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Comparisons of CapG and gelsolin- macrophages: demonstration of a unique role for CapG in receptor-mediated ruffling, phagocytosis, and vesicle rocketing.

Witke W, Li W, Kwiatkowski DJ, Southwick FS - J. Cell Biol. (2001)

Bottom Line: However, the loss of CapG in bone marrow macrophages profoundly inhibits macrophage colony stimulating factor-stimulated ruffling; reintroduction of CapG protein by microinjection fully restores this function.These motile functions are not impaired in gelsolin- macrophages and no additive effects are observed in CapG/gelsolin double- macrophages, establishing that CapG function is distinct from, and does not overlap with, gelsolin in macrophages.These primary effects on macrophage motile function suggest that CapG may be a useful target for the regulation of macrophage-mediated inflammatory responses.

View Article: PubMed Central - PubMed

Affiliation: Hematology Division, Department of Medicine, Brigham and Women's Hospital, Boston, MA 02115, USA.

ABSTRACT
Capping the barbed ends of actin filaments is a critical step for regulating actin-based motility in nonmuscle cells. The in vivo function of CapG, a calcium-sensitive barbed end capping protein and member of the gelsolin/villin family, has been assessed using a Capg allele engineered into mice. Both CapG- mice and CapG/gelsolin double- mice appear normal and have no gross functional abnormalities. However, the loss of CapG in bone marrow macrophages profoundly inhibits macrophage colony stimulating factor-stimulated ruffling; reintroduction of CapG protein by microinjection fully restores this function. CapG- macrophages also demonstrate approximately 50% impairment of immunoglobulin G, and complement-opsonized phagocytosis and lanthanum-induced vesicle rocketing. These motile functions are not impaired in gelsolin- macrophages and no additive effects are observed in CapG/gelsolin double- macrophages, establishing that CapG function is distinct from, and does not overlap with, gelsolin in macrophages. Our observations indicate that CapG is required for receptor-mediated ruffling, and that it is a major functional component of macrophage phagocytosis. These primary effects on macrophage motile function suggest that CapG may be a useful target for the regulation of macrophage-mediated inflammatory responses.

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Time-lapse phase micrographs of endosomal rocketing in wild-type and Capg−/− macrophages. Cells were treated with lanthanum and zinc chloride as described in the Materials and methods. This treatment induces the formation of vesicles that move through the cytoplasm by an actin-based motor. (A) The left-hand column shows a vesicle rocketing through a wild-type macrophage (arrow). (B) The right-hand column shows a vesicle migrating through a Capg−/− macrophage (arrow). Time is depicted in the upper left-hand corner. Note the long phase-dense actin tail in the wild-type cell (A) and the very short actin tail in the Capg−/− cell (B). Phase-dense tails were rarely seen behind rocketing vesicles in Capg−/−, but were commonly seen in wild-type cells. The average velocity of rocketing vesicles in Capg−/− macrophages was <1/2 that of wild-type cells. Bar, 10 mm.
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fig6: Time-lapse phase micrographs of endosomal rocketing in wild-type and Capg−/− macrophages. Cells were treated with lanthanum and zinc chloride as described in the Materials and methods. This treatment induces the formation of vesicles that move through the cytoplasm by an actin-based motor. (A) The left-hand column shows a vesicle rocketing through a wild-type macrophage (arrow). (B) The right-hand column shows a vesicle migrating through a Capg−/− macrophage (arrow). Time is depicted in the upper left-hand corner. Note the long phase-dense actin tail in the wild-type cell (A) and the very short actin tail in the Capg−/− cell (B). Phase-dense tails were rarely seen behind rocketing vesicles in Capg−/−, but were commonly seen in wild-type cells. The average velocity of rocketing vesicles in Capg−/− macrophages was <1/2 that of wild-type cells. Bar, 10 mm.

Mentions: When wild-type macrophages were exposed to lanthanum hydrogen chloride for 10 min followed by zinc, these cells formed multiple vesicles within their cytoplasm (see Materials and methods). After 20–30 min, ∼2% of the vesicles began moving within the cytoplasm at velocities ranging from 0.05 to 0.12 μm/s. Movement was associated with the formation of actin filament tails (Zeile et al., 2000). Similar treatment of Capg−/− macrophages led to the formation of fewer vesicles compared with wild-type macrophages, although this difference did not achieve statistical significance (mean of 46 ± 7.4 vesicles/Capg−/− cell vs. 77 ± 23/wild-type cell; n = 12–17 cells, P = 0.184). However, phase-dense rocket tails were shorter or absent in Capg−/− cells (Fig. 6) and the velocities of motile vesicles were less than half that of wild-type cells (mean of 0.08 ± 0.002 μm/s, n = 93 vs. Capg−/− cells 0.03 ± 0.002 μm/s; n = 66, P < 0.0001).


Comparisons of CapG and gelsolin- macrophages: demonstration of a unique role for CapG in receptor-mediated ruffling, phagocytosis, and vesicle rocketing.

Witke W, Li W, Kwiatkowski DJ, Southwick FS - J. Cell Biol. (2001)

Time-lapse phase micrographs of endosomal rocketing in wild-type and Capg−/− macrophages. Cells were treated with lanthanum and zinc chloride as described in the Materials and methods. This treatment induces the formation of vesicles that move through the cytoplasm by an actin-based motor. (A) The left-hand column shows a vesicle rocketing through a wild-type macrophage (arrow). (B) The right-hand column shows a vesicle migrating through a Capg−/− macrophage (arrow). Time is depicted in the upper left-hand corner. Note the long phase-dense actin tail in the wild-type cell (A) and the very short actin tail in the Capg−/− cell (B). Phase-dense tails were rarely seen behind rocketing vesicles in Capg−/−, but were commonly seen in wild-type cells. The average velocity of rocketing vesicles in Capg−/− macrophages was <1/2 that of wild-type cells. Bar, 10 mm.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2196452&req=5

fig6: Time-lapse phase micrographs of endosomal rocketing in wild-type and Capg−/− macrophages. Cells were treated with lanthanum and zinc chloride as described in the Materials and methods. This treatment induces the formation of vesicles that move through the cytoplasm by an actin-based motor. (A) The left-hand column shows a vesicle rocketing through a wild-type macrophage (arrow). (B) The right-hand column shows a vesicle migrating through a Capg−/− macrophage (arrow). Time is depicted in the upper left-hand corner. Note the long phase-dense actin tail in the wild-type cell (A) and the very short actin tail in the Capg−/− cell (B). Phase-dense tails were rarely seen behind rocketing vesicles in Capg−/−, but were commonly seen in wild-type cells. The average velocity of rocketing vesicles in Capg−/− macrophages was <1/2 that of wild-type cells. Bar, 10 mm.
Mentions: When wild-type macrophages were exposed to lanthanum hydrogen chloride for 10 min followed by zinc, these cells formed multiple vesicles within their cytoplasm (see Materials and methods). After 20–30 min, ∼2% of the vesicles began moving within the cytoplasm at velocities ranging from 0.05 to 0.12 μm/s. Movement was associated with the formation of actin filament tails (Zeile et al., 2000). Similar treatment of Capg−/− macrophages led to the formation of fewer vesicles compared with wild-type macrophages, although this difference did not achieve statistical significance (mean of 46 ± 7.4 vesicles/Capg−/− cell vs. 77 ± 23/wild-type cell; n = 12–17 cells, P = 0.184). However, phase-dense rocket tails were shorter or absent in Capg−/− cells (Fig. 6) and the velocities of motile vesicles were less than half that of wild-type cells (mean of 0.08 ± 0.002 μm/s, n = 93 vs. Capg−/− cells 0.03 ± 0.002 μm/s; n = 66, P < 0.0001).

Bottom Line: However, the loss of CapG in bone marrow macrophages profoundly inhibits macrophage colony stimulating factor-stimulated ruffling; reintroduction of CapG protein by microinjection fully restores this function.These motile functions are not impaired in gelsolin- macrophages and no additive effects are observed in CapG/gelsolin double- macrophages, establishing that CapG function is distinct from, and does not overlap with, gelsolin in macrophages.These primary effects on macrophage motile function suggest that CapG may be a useful target for the regulation of macrophage-mediated inflammatory responses.

View Article: PubMed Central - PubMed

Affiliation: Hematology Division, Department of Medicine, Brigham and Women's Hospital, Boston, MA 02115, USA.

ABSTRACT
Capping the barbed ends of actin filaments is a critical step for regulating actin-based motility in nonmuscle cells. The in vivo function of CapG, a calcium-sensitive barbed end capping protein and member of the gelsolin/villin family, has been assessed using a Capg allele engineered into mice. Both CapG- mice and CapG/gelsolin double- mice appear normal and have no gross functional abnormalities. However, the loss of CapG in bone marrow macrophages profoundly inhibits macrophage colony stimulating factor-stimulated ruffling; reintroduction of CapG protein by microinjection fully restores this function. CapG- macrophages also demonstrate approximately 50% impairment of immunoglobulin G, and complement-opsonized phagocytosis and lanthanum-induced vesicle rocketing. These motile functions are not impaired in gelsolin- macrophages and no additive effects are observed in CapG/gelsolin double- macrophages, establishing that CapG function is distinct from, and does not overlap with, gelsolin in macrophages. Our observations indicate that CapG is required for receptor-mediated ruffling, and that it is a major functional component of macrophage phagocytosis. These primary effects on macrophage motile function suggest that CapG may be a useful target for the regulation of macrophage-mediated inflammatory responses.

Show MeSH
Related in: MedlinePlus