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Assembly and positioning of actomyosin rings by contractility and planar cell polarity.

Sehring IM, Recho P, Denker E, Kourakis M, Mathiesen B, Hannezo E, Dong B, Jiang D - Elife (2015)

Bottom Line: Intriguingly, rings always form at the cells' anterior edge before migrating towards the center as contractility increases, reflecting a novel dynamical property of the cortex.We develop a simple model of the physical forces underlying this tug-of-war, which quantitatively reproduces our results.We thus propose a quantitative framework for dissecting the relative contribution of contractility and PCP to the self-assembly and repositioning of cytoskeletal structures, which should be applicable to other morphogenetic events.

View Article: PubMed Central - PubMed

Affiliation: Sars International Centre for Marine Molecular Biology, University of Bergen, Bergen, Norway.

ABSTRACT
The actomyosin cytoskeleton is a primary force-generating mechanism in morphogenesis, thus a robust spatial control of cytoskeletal positioning is essential. In this report, we demonstrate that actomyosin contractility and planar cell polarity (PCP) interact in post-mitotic Ciona notochord cells to self-assemble and reposition actomyosin rings, which play an essential role for cell elongation. Intriguingly, rings always form at the cells' anterior edge before migrating towards the center as contractility increases, reflecting a novel dynamical property of the cortex. Our drug and genetic manipulations uncover a tug-of-war between contractility, which localizes cortical flows toward the equator and PCP, which tries to reposition them. We develop a simple model of the physical forces underlying this tug-of-war, which quantitatively reproduces our results. We thus propose a quantitative framework for dissecting the relative contribution of contractility and PCP to the self-assembly and repositioning of cytoskeletal structures, which should be applicable to other morphogenetic events.

No MeSH data available.


Related in: MedlinePlus

Circumferential actin rings are shifted anteriorly in Halocynthia roretzi notochord cells with centrally localized nuclei.(A, B) Halocynthia notochord cells elongate from coin-shaped (A) to drum-shaped (B). A circumferential constriction appears at the equator of the cylindrical cell (arrowhead). The nucleus (asterisk) is localized in the center of each cell. (C, D) Cortical F-actin (arrow in C) and MRLC (arrow in D) accumulate at the equatorial region of the basal domain. (E, F) Notochord elongation (E, DMSO-treated) is abolished in Halocynthia embryos treated with 100 μM blebbistatin for 16 hr (F) starting at the onset of cell elongation (27 hpf). (G) Circumferential actin rings (white arrowheads) are shifted anteriorly after 3-hr blebbistatin treatment at 31 hpf (13°C). Similar to what is observed in Ciona notochord cells (Figure 2C', H), the shifted ring is associated with a circumferential bulge (white arrowheads), whereas the constriction is located posterior to the ring (red arrowheads). These results indicate (1) a conservation of the equatorial actomyosin contractile mechanism to drive notochord cell elongation in Halocynthia, (2) that the position of the ring does not influence the position of the nucleus, (3) the position of the nucleus does not influence the position of the ring, and (4) the nucleus position does not affect the direction of the ring shift. Anterior to the left in panels A–B, G. Scale bars in E and F, 50 μm; in all others, 10 μm.DOI:http://dx.doi.org/10.7554/eLife.09206.013
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fig3: Circumferential actin rings are shifted anteriorly in Halocynthia roretzi notochord cells with centrally localized nuclei.(A, B) Halocynthia notochord cells elongate from coin-shaped (A) to drum-shaped (B). A circumferential constriction appears at the equator of the cylindrical cell (arrowhead). The nucleus (asterisk) is localized in the center of each cell. (C, D) Cortical F-actin (arrow in C) and MRLC (arrow in D) accumulate at the equatorial region of the basal domain. (E, F) Notochord elongation (E, DMSO-treated) is abolished in Halocynthia embryos treated with 100 μM blebbistatin for 16 hr (F) starting at the onset of cell elongation (27 hpf). (G) Circumferential actin rings (white arrowheads) are shifted anteriorly after 3-hr blebbistatin treatment at 31 hpf (13°C). Similar to what is observed in Ciona notochord cells (Figure 2C', H), the shifted ring is associated with a circumferential bulge (white arrowheads), whereas the constriction is located posterior to the ring (red arrowheads). These results indicate (1) a conservation of the equatorial actomyosin contractile mechanism to drive notochord cell elongation in Halocynthia, (2) that the position of the ring does not influence the position of the nucleus, (3) the position of the nucleus does not influence the position of the ring, and (4) the nucleus position does not affect the direction of the ring shift. Anterior to the left in panels A–B, G. Scale bars in E and F, 50 μm; in all others, 10 μm.DOI:http://dx.doi.org/10.7554/eLife.09206.013

Mentions: As notochord cells are planarly polarized, and ring migration is always unidirectional in wild-type embryo, this prompted us to examine the effect of A/P polarity on actin ring migration. A conspicuous feature of the A/P polarity is the posterior localization of the nucleus, which is regulated by prickle, a core PCP component. The Ciona savignyi mutant aimless carries a deletion in prickle, resulting in a loss of A/P polarity manifested in the randomized localization of nucleus in addition to an earlier convergent extension defect (Jiang et al., 2005). We first explored the possibility that the posterior nucleus might influence the direction of ring movement mobilized by blebbistatin. To this end, we examined the notochord of the ascidian Halocynthia roretzi, which follows a remarkably similar early developmental process as in Ciona (Figure 3A,B), except at the cell elongation stage, the nuclei are positioned in the center of the cells (asterisks in Figure 3B). A conspicuous circumferential constriction is present at the equator, which is colocalized with a cortical ring of actin (Figure 3C) and activated myosin (Figure 3D), whose activity is essential for notochord cell elongation (Figure 3E,F). We next treated embryos with elongating notochord cells with blebbistatin for 3 hr. Similar to what was observed in Ciona, the actin ring was shifted invariably anteriorly (Figure 3G), indicating that the position of the nucleus does not influence the dynamic behavior of the ring. We thus used aimless embryos to explore the role of a compromised A/P polarity on the repositioning of the ring and its shifting upon blebbistatin treatment, independent from its influence on nuclear position. In wild-type C. savignyi embryos, elongated notochord cells possess a circumferential actin ring at the cell equator and a posterior nucleus (Figure 4A). Blebbistatin treatment shifts the ring towards the anterior pole (Figure 4B), mirroring exactly the events in C. intestinalis. In aimless embryos, the intercalation of notochord cells (outlined in Figure 4C) is impaired, except in the posterior region where the cells often align into a single file (Jiang et al., 2005). In these cells, the loss of A/P polarity is evident by the random position of the nuclei (Figure 4C insert). In 92% of mock-treated aimless cells in this region, the actin ring is positioned at the equator (Figure 4C insert), showing that equatorial ring positioning is independent from PCP. However, reducing contractility in polarity-deficient mutant cells led to a dramatically different phenotype: after 60-min blebbistatin treatment, no unidirectional migration towards the anterior was observed. The direction of the shift was randomized and independent of the localization of the nucleus: examples for anterior nucleus and ring, anterior nucleus and posterior ring, posterior nucleus and ring, and posterior nucleus and anterior ring could be found (Figure 4D,D'). In 56% of the cells, the ring was still positioned at the equator, and in the rest, the ring migrated either to the anterior or posterior side (18% and 26%) (n = 57) (Figure 4E). These results imply that A/P polarity by prickle is not necessary for the establishment of an equatorial actin ring; however, it is instrumental for the direction of movement of the ring upon blebbistatin treatment.10.7554/eLife.09206.013Figure 3.Circumferential actin rings are shifted anteriorly in Halocynthia roretzi notochord cells with centrally localized nuclei.


Assembly and positioning of actomyosin rings by contractility and planar cell polarity.

Sehring IM, Recho P, Denker E, Kourakis M, Mathiesen B, Hannezo E, Dong B, Jiang D - Elife (2015)

Circumferential actin rings are shifted anteriorly in Halocynthia roretzi notochord cells with centrally localized nuclei.(A, B) Halocynthia notochord cells elongate from coin-shaped (A) to drum-shaped (B). A circumferential constriction appears at the equator of the cylindrical cell (arrowhead). The nucleus (asterisk) is localized in the center of each cell. (C, D) Cortical F-actin (arrow in C) and MRLC (arrow in D) accumulate at the equatorial region of the basal domain. (E, F) Notochord elongation (E, DMSO-treated) is abolished in Halocynthia embryos treated with 100 μM blebbistatin for 16 hr (F) starting at the onset of cell elongation (27 hpf). (G) Circumferential actin rings (white arrowheads) are shifted anteriorly after 3-hr blebbistatin treatment at 31 hpf (13°C). Similar to what is observed in Ciona notochord cells (Figure 2C', H), the shifted ring is associated with a circumferential bulge (white arrowheads), whereas the constriction is located posterior to the ring (red arrowheads). These results indicate (1) a conservation of the equatorial actomyosin contractile mechanism to drive notochord cell elongation in Halocynthia, (2) that the position of the ring does not influence the position of the nucleus, (3) the position of the nucleus does not influence the position of the ring, and (4) the nucleus position does not affect the direction of the ring shift. Anterior to the left in panels A–B, G. Scale bars in E and F, 50 μm; in all others, 10 μm.DOI:http://dx.doi.org/10.7554/eLife.09206.013
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Related In: Results  -  Collection

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fig3: Circumferential actin rings are shifted anteriorly in Halocynthia roretzi notochord cells with centrally localized nuclei.(A, B) Halocynthia notochord cells elongate from coin-shaped (A) to drum-shaped (B). A circumferential constriction appears at the equator of the cylindrical cell (arrowhead). The nucleus (asterisk) is localized in the center of each cell. (C, D) Cortical F-actin (arrow in C) and MRLC (arrow in D) accumulate at the equatorial region of the basal domain. (E, F) Notochord elongation (E, DMSO-treated) is abolished in Halocynthia embryos treated with 100 μM blebbistatin for 16 hr (F) starting at the onset of cell elongation (27 hpf). (G) Circumferential actin rings (white arrowheads) are shifted anteriorly after 3-hr blebbistatin treatment at 31 hpf (13°C). Similar to what is observed in Ciona notochord cells (Figure 2C', H), the shifted ring is associated with a circumferential bulge (white arrowheads), whereas the constriction is located posterior to the ring (red arrowheads). These results indicate (1) a conservation of the equatorial actomyosin contractile mechanism to drive notochord cell elongation in Halocynthia, (2) that the position of the ring does not influence the position of the nucleus, (3) the position of the nucleus does not influence the position of the ring, and (4) the nucleus position does not affect the direction of the ring shift. Anterior to the left in panels A–B, G. Scale bars in E and F, 50 μm; in all others, 10 μm.DOI:http://dx.doi.org/10.7554/eLife.09206.013
Mentions: As notochord cells are planarly polarized, and ring migration is always unidirectional in wild-type embryo, this prompted us to examine the effect of A/P polarity on actin ring migration. A conspicuous feature of the A/P polarity is the posterior localization of the nucleus, which is regulated by prickle, a core PCP component. The Ciona savignyi mutant aimless carries a deletion in prickle, resulting in a loss of A/P polarity manifested in the randomized localization of nucleus in addition to an earlier convergent extension defect (Jiang et al., 2005). We first explored the possibility that the posterior nucleus might influence the direction of ring movement mobilized by blebbistatin. To this end, we examined the notochord of the ascidian Halocynthia roretzi, which follows a remarkably similar early developmental process as in Ciona (Figure 3A,B), except at the cell elongation stage, the nuclei are positioned in the center of the cells (asterisks in Figure 3B). A conspicuous circumferential constriction is present at the equator, which is colocalized with a cortical ring of actin (Figure 3C) and activated myosin (Figure 3D), whose activity is essential for notochord cell elongation (Figure 3E,F). We next treated embryos with elongating notochord cells with blebbistatin for 3 hr. Similar to what was observed in Ciona, the actin ring was shifted invariably anteriorly (Figure 3G), indicating that the position of the nucleus does not influence the dynamic behavior of the ring. We thus used aimless embryos to explore the role of a compromised A/P polarity on the repositioning of the ring and its shifting upon blebbistatin treatment, independent from its influence on nuclear position. In wild-type C. savignyi embryos, elongated notochord cells possess a circumferential actin ring at the cell equator and a posterior nucleus (Figure 4A). Blebbistatin treatment shifts the ring towards the anterior pole (Figure 4B), mirroring exactly the events in C. intestinalis. In aimless embryos, the intercalation of notochord cells (outlined in Figure 4C) is impaired, except in the posterior region where the cells often align into a single file (Jiang et al., 2005). In these cells, the loss of A/P polarity is evident by the random position of the nuclei (Figure 4C insert). In 92% of mock-treated aimless cells in this region, the actin ring is positioned at the equator (Figure 4C insert), showing that equatorial ring positioning is independent from PCP. However, reducing contractility in polarity-deficient mutant cells led to a dramatically different phenotype: after 60-min blebbistatin treatment, no unidirectional migration towards the anterior was observed. The direction of the shift was randomized and independent of the localization of the nucleus: examples for anterior nucleus and ring, anterior nucleus and posterior ring, posterior nucleus and ring, and posterior nucleus and anterior ring could be found (Figure 4D,D'). In 56% of the cells, the ring was still positioned at the equator, and in the rest, the ring migrated either to the anterior or posterior side (18% and 26%) (n = 57) (Figure 4E). These results imply that A/P polarity by prickle is not necessary for the establishment of an equatorial actin ring; however, it is instrumental for the direction of movement of the ring upon blebbistatin treatment.10.7554/eLife.09206.013Figure 3.Circumferential actin rings are shifted anteriorly in Halocynthia roretzi notochord cells with centrally localized nuclei.

Bottom Line: Intriguingly, rings always form at the cells' anterior edge before migrating towards the center as contractility increases, reflecting a novel dynamical property of the cortex.We develop a simple model of the physical forces underlying this tug-of-war, which quantitatively reproduces our results.We thus propose a quantitative framework for dissecting the relative contribution of contractility and PCP to the self-assembly and repositioning of cytoskeletal structures, which should be applicable to other morphogenetic events.

View Article: PubMed Central - PubMed

Affiliation: Sars International Centre for Marine Molecular Biology, University of Bergen, Bergen, Norway.

ABSTRACT
The actomyosin cytoskeleton is a primary force-generating mechanism in morphogenesis, thus a robust spatial control of cytoskeletal positioning is essential. In this report, we demonstrate that actomyosin contractility and planar cell polarity (PCP) interact in post-mitotic Ciona notochord cells to self-assemble and reposition actomyosin rings, which play an essential role for cell elongation. Intriguingly, rings always form at the cells' anterior edge before migrating towards the center as contractility increases, reflecting a novel dynamical property of the cortex. Our drug and genetic manipulations uncover a tug-of-war between contractility, which localizes cortical flows toward the equator and PCP, which tries to reposition them. We develop a simple model of the physical forces underlying this tug-of-war, which quantitatively reproduces our results. We thus propose a quantitative framework for dissecting the relative contribution of contractility and PCP to the self-assembly and repositioning of cytoskeletal structures, which should be applicable to other morphogenetic events.

No MeSH data available.


Related in: MedlinePlus