@article{14788, abstract = {Eukaryotic cells use clathrin-mediated endocytosis to take up a large range of extracellular cargo. During endocytosis, a clathrin coat forms on the plasma membrane, but it remains controversial when and how it is remodeled into a spherical vesicle. Here, we use 3D superresolution microscopy to determine the precise geometry of the clathrin coat at large numbers of endocytic sites. Through pseudo-temporal sorting, we determine the average trajectory of clathrin remodeling during endocytosis. We find that clathrin coats assemble first on flat membranes to 50% of the coat area before they become rapidly and continuously bent, and this mechanism is confirmed in three cell lines. We introduce the cooperative curvature model, which is based on positive feedback for curvature generation. It accurately describes the measured shapes and dynamics of the clathrin coat and could represent a general mechanism for clathrin coat remodeling on the plasma membrane.}, author = {Mund, Markus and Tschanz, Aline and Wu, Yu-Le and Frey, Felix F and Mehl, Johanna L. and Kaksonen, Marko and Avinoam, Ori and Schwarz, Ulrich S. and Ries, Jonas}, issn = {1540-8140}, journal = {Journal of Cell Biology}, keywords = {Cell Biology}, number = {3}, publisher = {Rockefeller University Press}, title = {{Clathrin coats partially preassemble and subsequently bend during endocytosis}}, doi = {10.1083/jcb.202206038}, volume = {222}, year = {2023}, } @article{12080, abstract = {Endocytosis is a multistep process involving the sequential recruitment and action of numerous proteins. This process can be divided into two phases: an early phase, in which sites of endocytosis are formed, and a late phase in which clathrin-coated vesicles are formed and internalized into the cytosol, but how these phases link to each other remains unclear. In this study, we demonstrate that anchoring the yeast Eps15-like protein Pan1p to the peroxisome triggers most of the events occurring during the late phase at the peroxisome. At this ectopic location, Pan1p recruits most proteins that function in the late phases—including actin nucleation promoting factors—and then initiates actin polymerization. Pan1p also recruited Prk1 kinase and actin depolymerizing factors, thereby triggering disassembly immediately after actin assembly and inducing dissociation of endocytic proteins from the peroxisome. These observations suggest that Pan1p is a key regulator for initiating, processing, and completing the late phase of endocytosis.}, author = {Enshoji, Mariko and Miyano, Yoshiko and Yoshida, Nao and Nagano, Makoto and Watanabe, Minami and Kunihiro, Mayumi and Siekhaus, Daria E and Toshima, Junko Y. and Toshima, Jiro}, issn = {1540-8140}, journal = {Journal of Cell Biology}, number = {10}, publisher = {Rockefeller University Press}, title = {{Eps15/Pan1p is a master regulator of the late stages of the endocytic pathway}}, doi = {10.1083/jcb.202112138}, volume = {221}, year = {2022}, } @article{12121, abstract = {Autophagosomes are double-membraned vesicles that traffic harmful or unwanted cellular macromolecules to the vacuole for recycling. Although autophagosome biogenesis has been extensively studied, autophagosome maturation, i.e., delivery and fusion with the vacuole, remains largely unknown in plants. Here, we have identified an autophagy adaptor, CFS1, that directly interacts with the autophagosome marker ATG8 and localizes on both membranes of the autophagosome. Autophagosomes form normally in Arabidopsis thaliana cfs1 mutants, but their delivery to the vacuole is disrupted. CFS1’s function is evolutionarily conserved in plants, as it also localizes to the autophagosomes and plays a role in autophagic flux in the liverwort Marchantia polymorpha. CFS1 regulates autophagic flux by bridging autophagosomes with the multivesicular body-localized ESCRT-I component VPS23A, leading to the formation of amphisomes. Similar to CFS1-ATG8 interaction, disrupting the CFS1-VPS23A interaction blocks autophagic flux and renders plants sensitive to nitrogen starvation. Altogether, our results reveal a conserved vacuolar sorting hub that regulates autophagic flux in plants.}, author = {Zhao, Jierui and Bui, Mai Thu and Ma, Juncai and Künzl, Fabian and Picchianti, Lorenzo and De La Concepcion, Juan Carlos and Chen, Yixuan and Petsangouraki, Sofia and Mohseni, Azadeh and García-Leon, Marta and Gomez, Marta Salas and Giannini, Caterina and Gwennogan, Dubois and Kobylinska, Roksolana and Clavel, Marion and Schellmann, Swen and Jaillais, Yvon and Friml, Jiří and Kang, Byung-Ho and Dagdas, Yasin}, issn = {1540-8140}, journal = {Journal of Cell Biology}, keywords = {Cell Biology}, number = {12}, publisher = {Rockefeller University Press}, title = {{Plant autophagosomes mature into amphisomes prior to their delivery to the central vacuole}}, doi = {10.1083/jcb.202203139}, volume = {221}, year = {2022}, } @article{12122, abstract = {Centrosomes play a crucial role during immune cell interactions and initiation of the immune response. In proliferating cells, centrosome numbers are tightly controlled and generally limited to one in G1 and two prior to mitosis. Defects in regulating centrosome numbers have been associated with cell transformation and tumorigenesis. Here, we report the emergence of extra centrosomes in leukocytes during immune activation. Upon antigen encounter, dendritic cells pass through incomplete mitosis and arrest in the subsequent G1 phase leading to tetraploid cells with accumulated centrosomes. In addition, cell stimulation increases expression of polo-like kinase 2, resulting in diploid cells with two centrosomes in G1-arrested cells. During cell migration, centrosomes tightly cluster and act as functional microtubule-organizing centers allowing for increased persistent locomotion along gradients of chemotactic cues. Moreover, dendritic cells with extra centrosomes display enhanced secretion of inflammatory cytokines and optimized T cell responses. Together, these results demonstrate a previously unappreciated role of extra centrosomes for regular cell and tissue homeostasis.}, author = {Weier, Ann-Kathrin and Homrich, Mirka and Ebbinghaus, Stephanie and Juda, Pavel and Miková, Eliška and Hauschild, Robert and Zhang, Lili and Quast, Thomas and Mass, Elvira and Schlitzer, Andreas and Kolanus, Waldemar and Burgdorf, Sven and Gruß, Oliver J. and Hons, Miroslav and Wieser, Stefan and Kiermaier, Eva}, issn = {1540-8140}, journal = {Journal of Cell Biology}, keywords = {Cell Biology}, number = {12}, publisher = {Rockefeller University Press}, title = {{Multiple centrosomes enhance migration and immune cell effector functions of mature dendritic cells}}, doi = {10.1083/jcb.202107134}, volume = {221}, year = {2022}, } @article{12272, abstract = {Reading, interpreting and crawling along gradients of chemotactic cues is one of the most complex questions in cell biology. In this issue, Georgantzoglou et al. (2022. J. Cell. Biol.https://doi.org/10.1083/jcb.202103207) use in vivo models to map the temporal sequence of how neutrophils respond to an acutely arising gradient of chemoattractant.}, author = {Stopp, Julian A and Sixt, Michael K}, issn = {1540-8140}, journal = {Journal of Cell Biology}, keywords = {Cell Biology}, number = {8}, publisher = {Rockefeller University Press}, title = {{Plan your trip before you leave: The neutrophils’ search-and-run journey}}, doi = {10.1083/jcb.202206127}, volume = {221}, year = {2022}, } @article{9094, abstract = {Dendritic cells (DCs) are crucial for the priming of naive T cells and the initiation of adaptive immunity. Priming is initiated at a heterologous cell–cell contact, the immunological synapse (IS). While it is established that F-actin dynamics regulates signaling at the T cell side of the contact, little is known about the cytoskeletal contribution on the DC side. Here, we show that the DC actin cytoskeleton is decisive for the formation of a multifocal synaptic structure, which correlates with T cell priming efficiency. DC actin at the IS appears in transient foci that are dynamized by the WAVE regulatory complex (WRC). The absence of the WRC in DCs leads to stabilized contacts with T cells, caused by an increase in ICAM1-integrin–mediated cell–cell adhesion. This results in lower numbers of activated and proliferating T cells, demonstrating an important role for DC actin in the regulation of immune synapse functionality.}, author = {Leithner, Alexander F and Altenburger, LM and Hauschild, R and Assen, Frank P and Rottner, K and TEB, Stradal and Diz-Muñoz, A and Stein, JV and Sixt, Michael K}, issn = {1540-8140}, journal = {Journal of Cell Biology}, number = {4}, publisher = {Rockefeller University Press}, title = {{Dendritic cell actin dynamics control contact duration and priming efficiency at the immunological synapse}}, doi = {10.1083/jcb.202006081}, volume = {220}, year = {2021}, } @article{10337, abstract = {The T cell receptor (TCR) pathway receives, processes, and amplifies the signal from pathogenic antigens to the activation of T cells. Although major components in this pathway have been identified, the knowledge on how individual components cooperate to effectively transduce signals remains limited. Phase separation emerges as a biophysical principle in organizing signaling molecules into liquid-like condensates. Here, we report that phospholipase Cγ1 (PLCγ1) promotes phase separation of LAT, a key adaptor protein in the TCR pathway. PLCγ1 directly cross-links LAT through its two SH2 domains. PLCγ1 also protects LAT from dephosphorylation by the phosphatase CD45 and promotes LAT-dependent ERK activation and SLP76 phosphorylation. Intriguingly, a nonmonotonic effect of PLCγ1 on LAT clustering was discovered. Computer simulations, based on patchy particles, revealed how the cluster size is regulated by protein compositions. Together, these results define a critical function of PLCγ1 in promoting phase separation of the LAT complex and TCR signal transduction.}, author = {Zeng, Longhui and Palaia, Ivan and Šarić, Anđela and Su, Xiaolei}, issn = {1540-8140}, journal = {Journal of Cell Biology}, keywords = {cell biology}, number = {6}, publisher = {Rockefeller University Press}, title = {{PLCγ1 promotes phase separation of T cell signaling components}}, doi = {10.1083/jcb.202009154}, volume = {220}, year = {2021}, } @article{11061, abstract = {Many adult tissues contain postmitotic cells as old as the host organism. The only organelle that does not turn over in these cells is the nucleus, and its maintenance represents a formidable challenge, as it harbors regulatory proteins that persist throughout adulthood. Here we developed strategies to visualize two classes of such long-lived proteins, histones and nucleoporins, to understand the function of protein longevity in nuclear maintenance. Genome-wide mapping of histones revealed specific enrichment of long-lived variants at silent gene loci. Interestingly, nuclear pores are maintained by piecemeal replacement of subunits, resulting in mosaic complexes composed of polypeptides with vastly different ages. In contrast, nondividing quiescent cells remove old nuclear pores in an ESCRT-dependent manner. Our findings reveal distinct molecular strategies of nuclear maintenance, linking lifelong protein persistence to gene regulation and nuclear integrity.}, author = {Toyama, Brandon H. and Arrojo e Drigo, Rafael and Lev-Ram, Varda and Ramachandra, Ranjan and Deerinck, Thomas J. and Lechene, Claude and Ellisman, Mark H. and HETZER, Martin W}, issn = {1540-8140}, journal = {Journal of Cell Biology}, keywords = {Cell Biology}, number = {2}, pages = {433--444}, publisher = {Rockefeller University Press}, title = {{Visualization of long-lived proteins reveals age mosaicism within nuclei of postmitotic cells}}, doi = {10.1083/jcb.201809123}, volume = {218}, year = {2019}, } @article{11069, abstract = {Repeated rounds of nuclear envelope (NE) rupture and repair have been observed in laminopathy and cancer cells and result in intermittent loss of nucleus compartmentalization. Currently, the causes of NE rupture are unclear. Here, we show that NE rupture in cancer cells relies on the assembly of contractile actin bundles that interact with the nucleus via the linker of nucleoskeleton and cytoskeleton (LINC) complex. We found that the loss of actin bundles or the LINC complex did not rescue nuclear lamina defects, a previously identified determinant of nuclear membrane stability, but did decrease the number and size of chromatin hernias. Finally, NE rupture inhibition could be rescued in cells treated with actin-depolymerizing drugs by mechanically constraining nucleus height. These data suggest a model of NE rupture where weak membrane areas, caused by defects in lamina organization, rupture because of an increase in intranuclear pressure from actin-based nucleus confinement.}, author = {Hatch, Emily M. and HETZER, Martin W}, issn = {0021-9525}, journal = {Journal of Cell Biology}, keywords = {Cell Biology}, number = {1}, pages = {27--36}, publisher = {Rockefeller University Press}, title = {{Nuclear envelope rupture is induced by actin-based nucleus confinement}}, doi = {10.1083/jcb.201603053}, volume = {215}, year = {2016}, } @article{11075, abstract = {Previously, we identified the nucleoporin gp210/Nup210 as a critical regulator of muscle and neuronal differentiation, but how this nucleoporin exerts its function and whether it modulates nuclear pore complex (NPC) activity remain unknown. Here, we show that gp210/Nup210 mediates muscle cell differentiation in vitro via its conserved N-terminal domain that extends into the perinuclear space. Removal of the C-terminal domain, which partially mislocalizes gp210/Nup210 away from NPCs, efficiently rescues the differentiation defect caused by the knockdown of endogenous gp210/Nup210. Unexpectedly, a gp210/Nup210 mutant lacking the NPC-targeting transmembrane and C-terminal domains is sufficient for C2C12 myoblast differentiation. We demonstrate that the endoplasmic reticulum (ER) stress-specific caspase cascade is exacerbated during Nup210 depletion and that blocking ER stress-mediated apoptosis rescues differentiation of Nup210-deficient cells. Our results suggest that the role of gp210/Nup210 in cell differentiation is mediated by its large luminal domain, which can act independently of NPC association and appears to play a pivotal role in the maintenance of nuclear envelope/ER homeostasis.}, author = {Gomez-Cavazos, J. Sebastian and HETZER, Martin W}, issn = {1540-8140}, journal = {Journal of Cell Biology}, keywords = {Cell Biology}, number = {6}, pages = {671--681}, publisher = {Rockefeller University Press}, title = {{The nucleoporin gp210/Nup210 controls muscle differentiation by regulating nuclear envelope/ER homeostasis}}, doi = {10.1083/jcb.201410047}, volume = {208}, year = {2015}, } @article{11081, abstract = {In eukaryotic cells the nuclear genome is enclosed by the nuclear envelope (NE). In metazoans, the NE breaks down in mitosis and it has been assumed that the physical barrier separating nucleoplasm and cytoplasm remains intact during the rest of the cell cycle and cell differentiation. However, recent studies suggest that nonmitotic NE remodeling plays a critical role in development, virus infection, laminopathies, and cancer. Although the mechanisms underlying these NE restructuring events are currently being defined, one common theme is activation of protein kinase C family members in the interphase nucleus to disrupt the nuclear lamina, demonstrating the importance of the lamina in maintaining nuclear integrity.}, author = {Hatch, Emily and HETZER, Martin W}, issn = {1540-8140}, journal = {Journal of Cell Biology}, keywords = {Cell Biology}, number = {2}, pages = {133--141}, publisher = {Rockefeller University Press}, title = {{Breaching the nuclear envelope in development and disease}}, doi = {10.1083/jcb.201402003}, volume = {205}, year = {2014}, } @article{11094, abstract = {Nuclear pore complexes (NPCs) assemble at the end of mitosis during nuclear envelope (NE) reformation and into an intact NE as cells progress through interphase. Although recent studies have shown that NPC formation occurs by two different molecular mechanisms at two distinct cell cycle stages, little is known about the molecular players that mediate the fusion of the outer and inner nuclear membranes to form pores. In this paper, we provide evidence that the transmembrane nucleoporin (Nup), POM121, but not the Nup107–160 complex, is present at new pore assembly sites at a time that coincides with inner nuclear membrane (INM) and outer nuclear membrane (ONM) fusion. Overexpression of POM121 resulted in juxtaposition of the INM and ONM. Additionally, Sun1, an INM protein that is known to interact with the cytoskeleton, was specifically required for interphase assembly and localized with POM121 at forming pores. We propose a model in which POM121 and Sun1 interact transiently to promote early steps of interphase NPC assembly.}, author = {Talamas, Jessica A. and HETZER, Martin W}, issn = {1540-8140}, journal = {Journal of Cell Biology}, keywords = {Cell Biology}, number = {1}, pages = {27--37}, publisher = {Rockefeller University Press}, title = {{POM121 and Sun1 play a role in early steps of interphase NPC assembly}}, doi = {10.1083/jcb.201012154}, volume = {194}, year = {2011}, } @article{11106, abstract = {Formation of the nuclear envelope (NE) around segregated chromosomes occurs by the reshaping of the endoplasmic reticulum (ER), a reservoir for disassembled nuclear membrane components during mitosis. In this study, we show that inner nuclear membrane proteins such as lamin B receptor (LBR), MAN1, Lap2β, and the trans-membrane nucleoporins Ndc1 and POM121 drive the spreading of ER membranes into the emerging NE via their capacity to bind chromatin in a collaborative manner. Despite their redundant functions, decreasing the levels of any of these trans-membrane proteins by RNAi-mediated knockdown delayed NE formation, whereas increasing the levels of any of them had the opposite effect. Furthermore, acceleration of NE formation interferes with chromosome separation during mitosis, indicating that the time frame over which chromatin becomes membrane enclosed is physiologically relevant and regulated. These data suggest that functionally distinct classes of chromatin-interacting membrane proteins, which are present at nonsaturating levels, collaborate to rapidly reestablish the nuclear compartment at the end of mitosis.}, author = {Anderson, Daniel J. and Vargas, Jesse D. and Hsiao, Joshua P. and HETZER, Martin W}, issn = {1540-8140}, journal = {Journal of Cell Biology}, keywords = {Cell Biology}, number = {2}, pages = {183--191}, publisher = {Rockefeller University Press}, title = {{Recruitment of functionally distinct membrane proteins to chromatin mediates nuclear envelope formation in vivo}}, doi = {10.1083/jcb.200901106}, volume = {186}, year = {2009}, } @article{11107, abstract = {Nucleocytoplasmic transport occurs exclusively through nuclear pore complexes (NPCs) embedded in pores formed by inner and outer nuclear membrane fusion. The mechanism for de novo pore and NPC biogenesis remains unclear. Reticulons (RTNs) and Yop1/DP1 are conserved membrane protein families required to form and maintain the tubular endoplasmic reticulum (ER) and the postmitotic nuclear envelope. In this study, we report that members of the RTN and Yop1/DP1 families are required for nuclear pore formation. Analysis of Saccharomyces cerevisiae prp20-G282S and nup133Δ NPC assembly mutants revealed perturbations in Rtn1–green fluorescent protein (GFP) and Yop1-GFP ER distribution and colocalization to NPC clusters. Combined deletion of RTN1 and YOP1 resulted in NPC clustering, nuclear import defects, and synthetic lethality with the additional absence of Pom34, Pom152, and Nup84 subcomplex members. We tested for a direct role in NPC biogenesis using Xenopus laevis in vitro assays and found that anti-Rtn4a antibodies specifically inhibited de novo nuclear pore formation. We hypothesize that these ER membrane–bending proteins mediate early NPC assembly steps.}, author = {Dawson, T. Renee and Lazarus, Michelle D. and HETZER, Martin W and Wente, Susan R.}, issn = {1540-8140}, journal = {Journal of Cell Biology}, keywords = {Cell Biology}, number = {5}, pages = {659--675}, publisher = {Rockefeller University Press}, title = {{ER membrane–bending proteins are necessary for de novo nuclear pore formation}}, doi = {10.1083/jcb.200806174}, volume = {184}, year = {2009}, } @article{11111, abstract = {During mitosis in metazoans, segregated chromosomes become enclosed by the nuclear envelope (NE), a double membrane that is continuous with the endoplasmic reticulum (ER). Recent in vitro data suggest that NE formation occurs by chromatin-mediated reorganization of the tubular ER; however, the basic principles of such a membrane-reshaping process remain uncharacterized. Here, we present a quantitative analysis of nuclear membrane assembly in mammalian cells using time-lapse microscopy. From the initial recruitment of ER tubules to chromatin, the formation of a membrane-enclosed, transport-competent nucleus occurs within ∼12 min. Overexpression of the ER tubule-forming proteins reticulon 3, reticulon 4, and DP1 inhibits NE formation and nuclear expansion, whereas their knockdown accelerates nuclear assembly. This suggests that the transition from membrane tubules to sheets is rate-limiting for nuclear assembly. Our results provide evidence that ER-shaping proteins are directly involved in the reconstruction of the nuclear compartment and that morphological restructuring of the ER is the principal mechanism of NE formation in vivo.}, author = {Anderson, Daniel J. and HETZER, Martin W}, issn = {1540-8140}, journal = {Journal of Cell Biology}, keywords = {Cell Biology}, number = {5}, pages = {911--924}, publisher = {Rockefeller University Press}, title = {{Reshaping of the endoplasmic reticulum limits the rate for nuclear envelope formation}}, doi = {10.1083/jcb.200805140}, volume = {182}, year = {2008}, } @article{3930, abstract = {An active involvement of blood-brain barrier endothelial cell basement membranes in development of inflammatory lesions in the central nervous system (CNS) has not been considered to date. Here we investigated the molecular composition and possible function of the extracellular matrix encountered by extravasating T lymphocytes during experimental autoimmune encephalomyelitis (EAE). Endothelial basement membranes contained laminin 8 (alpha4beta1gamma1) and/or 10 (alpha5beta1gamma1) and their expression was influenced by proinflammatory cytokines or angiostatic agents. T cells emigrating into the CNS during EAE encountered two biochemically distinct basement membranes, the endothelial (containing laminins 8 and 10) and the parenchymal (containing laminins 1 and 2) basement membranes. However, inflammatory cuffs occurred exclusively around endothelial basement membranes containing laminin 8, whereas in the presence of laminin 10 no infiltration was detectable. In vitro assays using encephalitogenic T cell lines revealed adhesion to laminins 8 and 10, whereas binding to laminins 1 and 2 could not be induced. Downregulation of integrin alpha6 on cerebral endothelium at sites of T cell infiltration, plus a high turnover of laminin 8 at these sites, suggested two possible roles for laminin 8 in the endothelial basement membrane: one at the level of the endothelial cells resulting in reduced adhesion and, thereby, increased penetrability of the monolayer; and secondly at the level of the T cells providing direct signals to the transmigrating cells.}, author = {Sixt, Michael K and Engelhardt, Britta and Pausch, Friederike and Hallmann, Rupert and Wendler, Olaf and Sorokin, Lydia}, issn = {0021-9525}, journal = {Journal of Cell Biology}, number = {5}, pages = {933 -- 946}, publisher = {Rockefeller University Press}, title = {{Endothelial cell laminin isoforms, laminins 8 and 10, play decisive roles in T cell recruitment across the blood-brain barrier in experimental autoimmune encephalomyelitis}}, doi = {10.1083/jcb.153.5.933 }, volume = {153}, year = {2001}, } @article{11126, abstract = {Nuclear import of the two uracil-rich small nuclear ribonucleoprotein (U snRNP) components U1A and U2B′′ is mediated by unusually long and complex nuclear localization signals (NLSs). Here we investigate nuclear import of U1A and U2B′′ in vitro and demonstrate that it occurs by an active, saturable process. Several lines of evidence suggest that import of the two proteins occurs by an import mechanism different to those characterized previously. No cross competition is seen with a variety of previously studied NLSs. In contrast to import mediated by members of the importin-β family of nucleocytoplasmic transport receptors, U1A/U2B′′ import is not inhibited by either nonhydrolyzable guanosine triphosphate (GTP) analogues or by a mutant of the GTPase Ran that is incapable of GTP hydrolysis. Adenosine triphosphate is capable of supporting U1A and U2B′′ import, whereas neither nonhydrolyzable adenosine triphosphate analogues nor GTP can do so. U1A and U2B′′ import in vitro does not require the addition of soluble cytosolic proteins, but a factor or factors required for U1A and U2B′′ import remains tightly associated with the nuclear fraction of conventionally permeabilized cells. This activity can be solubilized in the presence of elevated MgCl2. These data suggest that U1A and U2B′′ import into the nucleus occurs by a hitherto uncharacterized mechanism.}, author = {HETZER, Martin W and Mattaj, Iain W.}, issn = {1540-8140}, journal = {Journal of Cell Biology}, keywords = {Cell Biology}, number = {2}, pages = {293--304}, publisher = {Rockefeller University Press}, title = {{An Atp-dependent, Ran-independent mechanism for nuclear import of the U1a and U2b′′ spliceosome proteins}}, doi = {10.1083/jcb.148.2.293}, volume = {148}, year = {2000}, } @article{3756, abstract = {In many eukaryotic cells going through M-phase, a bipolar spindle is formed by microtubules nucleated from centrosomes. These microtubules, in addition to being `'captured” by kinetochores, may be stabilized by chromatin in two different ways: short-range stabilization effects may affect microtubules in close contact with the chromatin, while long-range stabilization effects may `'guide” microtubule growth towards the chromatin (e.g., by introducing a diffusive gradient of an enzymatic activity that affects microtubule assembly). Here, we use both meiotic and mitotic extracts from Xenopus laevis eggs to study microtubule aster formation and microtubule dynamics in the presence of chromatin. In `'low-speed” meiotic extracts, in the presence of salmon sperm chromatin, we find that short-range stabilization effects lead to a strong anisotropy of the microtubule asters. Analysis of the dynamic parameters of microtubule growth shows that this anisotropy arises from a decrease in the catastrophe frequency, an increase in the rescue frequency and a decrease in the growth velocity. In this system we also find evidence for long-range `'guidance” effects, which lead to a weak anisotropy of the asters. Statistically relevant results on these long-range effects are obtained in `'high-speed” mitotic extracts in the presence of artificially constructed chromatin stripes. We find that aster anisotropy is biased in the direction of the chromatin and that the catastrophe frequency is reduced in its vicinity. In this system we also find a surprising dependence of the catastrophe and the rescue frequencies on the length of microtubules nucleated from centrosomes: the catastrophe frequency increases and the rescue frequency decreases with microtubule length.}, author = {Dogterom, Marileen and Felix, M. and Guet, Calin C and Leibler, Stanislas}, issn = {0021-9525}, journal = {Journal of Cell Biology}, number = {1}, pages = {125 -- 140}, publisher = {Rockefeller University Press}, title = {{Influence of M-phase chromatin on the anisotropy of microtubule asters}}, doi = {doi: 10.1083/jcb.133.1.125 }, volume = {133}, year = {1996}, } @article{4177, abstract = {Thyroid hormones play an important role in brain development, but the mechanism(s) by which triiodothyronine (T3) mediates neuronal differentiation is poorly understood. Here we demonstrate that T3 regulates the neurotrophic factor, neurotrophin-3 (NT-3), in developing rat cerebellar granule cells both in cell culture and in vivo. In situ hybridization experiments showed that developing Purkinje cells do not express NT-3 mRNA but do express trkC, the putative neuronal receptor for NT-3. Addition of recombinant NT-3 to cerebellar cultures from embryonic rat brain induces hypertrophy and neurite sprouting of Purkinje cells, and upregulates the mRNA encoding the calcium-binding protein, calbindin-28 kD. The present study demonstrates a novel interaction between cerebellar granule neurons and developing Purkinje cells in which NT-3 induced by T3 in the granule cells promotes Purkinje cell differentiation.}, author = {Lindholm, Dan and Castrén, Eero and Tsoulfas, Pantelis and Kolbeck, Roland and Berzaghi, Maria and Leingärtner, Axel and Heisenberg, Carl-Philipp J and Tesarollo, Lino and Parada, Luis and Thoenen, Hans}, issn = {0021-9525}, journal = {Journal of Cell Biology}, number = {2}, pages = {443 -- 450}, publisher = {Rockefeller University Press}, title = {{Neurotrophin-3 induced by tri-iodothyronine in cerebellar granule cells promotes Purkinje cell differentiation}}, doi = {10.1083/jcb.122.2.443}, volume = {122}, year = {1993}, }