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Compartment-specific phosphoinositide dynamics in a central neuron

Prof. Dr. Dominik Oliver

Institute for Physiology und Pathophysiology
Philipps Universitity Marburg
Deutschhausstraße 1-2
D-35033 Marburg

phone: ++49 6421 2866444
fax:     ++49 6421 2862306

Homepage: http://www.uni-marburg.de/fb20/physiologie/ags/oliver


Project description 


Phosphoinositides (PIs) control many cellular processes including synaptic function and ion channel activity. We will use hippocampal pyramidal neurons as a model to investigate compartmentation of PI signaling. Changes in phosphatidylinositol(4,5)bisphosphate (PIP2) and phosphatidylinositol-(3,4,5)trisphosphate levels in response to a variety of physiological stimuli will be examined using FRET imaging. Particular attention will be paid to the highly compartmentalized dendritic compartments and postsynaptic spines. The dependence of ion channel activity and synaptic function on PIP2 will be probed with novel tools for spatiotemporally controlled depletion of PIP2.



Major goals

I. We want to obtain a spatiotemporal picture of synaptically induced PIP2 changes in the different compartments of the neuron. Thus, we will analyze changes in PIP2 concentrations during stimulation of modulatory synaptic inputs using fluorescent phosphoinositide probes, such as PH-PLCδ1-GFP or tubby-GFP.

Slice Results

PIP2-dynamics in acute hippocampal brain slices. (A) Expression of tubbyC-EGFP in CA1 pyramidal cells with a lentiviral vector. (B) Translocation of tubbyC from plasma membrane of a CA1 neuronal soma to the cytosol upon application of the muscarinic agonist Oxo-M indicates transient depletion of PIP2. (C) Example of mAChR-induced depletion of PIP2 in a dendrite. (D) Electrical stimulation in the stratum oriens of a hippocampal slice induced robust deplarization in a CA1 neuron, indicating efficient stimulation of cholinergic fibers and resulting synaptic activation of muscarinic receptors.

II. We aim at understanding the cellular signaling roles of the observed PIP2 dynamics. To this end, we will use recently developed methods to manipulate phosphoinositide concentrations in defined neuronal compartments. In particular we want to address the physiological function of phosphoinositides in dendritic/postsynaptic structures.




Manipulator Scheme


Genetically encoded tools for the manipulation of PI concentrations in living cells. (A) Voltage-activated PIP2-phosphatase Ci-VSP. (B) Chemical recruitment of the PIP2-degrading phosphatase Inp54p to the plasma membrane. Addition of rapamycin induces dimerization of the FRB domain of mTOR and the cytosolic FKBP protein fused to Inp54p. Because FRB fusion to the lyn11 provides membrane anchoring via palmitoylation and myristoylation, rapamycin-triggered dimerization forces the previously cytosolic Inp54p to the membrane. (C) Strategy for light-triggered recruitment of Inp54p. PhyB binds the chromophore phycocyanobilin (PCB) to form a light-sensitive holoprotein, which undergoes reversible conformational changes triggered by red and infrared light. Only the red-light induced state allows for association of the PIF domain. Plasma membrane anchoring by a C-terminal CAAX motif results in recruitment of PIF or PIF fusion constructs to the membrane upon illumination with red light.



Sandra Hackelberg, PhD student
Michael Leitner, PhD student
Olga Ivanova, PhD student
Anja Feuer, PhD student
Gisela Fischer, lab technician
Dr. Christian R. Halaszovich, Postdoc
Dr. Daniela N. Schreiber, Postdoc

Publications since 2007


Wilke BU, Linder M, Greifenberg L, Albus A, Kronimus Y, Bünemann M, Leitner MG, Oliver D (2014) Diacylglycerol mediates regulation of TASK potassium channels by Gq-coupled receptors. Nature Communications DOI:10.1038/ncomms6540

Bista P, Pawlowski M, Cerina M, Ehling P, Leist M, Meuth P, Aissaoui A, Borsotto M, Heurteaux C, Decher N, Pape HP, Oliver D, Meuth SG & Budde T (2014) Differential phospholipase C-dependent modulation of TWIK-related acid-sensitive K+ (TASK) and TWIK-related K+ (TREK) channels in rat thalamocortical relay neurons. J. Physiol  In Press.

Deng W, Nies F, Feuer A, Bočina I, Oliver D, Jiang D (2013) Anion Transport through Slc26 Family Protein Regulates Lumen Expansion during Tubulogenesis. Proc Natl Acad Sci USA 110:14972-14977. doi:10.1073/pnas.1220884110

Schiekel J, Lindner M, Hetzel A, Wemhöner K, Renigunta V, Schlichthörl G, Decher N, Oliver D*, Daut J* (2013) Inhibition of the potassium channel TASK-1 in rat cardiac muscle by endothelin-1 is mediated by phospholipase C. Cardiovasc Res. 97, 97-105.              *shared corresponding authorship

Halaszovich CR, Leitner MG, Le A, Frezza L, Feuer A, Schreiber DN, Villalba-Galea CA, Oliver D (2012) A human phospholipid phosphatase activated by a transmembrane control module. J Lip Res 53, 2266-2274

Leitner MG, Feuer A, Ebers O, Schreiber DN, Halaszovich CR, Oliver D (2012). Restoration of ion channel function in deafness-causing KCNQ4 mutants by synthetic channel openers. Br J Pharmacol 165:2244-2259.

Schaechinger TJ, Gorbunov D, Halaszovich CR, Moser T, Kugler S, Fakler B, Oliver D (2011). A synthetic prestin reveals protein domains and molecular operation of outer hair cell piezoelectricity. EMBO J 30:2793-2804

Lindner M., Leitner M.G., Halaszovich C.R., Hammond G.R., Oliver D. (2011). Probing the regulation of TASK potassium channels by PI(4,5)P2 with switchable phosphoinositide phosphatases.  J Physiol 589:3149-3162

Lacroix J, Halaszovich CR, Schreiber DN, Leitner MG, Bezanilla F, Oliver D, Villalba-Galea CA (2011). Controlling the Activity of a Phosphatase and Tensin Homolog (PTEN) by Membrane Potential. J Biol Chem 286:17945-17953

Leitner M.G., Halaszovich C.R., Oliver D. (2011). Aminoglycosides inhibit KCNQ4 channels in cochlear outer hair cells via depletion of phosphatidylinositol(4,5)bisphosphate. Mol Pharmacol 79:51-60

Reisinger E., Meintrup D., Oliver D., Fakler B. (2010). Gene expression associated with the onset of hearing detected by differential display in rat organ of Corti. Eur J Hum Genet 18:1327-1332.

Halaszovich C.R., Schreiber D.N., Oliver D. (2009). Ci-VSP is a depolarization-activated phosphatidylinositol-4,5-bisphosphate and phosphatidylinositol-3,4,5-trisphosphate 5'-phosphatase. J Biol Chem 284:2106-2113. 

Schaechinger T.J., Oliver D. (2007). Nonmammalian orthologs of prestin (SLC26A5) are electrogenic divalent/chloride anion exchangers. Proc Natl Acad Sci USA 104:7693-7698.

Zuletzt aktualisiert: 26.11.2014 · beimbort

SFB 593

Sonderforschungsbereich 593 - Institute of Cytobiology, Robert-Koch-Str. 6, 35037 Marburg, Germany
Tel. +49 6421/28-66899, Fax +49 6421/28-65482, E-Mail: sfb593@staff.uni-marburg.de

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