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Impaired T lymphocyte trafficking in mice deficient in an actin-nucleating protein, mDia1.

Sakata D, Taniguchi H, Yasuda S, Adachi-Morishima A, Hamazaki Y, Nakayama R, Miki T, Minato N, Narumiya S - J. Exp. Med. (2007)

Bottom Line: Although much is known about the latter, the physiological functions of mDia proteins are unclear.Similarly, mDia1(-/-) thymocytes showed reduced chemotaxis and impaired egression from the thymus.These results suggest that mDia1 plays a distinct role in chemotaxis in T lymphocyte trafficking.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmacology,Kyoto University Faculty of Medicine, Kyoto 606-8501, Japan.

ABSTRACT
Trafficking of immune cells is controlled by directed migration of relevant cells toward chemotactic signals. Actin cytoskeleton undergoes continuous remodeling and serves as machinery for cell migration. The mDia family of formins and the Wiskott-Aldrich syndrome protein (WASP)-Arp2/3 system are two major actin nucleating-polymerizing systems in mammalian cells, with the former producing long straight actin filaments and the latter producing branched actin meshwork. Although much is known about the latter, the physiological functions of mDia proteins are unclear. We generated mice deficient in one mDia isoform, mDia1. Although mDia1(-/-) mice were born and developed without apparent abnormality, mDia1(-/-) T lymphocytes exhibited impaired trafficking to secondary lymphoid organs in vivo and showed reduced chemotaxis, little actin filament formation, and impaired polarity in response to chemotactic stimuli in vitro. Similarly, mDia1(-/-) thymocytes showed reduced chemotaxis and impaired egression from the thymus. These results suggest that mDia1 plays a distinct role in chemotaxis in T lymphocyte trafficking.

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Impaired chemokine-induced actin polymerization and polarization of mDia1−/− T cells. (A) F-actin production to chemokine stimulation. T cells from mDia1+/+ (blue line) and mDia1−/− (red line) mice were stimulated with two concentrations of CCL21 for the indicated times, stained with Oregon green–phalloidin, and analyzed by flow cytometry. Mean fluorescence intensity of the entire cell population was determined for each group and is shown with that of unstimulated mDia1+/+ cells as 100%. (B) Impaired chemokine responses of mDia1−/− T cells. T cells from mDia1+/+ and mDia1−/− mice were stimulated with or without CCL21 and stained for talin, F-actin, and DNA, as indicated. Bar, 10 μm.
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fig3: Impaired chemokine-induced actin polymerization and polarization of mDia1−/− T cells. (A) F-actin production to chemokine stimulation. T cells from mDia1+/+ (blue line) and mDia1−/− (red line) mice were stimulated with two concentrations of CCL21 for the indicated times, stained with Oregon green–phalloidin, and analyzed by flow cytometry. Mean fluorescence intensity of the entire cell population was determined for each group and is shown with that of unstimulated mDia1+/+ cells as 100%. (B) Impaired chemokine responses of mDia1−/− T cells. T cells from mDia1+/+ and mDia1−/− mice were stimulated with or without CCL21 and stained for talin, F-actin, and DNA, as indicated. Bar, 10 μm.

Mentions: Given the function of mDia1 in actin nucleation and polymerization (3, 4), we next examined the cellular response of mDia1−/− T cells to chemokine stimulation. The cells were initially stained with fluorescent phalloidin and subjected to flow cytometry, which revealed that the addition of CCL21 to wild-type T cells induced robust filamentous actin (F-actin) production in a time- and concentration-dependent manner. (Fig. 3 A). In contrast, T cells from mDia1−/− mice exhibited a substantially attenuated response at all time points measured, and the extent of attenuation was more marked at the lower chemokine concentration. Then the cells were stained for F-actin and talin. Microscopic examination of these cells revealed that wild-type T cells exhibited not only robust F-actin formation but also cell polarization, and that both F-actin production and cell polarization were markedly suppressed in mDia1−/− T cells (Fig. 3 B). On the other hand, the addition of CXCL13 induced both actin filament production and polarization in mDia1−/− B cells (Fig. S2, available at http://www.jem.org/cgi/content/full/jem.20062647/DC1).


Impaired T lymphocyte trafficking in mice deficient in an actin-nucleating protein, mDia1.

Sakata D, Taniguchi H, Yasuda S, Adachi-Morishima A, Hamazaki Y, Nakayama R, Miki T, Minato N, Narumiya S - J. Exp. Med. (2007)

Impaired chemokine-induced actin polymerization and polarization of mDia1−/− T cells. (A) F-actin production to chemokine stimulation. T cells from mDia1+/+ (blue line) and mDia1−/− (red line) mice were stimulated with two concentrations of CCL21 for the indicated times, stained with Oregon green–phalloidin, and analyzed by flow cytometry. Mean fluorescence intensity of the entire cell population was determined for each group and is shown with that of unstimulated mDia1+/+ cells as 100%. (B) Impaired chemokine responses of mDia1−/− T cells. T cells from mDia1+/+ and mDia1−/− mice were stimulated with or without CCL21 and stained for talin, F-actin, and DNA, as indicated. Bar, 10 μm.
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Related In: Results  -  Collection

Show All Figures
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fig3: Impaired chemokine-induced actin polymerization and polarization of mDia1−/− T cells. (A) F-actin production to chemokine stimulation. T cells from mDia1+/+ (blue line) and mDia1−/− (red line) mice were stimulated with two concentrations of CCL21 for the indicated times, stained with Oregon green–phalloidin, and analyzed by flow cytometry. Mean fluorescence intensity of the entire cell population was determined for each group and is shown with that of unstimulated mDia1+/+ cells as 100%. (B) Impaired chemokine responses of mDia1−/− T cells. T cells from mDia1+/+ and mDia1−/− mice were stimulated with or without CCL21 and stained for talin, F-actin, and DNA, as indicated. Bar, 10 μm.
Mentions: Given the function of mDia1 in actin nucleation and polymerization (3, 4), we next examined the cellular response of mDia1−/− T cells to chemokine stimulation. The cells were initially stained with fluorescent phalloidin and subjected to flow cytometry, which revealed that the addition of CCL21 to wild-type T cells induced robust filamentous actin (F-actin) production in a time- and concentration-dependent manner. (Fig. 3 A). In contrast, T cells from mDia1−/− mice exhibited a substantially attenuated response at all time points measured, and the extent of attenuation was more marked at the lower chemokine concentration. Then the cells were stained for F-actin and talin. Microscopic examination of these cells revealed that wild-type T cells exhibited not only robust F-actin formation but also cell polarization, and that both F-actin production and cell polarization were markedly suppressed in mDia1−/− T cells (Fig. 3 B). On the other hand, the addition of CXCL13 induced both actin filament production and polarization in mDia1−/− B cells (Fig. S2, available at http://www.jem.org/cgi/content/full/jem.20062647/DC1).

Bottom Line: Although much is known about the latter, the physiological functions of mDia proteins are unclear.Similarly, mDia1(-/-) thymocytes showed reduced chemotaxis and impaired egression from the thymus.These results suggest that mDia1 plays a distinct role in chemotaxis in T lymphocyte trafficking.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmacology,Kyoto University Faculty of Medicine, Kyoto 606-8501, Japan.

ABSTRACT
Trafficking of immune cells is controlled by directed migration of relevant cells toward chemotactic signals. Actin cytoskeleton undergoes continuous remodeling and serves as machinery for cell migration. The mDia family of formins and the Wiskott-Aldrich syndrome protein (WASP)-Arp2/3 system are two major actin nucleating-polymerizing systems in mammalian cells, with the former producing long straight actin filaments and the latter producing branched actin meshwork. Although much is known about the latter, the physiological functions of mDia proteins are unclear. We generated mice deficient in one mDia isoform, mDia1. Although mDia1(-/-) mice were born and developed without apparent abnormality, mDia1(-/-) T lymphocytes exhibited impaired trafficking to secondary lymphoid organs in vivo and showed reduced chemotaxis, little actin filament formation, and impaired polarity in response to chemotactic stimuli in vitro. Similarly, mDia1(-/-) thymocytes showed reduced chemotaxis and impaired egression from the thymus. These results suggest that mDia1 plays a distinct role in chemotaxis in T lymphocyte trafficking.

Show MeSH
Related in: MedlinePlus