells to these agonists, integrin is released from cytoskeletal control by the Ca2+-induced activation of a calpain-like enzyme, and adhesive contact between cells is strengthened by means of the clustering of mobilized LFA-1 on the membrane. We propose that methods of leukocyte stimulation that cause Ca2+ fluxes induce LFA-1 adhesion by regulation of calpain activity. These findings suggest a mechanism whereby engagement of the TCR could promote adhesion strengthening at an early stage of interaction with an antigen-presenting cell.

Show MeSH
Related in: MedlinePlus