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A Sec14p-nodulin domain phosphatidylinositol transfer protein polarizes membrane growth of Arabidopsis thaliana root hairs.

Vincent P, Chua M, Nogue F, Fairbrother A, Mekeel H, Xu Y, Allen N, Bibikova TN, Gilroy S, Bankaitis VA - J. Cell Biol. (2005)

Bottom Line: Derangement of tip-directed Ca2+ gradients is also apparent and results from isotropic influx of Ca2+ from the extracellular milieu.We propose AtSfh1p regulates intracellular and plasma membrane phosphoinositide polarity landmarks that focus membrane trafficking, Ca2+ signaling, and cytoskeleton functions to the growing root hair apex.We further suggest that Sec14p-nodulin domain proteins represent a family of regulators of polarized membrane growth in plants.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell and Developmental Biology, Michael Hooker Microscopy Facility, University of North Carolina School of Medicine, Chapel Hill, NC 27599, USA. patrick_vincent@med.unc.edu

ABSTRACT
Phosphatidylinositol (PtdIns) transfer proteins (PITPs) regulate signaling interfaces between lipid metabolism and membrane trafficking. Herein, we demonstrate that AtSfh1p, a member of a large and uncharacterized Arabidopsis thaliana Sec14p-nodulin domain family, is a PITP that regulates a specific stage in root hair development. AtSfh1p localizes along the root hair plasma membrane and is enriched in discrete plasma membrane domains and in the root hair tip cytoplasm. This localization pattern recapitulates that visualized for PtdIns(4,5)P2 in developing root hairs. Gene ablation experiments show AtSfh1p izygosity compromises polarized root hair expansion in a manner that coincides with loss of tip-directed PtdIns(4,5)P2, dispersal of secretory vesicles from the tip cytoplasm, loss of the tip f-actin network, and manifest disorganization of the root hair microtubule cytoskeleton. Derangement of tip-directed Ca2+ gradients is also apparent and results from isotropic influx of Ca2+ from the extracellular milieu. We propose AtSfh1p regulates intracellular and plasma membrane phosphoinositide polarity landmarks that focus membrane trafficking, Ca2+ signaling, and cytoskeleton functions to the growing root hair apex. We further suggest that Sec14p-nodulin domain proteins represent a family of regulators of polarized membrane growth in plants.

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Defective Ca2+ signaling in AtSfh1p-deficient root hairs. (A) Ca2+ gradients. Ca2+ concentrations in wild-type (left) and mutant (right) root hairs are in pseudocolor calibrated with the inset scale shown at right. Bar, 25 μm. (B) Typical profile of Ca2+ fluxes around the root hairs of both wild-type and Atsfh1 izygous seedlings (3-d-old), as indicated. Measurements were taken at predefined locations, and the Ca2+ selective probe was positioned 2 μm away from the root hair surface at those locations. The excursion of the probe is perpendicular to the surface. Arrows show the magnitude and direction of the Ca2+ flux at the position on the root hair surface indicated. (C) Ca2+ flux profiling of wild-type and Atsfh1 izygous seedlings (3-d-old). The percentages were obtained by dividing the raw Ca2+ flux values (ΔμV) measured at each indicated position by the values measured at position 0 (inset). The averages of Ca2+ flux values from measurements of 15 independent root hairs are shown with standard errors. Single root hairs of AtSfh1::T-DNA plants were used in these analyses.
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fig7: Defective Ca2+ signaling in AtSfh1p-deficient root hairs. (A) Ca2+ gradients. Ca2+ concentrations in wild-type (left) and mutant (right) root hairs are in pseudocolor calibrated with the inset scale shown at right. Bar, 25 μm. (B) Typical profile of Ca2+ fluxes around the root hairs of both wild-type and Atsfh1 izygous seedlings (3-d-old), as indicated. Measurements were taken at predefined locations, and the Ca2+ selective probe was positioned 2 μm away from the root hair surface at those locations. The excursion of the probe is perpendicular to the surface. Arrows show the magnitude and direction of the Ca2+ flux at the position on the root hair surface indicated. (C) Ca2+ flux profiling of wild-type and Atsfh1 izygous seedlings (3-d-old). The percentages were obtained by dividing the raw Ca2+ flux values (ΔμV) measured at each indicated position by the values measured at position 0 (inset). The averages of Ca2+ flux values from measurements of 15 independent root hairs are shown with standard errors. Single root hairs of AtSfh1::T-DNA plants were used in these analyses.

Mentions: One critical contributing cue that controls polarized root hair growth is a tip-directed Ca2+ gradient. Defects in polarized membrane trafficking are expected to randomize ion (e.g., Ca2+) channel delivery to the plasma membrane, with the consequence that spatial regulation of Ca2+ signaling will be compromised. To investigate whether or not dysregulation of Ca2+ signaling was occurring in Atsfh1::T-DNA root hairs, we used Indo-1 loading strategies to image Ca2+ signaling in wild-type and isogenic Atsfh1::T-DNA root hairs. Striking derangements in Ca2+ signaling to the growing apex of the root hair plasma membrane were observed (Fig. 7 A). Ca2+ imaging recorded a single tip-focused Ca2+ gradient in wild-type root hairs, as reported previously (Wymer et al., 1997), with tip cytoplasmic Ca2+ reaching concentrations in excess of 600 nM. Cytoplasmic Ca2+ fell to concentrations below 100 nM very rapidly away from the root hair tip. In marked contrast, precocious Ca2+ signaling was evident along the Atsfh1 root hair cortical plasma membrane with local Ca2+ concentrations reaching 600 nM or greater (Fig. 7 A).


A Sec14p-nodulin domain phosphatidylinositol transfer protein polarizes membrane growth of Arabidopsis thaliana root hairs.

Vincent P, Chua M, Nogue F, Fairbrother A, Mekeel H, Xu Y, Allen N, Bibikova TN, Gilroy S, Bankaitis VA - J. Cell Biol. (2005)

Defective Ca2+ signaling in AtSfh1p-deficient root hairs. (A) Ca2+ gradients. Ca2+ concentrations in wild-type (left) and mutant (right) root hairs are in pseudocolor calibrated with the inset scale shown at right. Bar, 25 μm. (B) Typical profile of Ca2+ fluxes around the root hairs of both wild-type and Atsfh1 izygous seedlings (3-d-old), as indicated. Measurements were taken at predefined locations, and the Ca2+ selective probe was positioned 2 μm away from the root hair surface at those locations. The excursion of the probe is perpendicular to the surface. Arrows show the magnitude and direction of the Ca2+ flux at the position on the root hair surface indicated. (C) Ca2+ flux profiling of wild-type and Atsfh1 izygous seedlings (3-d-old). The percentages were obtained by dividing the raw Ca2+ flux values (ΔμV) measured at each indicated position by the values measured at position 0 (inset). The averages of Ca2+ flux values from measurements of 15 independent root hairs are shown with standard errors. Single root hairs of AtSfh1::T-DNA plants were used in these analyses.
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Related In: Results  -  Collection

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fig7: Defective Ca2+ signaling in AtSfh1p-deficient root hairs. (A) Ca2+ gradients. Ca2+ concentrations in wild-type (left) and mutant (right) root hairs are in pseudocolor calibrated with the inset scale shown at right. Bar, 25 μm. (B) Typical profile of Ca2+ fluxes around the root hairs of both wild-type and Atsfh1 izygous seedlings (3-d-old), as indicated. Measurements were taken at predefined locations, and the Ca2+ selective probe was positioned 2 μm away from the root hair surface at those locations. The excursion of the probe is perpendicular to the surface. Arrows show the magnitude and direction of the Ca2+ flux at the position on the root hair surface indicated. (C) Ca2+ flux profiling of wild-type and Atsfh1 izygous seedlings (3-d-old). The percentages were obtained by dividing the raw Ca2+ flux values (ΔμV) measured at each indicated position by the values measured at position 0 (inset). The averages of Ca2+ flux values from measurements of 15 independent root hairs are shown with standard errors. Single root hairs of AtSfh1::T-DNA plants were used in these analyses.
Mentions: One critical contributing cue that controls polarized root hair growth is a tip-directed Ca2+ gradient. Defects in polarized membrane trafficking are expected to randomize ion (e.g., Ca2+) channel delivery to the plasma membrane, with the consequence that spatial regulation of Ca2+ signaling will be compromised. To investigate whether or not dysregulation of Ca2+ signaling was occurring in Atsfh1::T-DNA root hairs, we used Indo-1 loading strategies to image Ca2+ signaling in wild-type and isogenic Atsfh1::T-DNA root hairs. Striking derangements in Ca2+ signaling to the growing apex of the root hair plasma membrane were observed (Fig. 7 A). Ca2+ imaging recorded a single tip-focused Ca2+ gradient in wild-type root hairs, as reported previously (Wymer et al., 1997), with tip cytoplasmic Ca2+ reaching concentrations in excess of 600 nM. Cytoplasmic Ca2+ fell to concentrations below 100 nM very rapidly away from the root hair tip. In marked contrast, precocious Ca2+ signaling was evident along the Atsfh1 root hair cortical plasma membrane with local Ca2+ concentrations reaching 600 nM or greater (Fig. 7 A).

Bottom Line: Derangement of tip-directed Ca2+ gradients is also apparent and results from isotropic influx of Ca2+ from the extracellular milieu.We propose AtSfh1p regulates intracellular and plasma membrane phosphoinositide polarity landmarks that focus membrane trafficking, Ca2+ signaling, and cytoskeleton functions to the growing root hair apex.We further suggest that Sec14p-nodulin domain proteins represent a family of regulators of polarized membrane growth in plants.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell and Developmental Biology, Michael Hooker Microscopy Facility, University of North Carolina School of Medicine, Chapel Hill, NC 27599, USA. patrick_vincent@med.unc.edu

ABSTRACT
Phosphatidylinositol (PtdIns) transfer proteins (PITPs) regulate signaling interfaces between lipid metabolism and membrane trafficking. Herein, we demonstrate that AtSfh1p, a member of a large and uncharacterized Arabidopsis thaliana Sec14p-nodulin domain family, is a PITP that regulates a specific stage in root hair development. AtSfh1p localizes along the root hair plasma membrane and is enriched in discrete plasma membrane domains and in the root hair tip cytoplasm. This localization pattern recapitulates that visualized for PtdIns(4,5)P2 in developing root hairs. Gene ablation experiments show AtSfh1p izygosity compromises polarized root hair expansion in a manner that coincides with loss of tip-directed PtdIns(4,5)P2, dispersal of secretory vesicles from the tip cytoplasm, loss of the tip f-actin network, and manifest disorganization of the root hair microtubule cytoskeleton. Derangement of tip-directed Ca2+ gradients is also apparent and results from isotropic influx of Ca2+ from the extracellular milieu. We propose AtSfh1p regulates intracellular and plasma membrane phosphoinositide polarity landmarks that focus membrane trafficking, Ca2+ signaling, and cytoskeleton functions to the growing root hair apex. We further suggest that Sec14p-nodulin domain proteins represent a family of regulators of polarized membrane growth in plants.

Show MeSH
Related in: MedlinePlus