@misc{LindowRedemannFabigetal., author = {Lindow, Norbert and Redemann, Stefanie and Fabig, Gunar and M{\"u}ller-Reichert, Thomas and Prohaska, Steffen}, title = {Quantification of Three-Dimensional Spindle Architecture}, issn = {1438-0064}, url = {http://nbn-resolving.de/urn:nbn:de:0297-zib-66562}, abstract = {Mitotic and meiotic spindles are microtubule-based structures to faithfully segregate chromosomes. Electron tomography is currently the method of choice to analyze the three-dimensional architecture of both types of spindles. Over the years, we have developed methods and software for automatic segmentation and stitching of microtubules in serial sections for large-scale reconstructions. Three-dimensional reconstruction of microtubules, however, is only the first step towards biological insight. The second step is the analysis of the structural data to derive measurable spindle properties. Here, we present a comprehensive set of techniques to quantify spindle parameters. These techniques provide quantitative analyses of specific microtubule classes and are applicable to a variety of tomographic reconstructions of spindles from different organisms.}, language = {en} } @article{JinKiralOzeletal., author = {Jin, Eugene Jennifer and Kiral, Ferdi Ridvan and Ozel, Mehmet Neset and Burchardt, Lara Sophie and Osterland, Marc and Epstein, Daniel and Wolfenberg, Heike and Prohaska, Steffen and Hiesinger, Peter Robin}, title = {Live Observation of Two Parallel Membrane Degradation Pathways at Axon Terminals}, series = {Current Biology}, volume = {28}, journal = {Current Biology}, number = {7}, doi = {10.1016/j.cub.2018.02.032}, pages = {1027 -- 1038.e4}, abstract = {Neurons are highly polarized cells that require continuous turnover of membrane proteins at axon terminals to develop, function, and survive. Yet, it is still unclear whether membrane protein degradation requires transport back to the cell body or whether degradation also occurs locally at the axon terminal, where live observation of sorting and degradation has remained a challenge. Here, we report direct observation of two cargo-specific membrane protein degradation mechanisms at axon terminals based on a live-imaging approach in intact Drosophila brains. We show that different acidification-sensing cargo probes are sorted into distinct classes of degradative ''hub'' compartments for synaptic vesicle proteins and plasma membrane proteins at axon terminals. Sorting and degradation of the two cargoes in the separate hubs are molecularly distinct. Local sorting of synaptic vesicle proteins for degradation at the axon terminal is, surprisingly, Rab7 independent, whereas sorting of plasma membrane proteins is Rab7 dependent. The cathepsin-like protease CP1 is specific to synaptic vesicle hubs, and its delivery requires the vesicle SNARE neuronal synaptobrevin. Cargo separation only occurs at the axon terminal, whereas degradative compartments at the cell body are mixed. These data show that at least two local, molecularly distinct pathways sort membrane cargo for degradation specifically at the axon terminal, whereas degradation can occur both at the terminal and en route to the cell body.}, language = {en} } @incollection{LindowRedemannBruenigetal., author = {Lindow, Norbert and Redemann, Stefanie and Br{\"u}nig, Florian and Fabig, Gunar and M{\"u}ller-Reichert, Thomas and Prohaska, Steffen}, title = {Quantification of three-dimensional spindle architecture}, series = {Methods in Cell Biology Part B}, volume = {145}, booktitle = {Methods in Cell Biology Part B}, publisher = {Academic Press}, issn = {0091-679X}, doi = {10.1016/bs.mcb.2018.03.012}, pages = {45 -- 64}, abstract = {Mitotic and meiotic spindles are microtubule-based structures to faithfully segregate chromosomes. Electron tomography is currently the method of choice to analyze the three-dimensional (3D) architecture of both types of spindles. Over the years, we have developed methods and software for automatic segmentation and stitching of microtubules in serial sections for large-scale reconstructions. 3D reconstruction of microtubules, however, is only the first step toward biological insight. The second step is the analysis of the structural data to derive measurable spindle properties. Here, we present a comprehensive set of techniques to quantify spindle parameters. These techniques provide quantitative analyses of specific microtubule classes and are applicable to a variety of tomographic reconstructions of spindles from different organisms.}, language = {en} } @misc{RedemannLantzschLindowetal., author = {Redemann, Stefanie and Lantzsch, Ina and Lindow, Norbert and Prohaska, Steffen and Srayko, Martin and M{\"u}ller-Reichert, Thomas}, title = {A switch in microtubule orientation during C. elegans meiosis}, issn = {1438-0064}, url = {http://nbn-resolving.de/urn:nbn:de:0297-zib-69855}, abstract = {In oocytes of many organisms, meiotic spindles form in the absence of centrosomes [1-5]. Such female meiotic spindles have a pointed appearance in metaphase with microtubules focused at acentrosomal spindle poles. At anaphase, the microtubules of acentrosomal spindles then transition to an inter- chromosomal array, while the spindle poles disappear. This transition is currently not understood. Previous studies have focused on this inter- chromosomal microtubule array and proposed a pushing model to drive chromosome segregation [6, 7]. This model includes an end-on orientation of microtubules with chromosomes. Alternatively, chromosomes were thought to associate along bundles of microtubules [8, 9]. Starting with metaphase, this second model proposed a pure lateral chromosome-to-microtubule association up to the final meiotic stages of anaphase. Here we applied large-scale electron tomography [10] of staged C. elegans oocytes in meiosis to analyze the orientation of microtubules in respect to chromosomes. We show that microtubules at metaphase I are primarily oriented laterally to the chromosomes and that microtubules switch to an end-on orientation during progression through anaphase. We further show that this switch in microtubule orientation involves a kinesin-13 microtubule depolymerase, KLP-7, which removes laterally associated microtubules around chromosomes. From this we conclude that both lateral and end-on modes of microtubule-to-chromosome orientations are successively used in C. elegans oocytes to segregate meiotic chromosomes.}, language = {en} } @article{RedemannLantzschLindowetal., author = {Redemann, Stefanie and Lantzsch, Ina and Lindow, Norbert and Prohaska, Steffen and Srayko, Martin and M{\"u}ller-Reichert, Thomas}, title = {A switch in microtubule orientation during C. elegans meiosis}, series = {Current Biology}, journal = {Current Biology}, issn = {0960-9822}, doi = {10.1016/j.cub.2018.07.012}, abstract = {In oocytes of many organisms, meiotic spindles form in the absence of centrosomes [1-5]. Such female meiotic spindles have a pointed appearance in metaphase with microtubules focused at acentrosomal spindle poles. At anaphase, the microtubules of acentrosomal spindles then transition to an inter- chromosomal array, while the spindle poles disappear. This transition is currently not understood. Previous studies have focused on this inter- chromosomal microtubule array and proposed a pushing model to drive chromosome segregation [6, 7]. This model includes an end-on orientation of microtubules with chromosomes. Alternatively, chromosomes were thought to associate along bundles of microtubules [8, 9]. Starting with metaphase, this second model proposed a pure lateral chromosome-to-microtubule association up to the final meiotic stages of anaphase. Here we applied large-scale electron tomography [10] of staged C. elegans oocytes in meiosis to analyze the orientation of microtubules in respect to chromosomes. We show that microtubules at metaphase I are primarily oriented laterally to the chromosomes and that microtubules switch to an end-on orientation during progression through anaphase. We further show that this switch in microtubule orientation involves a kinesin-13 microtubule depolymerase, KLP-7, which removes laterally associated microtubules around chromosomes. From this we conclude that both lateral and end-on modes of microtubule-to-chromosome orientations are successively used in C. elegans oocytes to segregate meiotic chromosomes.}, language = {en} }