title: Exocyst SEC3 and Phosphoinositides Define Sites of Exocytosis in Pollen Tube Initiation and Growth.
abstract: Polarized exocytosis is critical for pollen tube growth, but its localization and function are still under debate. The exocyst vesicle-tethering complex functions in polarized exocytosis. Here, we show that a sec3a exocyst subunit null mutant cannot be transmitted through the male gametophyte due to a defect in pollen tube growth. The green fluorescent protein (GFP)-SEC3a fusion protein is functional and accumulates at or proximal to the pollen tube tip plasma membrane. Partial complementation of sec3a resulted in the development of pollen with multiple tips, indicating that SEC3 is required to determine the site of pollen germination pore formation. Time-lapse imaging demonstrated that SEC3a and SEC8 were highly dynamic and that SEC3a localization on the apical plasma membrane predicts the direction of growth. At the tip, polar SEC3a domains coincided with cell wall deposition. Labeling of GFP-SEC3a-expressing pollen with the endocytic marker FM4-64 revealed the presence of subdomains on the apical membrane characterized by extensive exocytosis. In steady-state growing tobacco (Nicotiana tabacum) pollen tubes, SEC3a displayed amino-terminal Pleckstrin homology-like domain (SEC3a-N)-dependent subapical membrane localization. In agreement, SEC3a-N interacted with phosphoinositides in vitro and colocalized with a phosphatidylinositol 4,5-bisphosphate (PIP2) marker in pollen tubes. Correspondingly, molecular dynamics simulations indicated that SEC3a-N associates with the membrane by interacting with PIP2However, the interaction with PIP2is not required for polar localization and the function of SEC3a in Arabidopsis (Arabidopsis thaliana). Taken together, our findings indicate that SEC3a is a critical determinant of polar exocytosis during tip growth and suggest differential regulation of the exocytotic machinery depending on pollen tube growth modes.
authors: Department of Molecular Biology and Ecology of Plants, Tel Aviv University, Tel Aviv 69978, Israel (D.B., H.S., S.Y.);Institute of Experimental Botany, Czech Academy of Sciences, 165 02 Prague, Czech Republic (R.P., P.P., M.P., P.T., N.V., V.Ž.);J. Heyrovsky Institute of Physical Chemistry, Czech Academy of Sciences, 182 23 Prague, Czech Republic (L.C.); andDepartment of Experimental Plant Biology, Faculty of Science, Charles University, 128 44 Prague, Czech Republic (N.V., V.Ž.).
authors: Daria
authors: D
authors: Bloch
authors: Department of Molecular Biology and Ecology of Plants, Tel Aviv University, Tel Aviv 69978, Israel (D.B., H.S., S.Y.);Institute of Experimental Botany, Czech Academy of Sciences, 165 02 Prague, Czech Republic (R.P., P.P., M.P., P.T., N.V., V.Ž.);J. Heyrovsky Institute of Physical Chemistry, Czech Academy of Sciences, 182 23 Prague, Czech Republic (L.C.); andDepartment of Experimental Plant Biology, Faculty of Science, Charles University, 128 44 Prague, Czech Republic (N.V., V.Ž.).
authors: Roman
authors: R
authors: Pleskot
authors: Department of Molecular Biology and Ecology of Plants, Tel Aviv University, Tel Aviv 69978, Israel (D.B., H.S., S.Y.);Institute of Experimental Botany, Czech Academy of Sciences, 165 02 Prague, Czech Republic (R.P., P.P., M.P., P.T., N.V., V.Ž.);J. Heyrovsky Institute of Physical Chemistry, Czech Academy of Sciences, 182 23 Prague, Czech Republic (L.C.); andDepartment of Experimental Plant Biology, Faculty of Science, Charles University, 128 44 Prague, Czech Republic (N.V., V.Ž.).
authors: Přemysl
authors: 0000-0003-0488-7465
authors: P
authors: Pejchar
authors: Department of Molecular Biology and Ecology of Plants, Tel Aviv University, Tel Aviv 69978, Israel (D.B., H.S., S.Y.);Institute of Experimental Botany, Czech Academy of Sciences, 165 02 Prague, Czech Republic (R.P., P.P., M.P., P.T., N.V., V.Ž.);J. Heyrovsky Institute of Physical Chemistry, Czech Academy of Sciences, 182 23 Prague, Czech Republic (L.C.); andDepartment of Experimental Plant Biology, Faculty of Science, Charles University, 128 44 Prague, Czech Republic (N.V., V.Ž.).
authors: Martin
authors: 0000-0002-3699-7549
authors: M
authors: Potocký
authors: Department of Molecular Biology and Ecology of Plants, Tel Aviv University, Tel Aviv 69978, Israel (D.B., H.S., S.Y.);Institute of Experimental Botany, Czech Academy of Sciences, 165 02 Prague, Czech Republic (R.P., P.P., M.P., P.T., N.V., V.Ž.);J. Heyrovsky Institute of Physical Chemistry, Czech Academy of Sciences, 182 23 Prague, Czech Republic (L.C.); andDepartment of Experimental Plant Biology, Faculty of Science, Charles University, 128 44 Prague, Czech Republic (N.V., V.Ž.).
authors: Pavlína
authors: 0000-0001-8523-0099
authors: P
authors: Trpkošová
authors: Department of Molecular Biology and Ecology of Plants, Tel Aviv University, Tel Aviv 69978, Israel (D.B., H.S., S.Y.);Institute of Experimental Botany, Czech Academy of Sciences, 165 02 Prague, Czech Republic (R.P., P.P., M.P., P.T., N.V., V.Ž.);J. Heyrovsky Institute of Physical Chemistry, Czech Academy of Sciences, 182 23 Prague, Czech Republic (L.C.); andDepartment of Experimental Plant Biology, Faculty of Science, Charles University, 128 44 Prague, Czech Republic (N.V., V.Ž.).
authors: Lukasz
authors: 0000-0002-2083-8738
authors: L
authors: Cwiklik
authors: Department of Molecular Biology and Ecology of Plants, Tel Aviv University, Tel Aviv 69978, Israel (D.B., H.S., S.Y.);Institute of Experimental Botany, Czech Academy of Sciences, 165 02 Prague, Czech Republic (R.P., P.P., M.P., P.T., N.V., V.Ž.);J. Heyrovsky Institute of Physical Chemistry, Czech Academy of Sciences, 182 23 Prague, Czech Republic (L.C.); andDepartment of Experimental Plant Biology, Faculty of Science, Charles University, 128 44 Prague, Czech Republic (N.V., V.Ž.).
authors: Nemanja
authors: N
authors: Vukašinović
authors: Department of Molecular Biology and Ecology of Plants, Tel Aviv University, Tel Aviv 69978, Israel (D.B., H.S., S.Y.);Institute of Experimental Botany, Czech Academy of Sciences, 165 02 Prague, Czech Republic (R.P., P.P., M.P., P.T., N.V., V.Ž.);J. Heyrovsky Institute of Physical Chemistry, Czech Academy of Sciences, 182 23 Prague, Czech Republic (L.C.); andDepartment of Experimental Plant Biology, Faculty of Science, Charles University, 128 44 Prague, Czech Republic (N.V., V.Ž.).
authors: Hasana
authors: 0000-0002-5195-9487
authors: H
authors: Sternberg
authors: Department of Molecular Biology and Ecology of Plants, Tel Aviv University, Tel Aviv 69978, Israel (D.B., H.S., S.Y.);Institute of Experimental Botany, Czech Academy of Sciences, 165 02 Prague, Czech Republic (R.P., P.P., M.P., P.T., N.V., V.Ž.);J. Heyrovsky Institute of Physical Chemistry, Czech Academy of Sciences, 182 23 Prague, Czech Republic (L.C.); andDepartment of Experimental Plant Biology, Faculty of Science, Charles University, 128 44 Prague, Czech Republic (N.V., V.Ž.) shauly@tauex.tau.ac.il zarsky@ueb.cas.cz.
authors: Shaul
authors: 0000-0003-3264-0005
authors: S
authors: Yalovsky
authors: Department of Molecular Biology and Ecology of Plants, Tel Aviv University, Tel Aviv 69978, Israel (D.B., H.S., S.Y.);Institute of Experimental Botany, Czech Academy of Sciences, 165 02 Prague, Czech Republic (R.P., P.P., M.P., P.T., N.V., V.Ž.);J. Heyrovsky Institute of Physical Chemistry, Czech Academy of Sciences, 182 23 Prague, Czech Republic (L.C.); andDepartment of Experimental Plant Biology, Faculty of Science, Charles University, 128 44 Prague, Czech Republic (N.V., V.Ž.) shauly@tauex.tau.ac.il zarsky@ueb.cas.cz.
authors: Viktor
authors: 0000-0002-5301-0339
authors: V
authors: Žárský
mesh_terms: D000595:Amino Acid Sequence; D017360:Arabidopsis / Q000235:genetics / Q000254:growth & development / Q000378:metabolism; D029681:Arabidopsis Proteins / Q000145:classification / Q000235:genetics / Q000378:metabolism*; D001483:Base Sequence; D001665:Binding Sites / Q000235:genetics; D002462:Cell Membrane / Q000378:metabolism; D005089:Exocytosis; D020869:Gene Expression Profiling / Q000379:methods; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D018613:Microscopy, Confocal; D056004:Molecular Dynamics Simulation; D009154:Mutation; D019269:Phosphatidylinositol 4,5-Diphosphate / Q000378:metabolism; D010716:Phosphatidylinositols / Q000378:metabolism*; D010802:Phylogeny; D030821:Plants, Genetically Modified; D011058:Pollen / Q000235:genetics / Q000254:growth & development / Q000378:metabolism; D053205:Pollen Tube / Q000235:genetics / Q000254:growth & development / Q000378:metabolism*; D011485:Protein Binding; D020033:Protein Isoforms / Q000235:genetics / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017386:Sequence Homology, Amino Acid; D012689:Sequence Homology, Nucleic Acid; D059008:Time-Lapse Imaging / Q000379:methods; D033921:Vesicular Transport Proteins / Q000145:classification / Q000235:genetics / Q000378:metabolism*
chemical_list: D029681:Arabidopsis Proteins; D019269:Phosphatidylinositol 4,5-Diphosphate; D010716:Phosphatidylinositols; D020033:Protein Isoforms; D033921:Vesicular Transport Proteins; C000623166:exocyst complex component sec3A, Arabidopsis; D049452:Green Fluorescent Proteins