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Phosphatidylinositol 3-kinase activity and asymmetrical accumulation of F-actin are necessary for establishment of cell polarity in the early development of monospores from the marine red alga Porphyra yezoensis.

Li L, Saga N, Mikami K - J. Exp. Bot. (2008)

Bottom Line: Our results indicate that the asymmetrical localization of F-actin at the leading edge is fixed by the establishment of the anterior-posterior axis in migrating monospores, which is PI3K-dependent and protein synthesis-independent.These findings suggest that the establishment of anterior-posterior and apical-basal axes are differentially regulated during the early development of monospores.Our results also indicate that PI3K-dependent F-actin asymmetry is evolutionally conserved in relation to the establishment of cell polarity in migrating eukaryotic cells.

View Article: PubMed Central - PubMed

Affiliation: Graduate School of Fisheries Sciences, Hokkaido University, 3-1-1 Minato-cho, Hakodate 041-8611, Japan.

ABSTRACT
The polarized distribution of F-actin is important in providing the driving force for directional migration in mammalian leukocytes and Dictyostelium cells, in which compartmentation of phosphatidylinositol 3-kinase (PI3K) and phosphatidylinositol phosphatase is critical for the establishment of cell polarity. Since monospores from the red alga Porphyra yezoensis are a real example of migrating plant cells, the involvement of the cytoskeleton and PI3K was investigated during their early development. Our results indicate that the asymmetrical localization of F-actin at the leading edge is fixed by the establishment of the anterior-posterior axis in migrating monospores, which is PI3K-dependent and protein synthesis-independent. After migration, monospores adhere to the substratum and then become upright, developing into multicellular thalli via the establishment of the apical-basal axis. In this process, F-actin usually accumulates at the bottom of the basal cell and development after migration requires new protein synthesis. These findings suggest that the establishment of anterior-posterior and apical-basal axes are differentially regulated during the early development of monospores. Our results also indicate that PI3K-dependent F-actin asymmetry is evolutionally conserved in relation to the establishment of cell polarity in migrating eukaryotic cells.

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Effects of the PI3K inhibitor on the motility and becoming upright of monospores. (A) Effect of LY294002 on morphology and asymmetrical F-actin accumulation. Freshly released monospores were treated with LY294002 for 3 h. F-actin was stained with Alex Flour 488 phalloidin. Upper and lower photographs in each panel show bright-field and fluorescent images, respectively. (a) Migrating monospores grown in ESL medium containing 0.3% DMSO. (b–f) Monospores incubated with an increasing concentration of LY294002 (b, 0.5 μM; c, 2.5 μM; d, 5 μM; e, 10 μM; f, 15 μM). Scale bar = 5 μm. (B) Effects of LY294002 and its analogue LY303511 on the early development of monospores. Freshly released monospores were treated with an increasing concentration of LY294002 (a, c) and LY303511(b, d) for 3 h (a, b) and 24 h (c, d). (a, b) Effects on migration. (c, d) Effects on formation of germlings. Data are presented as mean ±SD (n=3).
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fig4: Effects of the PI3K inhibitor on the motility and becoming upright of monospores. (A) Effect of LY294002 on morphology and asymmetrical F-actin accumulation. Freshly released monospores were treated with LY294002 for 3 h. F-actin was stained with Alex Flour 488 phalloidin. Upper and lower photographs in each panel show bright-field and fluorescent images, respectively. (a) Migrating monospores grown in ESL medium containing 0.3% DMSO. (b–f) Monospores incubated with an increasing concentration of LY294002 (b, 0.5 μM; c, 2.5 μM; d, 5 μM; e, 10 μM; f, 15 μM). Scale bar = 5 μm. (B) Effects of LY294002 and its analogue LY303511 on the early development of monospores. Freshly released monospores were treated with an increasing concentration of LY294002 (a, c) and LY303511(b, d) for 3 h (a, b) and 24 h (c, d). (a, b) Effects on migration. (c, d) Effects on formation of germlings. Data are presented as mean ±SD (n=3).

Mentions: As show in Fig. 3Aa, migrating monospores formed a tapered shape as in Dictyostelium cells and leukocytes, which prompted us to examine the possible involvement of PI3K and PI(3,4,5)P3-phosphatase in the movement of P. yezoensis monospores. First, monospores were treated with a gradually increasing concentration of LY294002 and LY303511, the specific inhibitor of PI3K and its analogue, respectively. After 3 h incubation, the polarized localization of F-actin was gradually prevented with an increasing concentration of LY294002 along with morphological changes, whereas about 84% of the monospores started movement in the control medium (Fig. 4A). In parallel with this, the migration of monospores and the formation of germlings decreased in a dose-dependent manner during treatment with LY294002 for 3 h and 24 h (Fig. 4Ba, c). By contrast, LY303511 had no effect (Fig. 4Bb, d). Moreover, the effect of LY294002 was reversible after washing the monospores (data not shown). These results indicate that PI3K activity is involved in migration through the formation of the anterior–posterior axis and the regulation of asymmetrical F-actin accumulation at leading edges. However, PI3K does not play a role in adhesion and becoming upright of monospores, since migrating monospores developed to germlings in the presence of LY294002 (Fig. 4Bc).


Phosphatidylinositol 3-kinase activity and asymmetrical accumulation of F-actin are necessary for establishment of cell polarity in the early development of monospores from the marine red alga Porphyra yezoensis.

Li L, Saga N, Mikami K - J. Exp. Bot. (2008)

Effects of the PI3K inhibitor on the motility and becoming upright of monospores. (A) Effect of LY294002 on morphology and asymmetrical F-actin accumulation. Freshly released monospores were treated with LY294002 for 3 h. F-actin was stained with Alex Flour 488 phalloidin. Upper and lower photographs in each panel show bright-field and fluorescent images, respectively. (a) Migrating monospores grown in ESL medium containing 0.3% DMSO. (b–f) Monospores incubated with an increasing concentration of LY294002 (b, 0.5 μM; c, 2.5 μM; d, 5 μM; e, 10 μM; f, 15 μM). Scale bar = 5 μm. (B) Effects of LY294002 and its analogue LY303511 on the early development of monospores. Freshly released monospores were treated with an increasing concentration of LY294002 (a, c) and LY303511(b, d) for 3 h (a, b) and 24 h (c, d). (a, b) Effects on migration. (c, d) Effects on formation of germlings. Data are presented as mean ±SD (n=3).
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
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fig4: Effects of the PI3K inhibitor on the motility and becoming upright of monospores. (A) Effect of LY294002 on morphology and asymmetrical F-actin accumulation. Freshly released monospores were treated with LY294002 for 3 h. F-actin was stained with Alex Flour 488 phalloidin. Upper and lower photographs in each panel show bright-field and fluorescent images, respectively. (a) Migrating monospores grown in ESL medium containing 0.3% DMSO. (b–f) Monospores incubated with an increasing concentration of LY294002 (b, 0.5 μM; c, 2.5 μM; d, 5 μM; e, 10 μM; f, 15 μM). Scale bar = 5 μm. (B) Effects of LY294002 and its analogue LY303511 on the early development of monospores. Freshly released monospores were treated with an increasing concentration of LY294002 (a, c) and LY303511(b, d) for 3 h (a, b) and 24 h (c, d). (a, b) Effects on migration. (c, d) Effects on formation of germlings. Data are presented as mean ±SD (n=3).
Mentions: As show in Fig. 3Aa, migrating monospores formed a tapered shape as in Dictyostelium cells and leukocytes, which prompted us to examine the possible involvement of PI3K and PI(3,4,5)P3-phosphatase in the movement of P. yezoensis monospores. First, monospores were treated with a gradually increasing concentration of LY294002 and LY303511, the specific inhibitor of PI3K and its analogue, respectively. After 3 h incubation, the polarized localization of F-actin was gradually prevented with an increasing concentration of LY294002 along with morphological changes, whereas about 84% of the monospores started movement in the control medium (Fig. 4A). In parallel with this, the migration of monospores and the formation of germlings decreased in a dose-dependent manner during treatment with LY294002 for 3 h and 24 h (Fig. 4Ba, c). By contrast, LY303511 had no effect (Fig. 4Bb, d). Moreover, the effect of LY294002 was reversible after washing the monospores (data not shown). These results indicate that PI3K activity is involved in migration through the formation of the anterior–posterior axis and the regulation of asymmetrical F-actin accumulation at leading edges. However, PI3K does not play a role in adhesion and becoming upright of monospores, since migrating monospores developed to germlings in the presence of LY294002 (Fig. 4Bc).

Bottom Line: Our results indicate that the asymmetrical localization of F-actin at the leading edge is fixed by the establishment of the anterior-posterior axis in migrating monospores, which is PI3K-dependent and protein synthesis-independent.These findings suggest that the establishment of anterior-posterior and apical-basal axes are differentially regulated during the early development of monospores.Our results also indicate that PI3K-dependent F-actin asymmetry is evolutionally conserved in relation to the establishment of cell polarity in migrating eukaryotic cells.

View Article: PubMed Central - PubMed

Affiliation: Graduate School of Fisheries Sciences, Hokkaido University, 3-1-1 Minato-cho, Hakodate 041-8611, Japan.

ABSTRACT
The polarized distribution of F-actin is important in providing the driving force for directional migration in mammalian leukocytes and Dictyostelium cells, in which compartmentation of phosphatidylinositol 3-kinase (PI3K) and phosphatidylinositol phosphatase is critical for the establishment of cell polarity. Since monospores from the red alga Porphyra yezoensis are a real example of migrating plant cells, the involvement of the cytoskeleton and PI3K was investigated during their early development. Our results indicate that the asymmetrical localization of F-actin at the leading edge is fixed by the establishment of the anterior-posterior axis in migrating monospores, which is PI3K-dependent and protein synthesis-independent. After migration, monospores adhere to the substratum and then become upright, developing into multicellular thalli via the establishment of the apical-basal axis. In this process, F-actin usually accumulates at the bottom of the basal cell and development after migration requires new protein synthesis. These findings suggest that the establishment of anterior-posterior and apical-basal axes are differentially regulated during the early development of monospores. Our results also indicate that PI3K-dependent F-actin asymmetry is evolutionally conserved in relation to the establishment of cell polarity in migrating eukaryotic cells.

Show MeSH