Deutsche Forschungsgemeinschaft Research Center for Molecular Physiology of the Brain

Göttingen, Germany

Deutsche Forschungsgemeinschaft Research Center for Molecular Physiology of the Brain

Göttingen, Germany

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Lipstein N.,Max Planck Institute for Experimental Medicine | Lipstein N.,Tel Aviv University | Schaks S.,Martin Luther University of Halle Wittenberg | Dimova K.,Max Planck Institute for Experimental Medicine | And 11 more authors.
Molecular and Cellular Biology | Year: 2012

Munc13s are presynaptic proteins that mediate synaptic vesicle priming and thereby control the size of the readily releasable pool of vesicles. During high synaptic activity, Munc13-1 and its closely related homolog, ubMunc13-2, bind Ca2+/calmodulin, resulting in enhanced priming activity and in changes of short-term synaptic plasticity characteristics. Here, we studied whether bMunc13-2 and Munc13-3, two remote isoforms of Munc13-1 with a neuronal subtype-specific expression pattern, mediate synaptic vesicle priming and regulate short-term synaptic plasticity in a Ca2+/calmodulin-dependent manner. We identified a single functional Ca2+/calmodulin binding site in these isoforms and provide structural evidence that all Munc13s employ a common mode of interaction with calmodulin despite the lack of sequence homology between their Ca2+/calmodulin binding sites. Electrophysiological analysis showed that, during high-frequency activity, Ca2+/calmodulin binding positively regulates the priming activity of bMunc13-2 and Munc13-3, resulting in an increase in the size of the readily releasable pool of vesicles and subsequently in strong short-term synaptic enhancement of neurotransmission. We conclude that Ca2+/calmodulin-dependent regulation of priming activity is structurally and functionally conserved in all Munc13 proteins, and that the composition of Munc13 isoforms in a neuron differentially controls its short-term synaptic plasticity characteristics. © 2012, American Society for Microbiology.

Dihazi H.,University of Gottingen | Dihazi G.H.,University of Gottingen | Jahn O.,Max Planck Institute for Experimental Medicine | Jahn O.,Deutsche Forschungsgemeinschaft Research Center for Molecular Physiology of the Brain | And 5 more authors.
Journal of Proteome Research | Year: 2011

Multipotent adult germline stem cells (maGSCs) are pluripotent cells that can be differentiated into somatic cells of the three primary germ layers. To highlight the protein profile changes associated with stem cell differentiation, retinoic acid (RA) treated mouse stem cells (maGSCs and ESCs) were compared to nontreated stem cells. 2-DE and DIGE reference maps were created, and differentially expressed proteins were further processed for identification. In both stem cell types, the RA induced differentiation resulted in an alteration of 36 proteins of which 18 were down-regulated and might be potential pluripotency associated proteins, whereas the other 18 proteins were up-regulated. These might be correlated to stem cell differentiation. Surprisingly, eukaryotic initiation factor 5A (Eif5a), a protein which is essential for cell proliferation and differentiation, was significantly down-regulated under RA treatment. A time-dependent investigation of Eif5a showed that the RA treatment of stem cells resulted in a significant up-regulation of the Eif5a in the first 48 h followed by a progressive down-regulation thereafter. This effect could be blocked by the hypusination inhibitor ciclopirox olamine (CPX). The alteration of Eif5a hypusination, as confirmed by mass spectrometry, exerts an antiproliferative effect on ESCs and maGSCs in vitro, but does not affect the cell pluripotency. Our data highlights the important role of Eif5a and its hypusination for stem cell differentiation and proliferation. © 2011 American Chemical Society.

Gomez-Varela D.,Max Planck Institute for Experimental Medicine | Kohl T.,Max Planck Institute for Experimental Medicine | Schmidt M.,European Neuroscience Institute | Schmidt M.,Deutsche Forschungsgemeinschaft Research Center for Molecular Physiology of the Brain | And 7 more authors.
PLoS ONE | Year: 2010

Voltage-gated ion channels are main players involved in fast synaptic events. However, only slow intracellular mechanisms have so far been described for controlling their localization as real-time visualization of endogenous voltage-gated channels at high temporal and spatial resolution has not been achieved yet. Using a specific extracellular antibody and quantum dots we reveal and characterize lateral mobility as a faster mechanism to dynamically control the number of endogenous ether-a-go-go (Eag)1 ion channels inside synapses. We visualize Eag1 entering and leaving synapses by lateral diffusion in the plasma membrane of rat hippocampal neurons. Mathematical analysis of their trajectories revealed how the motion of Eag1 gets restricted when the channels diffuse into the synapse, suggesting molecular interactions between Eag1 and synaptic components. In contrast, Eag1 channels switch to Brownian movement when they exit synapses and diffuse into extrasynaptic membranes. Furthermore, we demonstrate that the mobility of Eag1 channels is specifically regulated inside synapses by actin filaments, microtubules and electrical activity. In summary, using single-particle-tracking techniques with quantum dots nanocrystals, our study shows for the first time the lateral diffusion of an endogenous voltage-gated ion channel in neurons. The location-dependent constraints imposed by cytoskeletal elements together with the regulatory role of electrical activity strongly suggest a pivotal role for the mobility of voltage-gated ion channels in synaptic activity. © 2010 Gómez-Varela et al.

Breunig E.,University of Gottingen | Manzini I.,University of Gottingen | Manzini I.,Deutsche Forschungsgemeinschaft Research Center for Molecular Physiology of the Brain | Piscitelli F.,National Research Council Italy | And 5 more authors.
Journal of Neuroscience | Year: 2010

Cannabinoids modulate the activity of many neuronal cells, among them sensory neurons in the olfactory epithelium. Here we show that the endocannabinoid 2-arachidonoyl-glycerol (2-AG) is synthesized in both olfactory receptor neurons and glia-like sustentacular cells in larval Xenopus laevis. Its production in the latter depends on the hunger state of the animal. The essential effect of 2-AG in olfactory receptor neurons is the control of odorant detection thresholds via cannabinoid CB1 receptor activation. Hunger renders olfactory neurons more sensitive. Endocannabinoid modulation in the nose may therefore substantially influence food-seeking behavior. Copyright©2010 the authors.

Lakamper S.,University of Gottingen | Lakamper S.,University of Amsterdam | Lakamper S.,Deutsche Forschungsgemeinschaft Research Center for Molecular Physiology of the Brain | Thiede C.,University of Gottingen | And 9 more authors.
Journal of Molecular Biology | Year: 2010

Controlled activity of several kinesin motors is required for the proper assembly of the mitotic spindle. Eg5, a homotetrameric bipolar kinesin-5 from Xenopus laevis, can cross-link and slide anti-parallel microtubules apart by a motility mechanism comprising diffusional and directional modes. How this mechanism is regulated, possibly by the tail domains of the opposing motors, is poorly understood. In order to explore the basic unregulated kinesin-5 motor activity, we generated a stably dimeric kinesin-5 construct, Eg5Kin, consisting of the motor domain and neck linker of Eg5 and the neck coiled coil of Drosophila melanogaster kinesin-1 (DmKHC). In single-molecule motility assays, we found this chimera to be highly processive. In addition, we studied the effect of the kinesin-5-specific inhibitor monastrol using single-molecule fluorescence assays. We found that monastrol reduced the length of processive runs, but strikingly did not affect velocity. Quantitative analysis of monastrol dose dependence suggests that two bound monastrol molecules are required to be bound to an Eg5Kin dimer to terminate a run. © 2010 Elsevier Ltd.

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