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Visualization of PtdIns3P dynamics in living plant cells.
Plant J. 2006 Sep; 47(5):687-700.PJ

Abstract

To investigate PtdIns3P localization and function in plants, a fluorescent PtdIns3P-specific biosensor (YFP-2xFYVE) was created. On lipid dot blots it bound specifically and with high affinity to PtdIns3P. Transient expression in cowpea protoplasts labelled vacuolar membranes and highly motile structures undergoing fusion and fission. Stable expression in tobacco BY-2 cells labelled similar motile structures, but labelled vacuolar membranes hardly at all. YFP-2xFYVE fluorescence strongly co-localized with the pre-vacuolar marker AtRABF2b, partially co-localized with the endosomal tracer FM4-64, but showed no overlap with the Golgi marker STtmd-CFP. Treatment of cells with wortmannin, a PI3 kinase inhibitor, caused the YFP-2xFYVE fluorescence to redistribute into the cytosol and nucleus within 15 min. BY-2 cells expressing YFP-2xFYVE contained twice as much PtdIns3P as YFP-transformed cells, but this had no effect on cell-growth or stress-induced phospholipid signalling responses. Upon treatment with wortmannin, PtdIns3P levels were reduced by approximately 40% within 15 min in both cell lines. Stable expression of YFP-2xFYVE in Arabidopsis plants labelled different subcellular structures in root compared with shoot tissues. In addition labelling the motile structures common to all cells, YFP-2xFYVE strongly labelled the vacuolar membrane in leaf epidermal and guard cells, suggesting that cell differentiation alters the distribution of PtdIns3P. In dividing BY-2 cells, YFP-2xFYVE-labelled vesicles surrounded the newly formed cell plate, suggesting a role for PtdIns3P in cytokinesis. Together, these data show that YFP-2xFYVE may be used as a biosensor to specifically visualize PtdIns3P in living plant cells.

Authors+Show Affiliations

Section of Molecular Cytology, Swammerdam Institute for Life Sciences, University of Amsterdam, Kruislaan 316, Amsterdam, The Netherlands.No affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info available

Pub Type(s)

Journal Article
Research Support, Non-U.S. Gov't

Language

eng

PubMed ID

16856980

Citation

Vermeer, Joop E M., et al. "Visualization of PtdIns3P Dynamics in Living Plant Cells." The Plant Journal : for Cell and Molecular Biology, vol. 47, no. 5, 2006, pp. 687-700.
Vermeer JE, van Leeuwen W, Tobeña-Santamaria R, et al. Visualization of PtdIns3P dynamics in living plant cells. Plant J. 2006;47(5):687-700.
Vermeer, J. E., van Leeuwen, W., Tobeña-Santamaria, R., Laxalt, A. M., Jones, D. R., Divecha, N., Gadella, T. W., & Munnik, T. (2006). Visualization of PtdIns3P dynamics in living plant cells. The Plant Journal : for Cell and Molecular Biology, 47(5), 687-700.
Vermeer JE, et al. Visualization of PtdIns3P Dynamics in Living Plant Cells. Plant J. 2006;47(5):687-700. PubMed PMID: 16856980.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Visualization of PtdIns3P dynamics in living plant cells. AU - Vermeer,Joop E M, AU - van Leeuwen,Wessel, AU - Tobeña-Santamaria,Rafa, AU - Laxalt,Ana M, AU - Jones,David R, AU - Divecha,Nullin, AU - Gadella,Theodorus W J,Jr AU - Munnik,Teun, Y1 - 2006/07/19/ PY - 2006/7/22/pubmed PY - 2006/11/14/medline PY - 2006/7/22/entrez SP - 687 EP - 700 JF - The Plant journal : for cell and molecular biology JO - Plant J. VL - 47 IS - 5 N2 - To investigate PtdIns3P localization and function in plants, a fluorescent PtdIns3P-specific biosensor (YFP-2xFYVE) was created. On lipid dot blots it bound specifically and with high affinity to PtdIns3P. Transient expression in cowpea protoplasts labelled vacuolar membranes and highly motile structures undergoing fusion and fission. Stable expression in tobacco BY-2 cells labelled similar motile structures, but labelled vacuolar membranes hardly at all. YFP-2xFYVE fluorescence strongly co-localized with the pre-vacuolar marker AtRABF2b, partially co-localized with the endosomal tracer FM4-64, but showed no overlap with the Golgi marker STtmd-CFP. Treatment of cells with wortmannin, a PI3 kinase inhibitor, caused the YFP-2xFYVE fluorescence to redistribute into the cytosol and nucleus within 15 min. BY-2 cells expressing YFP-2xFYVE contained twice as much PtdIns3P as YFP-transformed cells, but this had no effect on cell-growth or stress-induced phospholipid signalling responses. Upon treatment with wortmannin, PtdIns3P levels were reduced by approximately 40% within 15 min in both cell lines. Stable expression of YFP-2xFYVE in Arabidopsis plants labelled different subcellular structures in root compared with shoot tissues. In addition labelling the motile structures common to all cells, YFP-2xFYVE strongly labelled the vacuolar membrane in leaf epidermal and guard cells, suggesting that cell differentiation alters the distribution of PtdIns3P. In dividing BY-2 cells, YFP-2xFYVE-labelled vesicles surrounded the newly formed cell plate, suggesting a role for PtdIns3P in cytokinesis. Together, these data show that YFP-2xFYVE may be used as a biosensor to specifically visualize PtdIns3P in living plant cells. SN - 0960-7412 UR - https://www.unboundmedicine.com/medline/citation/16856980/Visualization_of_PtdIns3P_dynamics_in_living_plant_cells_ L2 - https://doi.org/10.1111/j.1365-313X.2006.02830.x DB - PRIME DP - Unbound Medicine ER -