Exploring the phosphoinositide network to modulate plant cell polarity
Final Report Abstract
Based on previous results, the roles of phosphoinositides in the establishment of cell polarity were studied in Arabidopsis. Focus of this work was on the production of the multifunctional signaling lipid, phosphatidylinositol-4,5-bisphosphate (PtdIns(4,5)P2) by PI4P 5-kinases. One important aspect of previous work was the role of PtdIns(4,5)P2-formation by different PI4P 5-kinase isoforms in the control of pollen tube growth. We had previously demonstrated that the type B PI4P5-kinases, PIP5K4 and PIP5K5, exerted relevant effects on apical pectin secretion in pollen tubes. It was an important goal of the last funding period to complete our work on the roles of type A PI4P5-kinases in pollen tubes. To this end, the data obtained indicate that type A enzymes have a different role in pollen tube growth and are important in controlling the actin cytoskeleton. Considering that PtdIns(4,5)P2 generated by PI4P5-kinases of the types A and B exerted such different functions, another important aspect of our work was aimed at defining possible channeling of phosphoinositide intermediates between enzymes towards one or the other effect. A systematic coexpression study was initiated, in which various enzymes of the phosphoinositide pathway were tested for synergistic effects on pollen tube growth. Enzymes tested included PI-synthases (PIS), PI-kinases (PI4K) and PI4P 5-kinases. The data indicate that a pathway defined by the enzymes, PIS2, PI4Kβ1 and PIP5K5 is involved in the comntrol of apical pectin secretion in pollen tubes. These results were also published. While much of our previous work had been directed towards understanding the workings of single cells exhibiting polar tip growth, such as root hairs or pollen tubes, another aspect of our work was aimed at understanding polarity at the tissue level in the vegetative parts of Arabidopsis. Two ubiquitously expressed PI4P 5-kinase isoforms, PIP5K1 and PIP5K2, were identified and biochemically characterized. T-DNA insertion mutants for each gene did not show obvious phenotypes. However, the pip5k1 pip5k2 double mutant was severely dwarfed. Experiments devised to explain the growth phenotype indicate that reduced levels of PtdIns(4,5)P 2 in the mutants result in mislocalization and impaired membrane recycling of auxin-efflux-carriers (PIN-proteins). Direct analysis of auxin transport rates indicates that the double mutant has reduced capability for auxin transport compared to wild type controls. Furthermore, the pip5k1 pip5k2 double mutant is agravitropic, whereas over expressors of PIP5K1 or PIP5K2 exhibit random, non gravitropic growth. The dwarf phenotype might be a consequence of reduced meristem activity in the mutants, as was determined according to an expressed cyclin:CDB:GUS-reporter, which is possible due to reduced auxin in the root tip. Root apices did not accumulate radiolabel from [3H]sucrose, suggesting reduced sugar consumption in root meristems. Tissue patterning and vascular development was severely perturbed in root tips of the pip5k1 pip5k2 double mutant, indicating that the defective meristems have an impact on plant development. Overall, the last funding period has yielded important data that brought closure to a number of previously unfinished aspects of our work. Furthermore, the analysis of the pip5k1 pip5k2 double mutant has provided profound insights into the roles of PtdIns(4,5)P 2 in the vegetative protion of the plant and supports the notion that PtdIns(4,5)P 2 is of key importance for plant growth and development. While many questions have been answered, more and new questions have risen and will provide us with ample opportunity for discovery in our chosen field in the future.
Publications
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(2010) "Plant Physiology Classics Collection" Online-Kommentar zu Müller-Röber B und Pical C (2002) Plant Physiol 130: 22-46
Heilmann I, Im YJ, Dieck C, Perera IY, Boss WF
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(2010) At the poles across kingdoms: phosphoinositides and polar tip growth. Protoplasma 240: 13-31
Ischebeck T, Seiler S, Heilmann I
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(2010) Elevated phosphatidylinositol 3,4,5 trisphosphate in glia triggers cell autonomous membrane wrapping and myelination. J Neurosci 30: 8953-8964
Goebbels S, Oltrogge J, Kemper R, Heilmann I, Bormuth I, Wolfer S, Wichert S, Moebius W, Liu X, Lappe-Siefke C, Rossner M, Groszer M, Suter U, Frahm J, Boretius, S, Nave K-A
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(2010) Functional cooperativity of enzymes of phosphoinositide conversion according to synergistic effects on pectin secretion in tobacco pollen tubes. Mol Plant 3: 870-881
Ischebeck T, Vu LH, Jin X, Stenzel I, Löfke C, Heilmann I
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(2010) Phosphoinositide signaling in plants. The Lipid Library (website of the American Oil Chemists’ Society, AOCS)
Heilmann I
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(2010) Polyphosphoinositides are enriched in plant membrane rafts and form microdomains in the plasma membrane. Plant Physiol 152: 2173-2187
Furt F, König S, Bessoule J-J, Sargueil F, Zallot R, Stanislas T, Noirot E, Lherminier J, Simon- Plas F, Heilmann I, Mongrand S
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(2011) Phosphatidylinositol-4,5-bisphosphate influences Nt-Rac5-mediated cell expansion in pollen tubes of Nicotiana tabacum. Plant J 65: 453-468
Ischebeck T, Stenzel I, Hempel F, Jin X, Mosblech A, Heilmann I
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(2011) PIPKs are essential for rhizoid elongation and caulonemal cell development in the moss Physcomitrella patens. Plant J
Saavedra L, Balbi V, Lerche J, Mikami K, Heilmann I, Sommarin M
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(2011) The Hull of Fame: Lipid Signaling in the Plasma Membrane. In The Plant Plasma Membrane, Plant Cell Monographs, Murphy AS, Peer W, Schulz B, eds. Springer Academic Publishers, Berlin/Heidelberg, pp. 437-456
Im YJ, Heilmann I, Perera IY
