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Dynamic F-actin movement is essential for fertilization in Arabidopsis thaliana.

Kawashima T, Maruyama D, Shagirov M, Li J, Hamamura Y, Yelagandula R, Toyama Y, Berger F - Elife (2014)

Bottom Line: Live imaging shows that F-actin structures assist the male nucleus during its migration towards the female nucleus.We identify a female gamete-specific Rho-GTPase that regulates F-actin dynamics and further show that actin-myosin interactions are also involved in male gamete nucleus migration.The innovation of a novel actin-based mechanism of fertilization during plant evolution might account for the complete loss of the centrosome in flowering plants.

View Article: PubMed Central - PubMed

Affiliation: Temasek Life Sciences Laboratory, National University of Singapore, Singapore, Singapore.

ABSTRACT
In animals, microtubules and centrosomes direct the migration of gamete pronuclei for fertilization. By contrast, flowering plants have lost essential components of the centrosome, raising the question of how flowering plants control gamete nuclei migration during fertilization. Here, we use Arabidopsis thaliana to document a novel mechanism that regulates F-actin dynamics in the female gametes and is essential for fertilization. Live imaging shows that F-actin structures assist the male nucleus during its migration towards the female nucleus. We identify a female gamete-specific Rho-GTPase that regulates F-actin dynamics and further show that actin-myosin interactions are also involved in male gamete nucleus migration. Genetic analyses and imaging indicate that microtubules are dispensable for migration and fusion of male and female gamete nuclei. The innovation of a novel actin-based mechanism of fertilization during plant evolution might account for the complete loss of the centrosome in flowering plants.

No MeSH data available.


Controls of F-actin dynamics in the central cell.(A) A scheme of the mature Arabidopsis ovule. F-actin (light green cables) velocities of the entire central cell and around the nucleus (dashed box) were analyzed. ccn, central cell nucleus; cz, chalaza; ec/sy, egg/synergid cells complex; mp, micropyle. (B) F-actin velocity around the central cell nucleus. Inward and outward represent F-actin movement direction towards and away from the nucleus, respectively. a.u., arbitral unit. (C) The representative probability distributions of velocities integrated over the entire central cell area and time. Solid lines represent the fitted gamma distribution. (D) The mean velocities of central cell F-actin movement. Error bars represent the standard deviation of five to seven biological replicates. n.s., not significant; ***, p < 0.001; **, p < 0.01, student t-test. (E) Stacks of central cell F-actin time-lapse. F-actin movements are marked by rainbow colors while white color results from overlapping multiple colors, indicating less movement. Scale bar = 10 µm.DOI:http://dx.doi.org/10.7554/eLife.04501.015
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fig6: Controls of F-actin dynamics in the central cell.(A) A scheme of the mature Arabidopsis ovule. F-actin (light green cables) velocities of the entire central cell and around the nucleus (dashed box) were analyzed. ccn, central cell nucleus; cz, chalaza; ec/sy, egg/synergid cells complex; mp, micropyle. (B) F-actin velocity around the central cell nucleus. Inward and outward represent F-actin movement direction towards and away from the nucleus, respectively. a.u., arbitral unit. (C) The representative probability distributions of velocities integrated over the entire central cell area and time. Solid lines represent the fitted gamma distribution. (D) The mean velocities of central cell F-actin movement. Error bars represent the standard deviation of five to seven biological replicates. n.s., not significant; ***, p < 0.001; **, p < 0.01, student t-test. (E) Stacks of central cell F-actin time-lapse. F-actin movements are marked by rainbow colors while white color results from overlapping multiple colors, indicating less movement. Scale bar = 10 µm.DOI:http://dx.doi.org/10.7554/eLife.04501.015

Mentions: We measured the dynamics of F-actin around the central cell nucleus, where the sperm cell nucleus migration occurs, and observed a predominant F-actin movement from the cell periphery towards the nucleus (Video 3), hereafter referred to as inward movement (Figure 6A,B). By contrast, F-actin movement was extremely reduced in central cells expressing DN-ROP8 (Figure 6C–E; Videos 3; Video 4; Video 5; 45% and 30% defects in DN-ROP8 Line 1 [n = 11] and Line 2 [n = 10], respectively, compared to 0% defect in WT [n = 7] and CA-ROP8 Line 1 [n = 5] and Line 2 [n = 6]). Taken together, these results show that ROP8 controls novel constant inward dynamics of F-actin movement in the central cell and that this ROP8-dependent inward F-actin movement is required for the proper transport of the sperm cell nucleus to the central cell nucleus. Although F-actin flows towards the central cell nucleus, we did not detect accumulation of F-actin around the nucleus (Video 3), suggesting that F-actin disassembly takes place around the central cell nucleus.Video 3.Time-lapse image of F-actin dynamics in the mature WT central cell.


Dynamic F-actin movement is essential for fertilization in Arabidopsis thaliana.

Kawashima T, Maruyama D, Shagirov M, Li J, Hamamura Y, Yelagandula R, Toyama Y, Berger F - Elife (2014)

Controls of F-actin dynamics in the central cell.(A) A scheme of the mature Arabidopsis ovule. F-actin (light green cables) velocities of the entire central cell and around the nucleus (dashed box) were analyzed. ccn, central cell nucleus; cz, chalaza; ec/sy, egg/synergid cells complex; mp, micropyle. (B) F-actin velocity around the central cell nucleus. Inward and outward represent F-actin movement direction towards and away from the nucleus, respectively. a.u., arbitral unit. (C) The representative probability distributions of velocities integrated over the entire central cell area and time. Solid lines represent the fitted gamma distribution. (D) The mean velocities of central cell F-actin movement. Error bars represent the standard deviation of five to seven biological replicates. n.s., not significant; ***, p < 0.001; **, p < 0.01, student t-test. (E) Stacks of central cell F-actin time-lapse. F-actin movements are marked by rainbow colors while white color results from overlapping multiple colors, indicating less movement. Scale bar = 10 µm.DOI:http://dx.doi.org/10.7554/eLife.04501.015
© Copyright Policy - open-access
Related In: Results  -  Collection

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fig6: Controls of F-actin dynamics in the central cell.(A) A scheme of the mature Arabidopsis ovule. F-actin (light green cables) velocities of the entire central cell and around the nucleus (dashed box) were analyzed. ccn, central cell nucleus; cz, chalaza; ec/sy, egg/synergid cells complex; mp, micropyle. (B) F-actin velocity around the central cell nucleus. Inward and outward represent F-actin movement direction towards and away from the nucleus, respectively. a.u., arbitral unit. (C) The representative probability distributions of velocities integrated over the entire central cell area and time. Solid lines represent the fitted gamma distribution. (D) The mean velocities of central cell F-actin movement. Error bars represent the standard deviation of five to seven biological replicates. n.s., not significant; ***, p < 0.001; **, p < 0.01, student t-test. (E) Stacks of central cell F-actin time-lapse. F-actin movements are marked by rainbow colors while white color results from overlapping multiple colors, indicating less movement. Scale bar = 10 µm.DOI:http://dx.doi.org/10.7554/eLife.04501.015
Mentions: We measured the dynamics of F-actin around the central cell nucleus, where the sperm cell nucleus migration occurs, and observed a predominant F-actin movement from the cell periphery towards the nucleus (Video 3), hereafter referred to as inward movement (Figure 6A,B). By contrast, F-actin movement was extremely reduced in central cells expressing DN-ROP8 (Figure 6C–E; Videos 3; Video 4; Video 5; 45% and 30% defects in DN-ROP8 Line 1 [n = 11] and Line 2 [n = 10], respectively, compared to 0% defect in WT [n = 7] and CA-ROP8 Line 1 [n = 5] and Line 2 [n = 6]). Taken together, these results show that ROP8 controls novel constant inward dynamics of F-actin movement in the central cell and that this ROP8-dependent inward F-actin movement is required for the proper transport of the sperm cell nucleus to the central cell nucleus. Although F-actin flows towards the central cell nucleus, we did not detect accumulation of F-actin around the nucleus (Video 3), suggesting that F-actin disassembly takes place around the central cell nucleus.Video 3.Time-lapse image of F-actin dynamics in the mature WT central cell.

Bottom Line: Live imaging shows that F-actin structures assist the male nucleus during its migration towards the female nucleus.We identify a female gamete-specific Rho-GTPase that regulates F-actin dynamics and further show that actin-myosin interactions are also involved in male gamete nucleus migration.The innovation of a novel actin-based mechanism of fertilization during plant evolution might account for the complete loss of the centrosome in flowering plants.

View Article: PubMed Central - PubMed

Affiliation: Temasek Life Sciences Laboratory, National University of Singapore, Singapore, Singapore.

ABSTRACT
In animals, microtubules and centrosomes direct the migration of gamete pronuclei for fertilization. By contrast, flowering plants have lost essential components of the centrosome, raising the question of how flowering plants control gamete nuclei migration during fertilization. Here, we use Arabidopsis thaliana to document a novel mechanism that regulates F-actin dynamics in the female gametes and is essential for fertilization. Live imaging shows that F-actin structures assist the male nucleus during its migration towards the female nucleus. We identify a female gamete-specific Rho-GTPase that regulates F-actin dynamics and further show that actin-myosin interactions are also involved in male gamete nucleus migration. Genetic analyses and imaging indicate that microtubules are dispensable for migration and fusion of male and female gamete nuclei. The innovation of a novel actin-based mechanism of fertilization during plant evolution might account for the complete loss of the centrosome in flowering plants.

No MeSH data available.