Limits...
Rop GTPase-dependent dynamics of tip-localized F-actin controls tip growth in pollen tubes.

Fu Y, Wu G, Yang Z - J. Cell Biol. (2001)

Bottom Line: Moreover, overexpression of RopGAP1 alone disrupted SABs and inhibited growth.Finally, SAB oscillates and appears at the tip before growth.Together, these results indicate that the dynamics of tip actin are essential for tip growth and provide the first direct evidence to link Rho GTPase to actin organization in controlling cell polarity and polar growth in plants.

View Article: PubMed Central - PubMed

Affiliation: Department of Botany and Plant Sciences, University of California, Riverside, California 92521, USA.

ABSTRACT
Tip-growing pollen tubes provide a useful model system to study polar growth. Although roles for tip-focused calcium gradient and tip-localized Rho-family GTPase in pollen tube growth is established, the existence and function of tip-localized F-actin have been controversial. Using the green fluorescent protein-tagged actin-binding domain of mouse talin, we found a dynamic form of tip-localized F-actin in tobacco pollen tubes, termed short actin bundles (SABs). The dynamics of SABs during polar growth in pollen tubes is regulated by Rop1At, a Rop GTPase belonging to the Rho family. When overexpressed, Rop1At transformed SAB into a network of fine filaments and induced a transverse actin band behind the tip, leading to depolarized growth. These changes were due to ectopic Rop1At localization to the apical region of the plasma membrane and were suppressed by guanine dissociation inhibitor overexpression, which removed ectopically localized Rop1At. Rop GTPase-activating protein (RopGAP1) overexpression, or Latrunculin B treatments, also recovered normal actin organization and tip growth in Rop1At-overexpressing tubes. Moreover, overexpression of RopGAP1 alone disrupted SABs and inhibited growth. Finally, SAB oscillates and appears at the tip before growth. Together, these results indicate that the dynamics of tip actin are essential for tip growth and provide the first direct evidence to link Rho GTPase to actin organization in controlling cell polarity and polar growth in plants.

Show MeSH

Related in: MedlinePlus

DN-rop1At overexpression disrupts SAB and induces a transverse actin band behind the tip. Tobacco pollen grains were cobombarded with LAT52::GFP-mTalin and LAT52::DN-rop1At constructs (A and B) or bombarded with LAT52::GFP-mTalin alone before treatment with 5 nM Latrunculin B (C) or 300 nM caffeine (D). GFP-mTalin was analyzed by confocal microscopy as described in the legend to Fig. 1. All images shown are projections of 1-μm confocal sections. Arrow, transverse actin bands behind the apex; asterisk, F-actin free apex; diamond, actin cables. Bar, 10 μm.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC2198818&req=5

Figure 2: DN-rop1At overexpression disrupts SAB and induces a transverse actin band behind the tip. Tobacco pollen grains were cobombarded with LAT52::GFP-mTalin and LAT52::DN-rop1At constructs (A and B) or bombarded with LAT52::GFP-mTalin alone before treatment with 5 nM Latrunculin B (C) or 300 nM caffeine (D). GFP-mTalin was analyzed by confocal microscopy as described in the legend to Fig. 1. All images shown are projections of 1-μm confocal sections. Arrow, transverse actin bands behind the apex; asterisk, F-actin free apex; diamond, actin cables. Bar, 10 μm.

Mentions: To assess the involvement of Rop in the regulation of tip F-actin, we coexpressed GFP-mTalin with a DN-rop1At mutant in tobacco pollen tubes, which has been shown to inhibit tube growth and cause slight tube expansion when overexpressed in Arabidopsis pollen (Li et al. 1999). DN-rop1At transient expression caused similar phenotypes in tobacco pollen tubes (Fig. 2A and Fig. B). Importantly, DN-rop1At expression also disrupted the formation of apical SAB, suggesting Rop may control SAB formation. Nearly 60% of DN-rop1At tubes contained no SAB, and ∼32% of them had drastically reduced SAB levels; only 8% of pollen tubes were seen with a similar amount of apical actin as wild-type tubes. We did not observe significant changes in axial actin cables in DN-rop1At tubes (Fig. 2 B). Surprisingly, we also observed a transverse actin band composed of cortical actin hoops just behind the apical dome in DN-rop1At cells. Time–sequence analyses showed that the transverse actin band was quite stable and actually widened over time, apparently due to actin polymerization behind the apical dome (data not shown). The DN-rop1At–induced transverse actin band could be caused by DN-rop1At–induced growth inhibition, direct effects of DN-rop1At, or indirect effects of DN-rop1At that result in increased Rop activity behind the tip, because DN-rop1At overexpression causes ectopic localization of wild-type Rop proteins in the subapical region of the PM (Wu, G., Y. Fu, and Z. Yang, unpublished data).


Rop GTPase-dependent dynamics of tip-localized F-actin controls tip growth in pollen tubes.

Fu Y, Wu G, Yang Z - J. Cell Biol. (2001)

DN-rop1At overexpression disrupts SAB and induces a transverse actin band behind the tip. Tobacco pollen grains were cobombarded with LAT52::GFP-mTalin and LAT52::DN-rop1At constructs (A and B) or bombarded with LAT52::GFP-mTalin alone before treatment with 5 nM Latrunculin B (C) or 300 nM caffeine (D). GFP-mTalin was analyzed by confocal microscopy as described in the legend to Fig. 1. All images shown are projections of 1-μm confocal sections. Arrow, transverse actin bands behind the apex; asterisk, F-actin free apex; diamond, actin cables. Bar, 10 μm.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2198818&req=5

Figure 2: DN-rop1At overexpression disrupts SAB and induces a transverse actin band behind the tip. Tobacco pollen grains were cobombarded with LAT52::GFP-mTalin and LAT52::DN-rop1At constructs (A and B) or bombarded with LAT52::GFP-mTalin alone before treatment with 5 nM Latrunculin B (C) or 300 nM caffeine (D). GFP-mTalin was analyzed by confocal microscopy as described in the legend to Fig. 1. All images shown are projections of 1-μm confocal sections. Arrow, transverse actin bands behind the apex; asterisk, F-actin free apex; diamond, actin cables. Bar, 10 μm.
Mentions: To assess the involvement of Rop in the regulation of tip F-actin, we coexpressed GFP-mTalin with a DN-rop1At mutant in tobacco pollen tubes, which has been shown to inhibit tube growth and cause slight tube expansion when overexpressed in Arabidopsis pollen (Li et al. 1999). DN-rop1At transient expression caused similar phenotypes in tobacco pollen tubes (Fig. 2A and Fig. B). Importantly, DN-rop1At expression also disrupted the formation of apical SAB, suggesting Rop may control SAB formation. Nearly 60% of DN-rop1At tubes contained no SAB, and ∼32% of them had drastically reduced SAB levels; only 8% of pollen tubes were seen with a similar amount of apical actin as wild-type tubes. We did not observe significant changes in axial actin cables in DN-rop1At tubes (Fig. 2 B). Surprisingly, we also observed a transverse actin band composed of cortical actin hoops just behind the apical dome in DN-rop1At cells. Time–sequence analyses showed that the transverse actin band was quite stable and actually widened over time, apparently due to actin polymerization behind the apical dome (data not shown). The DN-rop1At–induced transverse actin band could be caused by DN-rop1At–induced growth inhibition, direct effects of DN-rop1At, or indirect effects of DN-rop1At that result in increased Rop activity behind the tip, because DN-rop1At overexpression causes ectopic localization of wild-type Rop proteins in the subapical region of the PM (Wu, G., Y. Fu, and Z. Yang, unpublished data).

Bottom Line: Moreover, overexpression of RopGAP1 alone disrupted SABs and inhibited growth.Finally, SAB oscillates and appears at the tip before growth.Together, these results indicate that the dynamics of tip actin are essential for tip growth and provide the first direct evidence to link Rho GTPase to actin organization in controlling cell polarity and polar growth in plants.

View Article: PubMed Central - PubMed

Affiliation: Department of Botany and Plant Sciences, University of California, Riverside, California 92521, USA.

ABSTRACT
Tip-growing pollen tubes provide a useful model system to study polar growth. Although roles for tip-focused calcium gradient and tip-localized Rho-family GTPase in pollen tube growth is established, the existence and function of tip-localized F-actin have been controversial. Using the green fluorescent protein-tagged actin-binding domain of mouse talin, we found a dynamic form of tip-localized F-actin in tobacco pollen tubes, termed short actin bundles (SABs). The dynamics of SABs during polar growth in pollen tubes is regulated by Rop1At, a Rop GTPase belonging to the Rho family. When overexpressed, Rop1At transformed SAB into a network of fine filaments and induced a transverse actin band behind the tip, leading to depolarized growth. These changes were due to ectopic Rop1At localization to the apical region of the plasma membrane and were suppressed by guanine dissociation inhibitor overexpression, which removed ectopically localized Rop1At. Rop GTPase-activating protein (RopGAP1) overexpression, or Latrunculin B treatments, also recovered normal actin organization and tip growth in Rop1At-overexpressing tubes. Moreover, overexpression of RopGAP1 alone disrupted SABs and inhibited growth. Finally, SAB oscillates and appears at the tip before growth. Together, these results indicate that the dynamics of tip actin are essential for tip growth and provide the first direct evidence to link Rho GTPase to actin organization in controlling cell polarity and polar growth in plants.

Show MeSH
Related in: MedlinePlus