scholarly journals Growth Cone-Localized Microtubule Organizing Center Establishes Microtubule Orientation in Dendrites

2019 ◽  
Author(s):  
Xing Liang ◽  
Marcela Kokes ◽  
Richard Fetter ◽  
Melissa A. Pickett ◽  
Maria D. Sallee ◽  
...  
Keyword(s):  
Growth Cone ◽  
Membrane Trafficking ◽  
Dendritic Growth ◽  
Microtubule Organizing Center ◽  
Microtubule Nucleation ◽  
Recycling Endosomes ◽  
Microtubule Orientation ◽  
Microtubule Polarity ◽  
Organizing Center ◽  
Subcellular Compartmentalization

AbstractA polarized arrangement of neuronal microtubule arrays is the foundation of membrane trafficking and subcellular compartmentalization. Conserved among both invertebrates and vertebrates, axons contain exclusively “plus-end-out” microtubules while dendrites contain a high percentage of “minus-end-out” microtubules, the origins of which have been a mystery. Here we show that the dendritic growth cone contains a non-centrosomal microtubule organizing center (ncMTOC), which generates minus-end-out microtubules along outgrowing dendrites and plus-end-out microtubules in the growth cone. RAB-11-positive recycling endosomes accumulate in this region and are responsible for localizing the microtubule nucleation complex γ-TuRC. The MTOC tracks the extending growth cone by kinesin-1/UNC-116-mediated endosome movements on distal plus-end-out microtubules and dynein-mediated endosome clustering near MTOC. Critically, perturbation of the function or localization of the MTOC causes reversed microtubule polarity in dendrites. These findings unveil the dendritic MTOC as a critical organelle for establishing axon-dendrite polarity.

scholarly journals Growth cone-localized microtubule organizing center establishes microtubule orientation in dendrites

eLife ◽  
2020 ◽  
Vol 9 ◽  
Author(s):  
Xing Liang ◽  
Marcela Kokes ◽  
Richard D Fetter ◽  
Maria Danielle Sallee ◽  
Adrian W Moore ◽  
...  
Keyword(s):  
Caenorhabditis Elegans ◽  
Growth Cone ◽  
Membrane Trafficking ◽  
Dendritic Growth ◽  
Microtubule Organizing Center ◽  
Microtubule Nucleation ◽  
Microtubule Orientation ◽  
Microtubule Polarity ◽  
Organizing Center ◽  
Subcellular Compartmentalization

A polarized arrangement of neuronal microtubule arrays is the foundation of membrane trafficking and subcellular compartmentalization. Conserved among both invertebrates and vertebrates, axons contain exclusively ‘plus-end-out’ microtubules while dendrites contain a high percentage of ‘minus-end-out’ microtubules, the origins of which have been a mystery. Here we show that in Caenorhabditis elegans the dendritic growth cone contains a non-centrosomal microtubule organizing center (MTOC), which generates minus-end-out microtubules along outgrowing dendrites and plus-end-out microtubules in the growth cone. RAB-11-positive endosomes accumulate in this region and co-migrate with the microtubule nucleation complex γ-TuRC. The MTOC tracks the extending growth cone by kinesin-1/UNC-116-mediated endosome movements on distal plus-end-out microtubules and dynein clusters this advancing MTOC. Critically, perturbation of the function or localization of the MTOC causes reversed microtubule polarity in dendrites. These findings unveil the endosome-localized dendritic MTOC as a critical organelle for establishing axon-dendrite polarity.

scholarly journals Author response: Growth cone-localized microtubule organizing center establishes microtubule orientation in dendrites

2020 ◽  
Author(s):  
Xing Liang ◽  
Marcela Kokes ◽  
Richard D Fetter ◽  
Maria Danielle Sallee ◽  
Adrian W Moore ◽  
...  
Keyword(s):  
Growth Cone ◽  
Author Response ◽  
Microtubule Organizing Center ◽  
Microtubule Orientation ◽  
Organizing Center

scholarly journals Growth Cone-Localized Microtubule Organizing Center Establishes Microtubule Orientation in Dendrites

2020 ◽  
Author(s):  
Xing Liang ◽  
Marcela Kokes ◽  
Richard Fetter ◽  
Maria D. Sallee ◽  
Adrian W. Moore ◽  
...  
Keyword(s):  
Growth Cone ◽  
Microtubule Organizing Center ◽  
Microtubule Orientation ◽  
Organizing Center

scholarly journals PTRN-1/CAMSAP and NOCA-2/NINEIN are required for microtubule polarity in Caenorhabditis elegans dendrites

2021 ◽  
Author(s):  
liu he ◽  
Lotte van Beem ◽  
Casper Hoogenraad ◽  
Martin Harterink
Keyword(s):  
Caenorhabditis Elegans ◽  
Dendritic Growth ◽  
Double Mutant ◽  
Microtubule Organizing Center ◽  
Microtubule Organization ◽  
Microtubule Stabilization ◽  
Microtubule Polarity ◽  
Organizing Center ◽  
Related Proteins

The neuronal microtubule cytoskeleton is key to establish axon-dendrite polarity. Dendrites are characterized by the presence of minus-end out microtubules, however the mechanisms that organize these microtubules minus-end out is still poorly understood. Here, we characterized the role of two microtubule minus-end related proteins in this process in Caenorhabditis elegans, the microtubule minus-end stabilizing protein CAMSAP (PTRN-1) and a NINEIN homologue (NOCA-2). We found that CAMSAP and NINEIN function in parallel to mediate microtubule organization in dendrites. During dendrite outgrowth, RAB-11 positive vesicles localized to the dendrite tip function as a microtubule organizing center (MTOC) to nucleate microtubules. In the absence of either CAMSAP or NINEIN, we observed a low penetrance MTOC vesicles mis-localization to the cell body, and a nearly fully penetrant phenotype in double mutant animals. This suggests that both proteins are important for localizing the MTOC vesicles to the growing dendrite tip to organize microtubules minus-end out. Whereas NINEIN localizes to the MTOC vesicles where it is important for the recruitment of the microtubule nucleator ?-tubulin, CAMSAP localizes around the MTOC vesicles and is co-translocated forward with the MTOC vesicles upon dendritic growth. Together, these results indicate that microtubule nucleation from the MTOC vesicles and microtubule stabilization are both important to localize the MTOC vesicles distally to organize dendritic microtubules minus-end out.

scholarly journals Mobility, Microtubule Nucleation and Structure of Microtubule-organizing Centers in Multinucleated Hyphae ofAshbya gossypii

2010 ◽  
Vol 21 (1) ◽  
pp. 18-28 ◽  
Author(s):  
Claudia Lang ◽  
Sandrine Grava ◽  
Tineke van den Hoorn ◽  
Rhonda Trimble ◽  
Peter Philippsen ◽  
...  
Keyword(s):  
Spindle Pole Body ◽  
Hyphal Growth ◽  
Spindle Pole ◽  
Microtubule Organizing Center ◽  
Microtubule Nucleation ◽  
Cytoplasmic Microtubule ◽  
Organizing Center ◽  
Cytoplasmic Side ◽  
Independent Mode

We investigated the migration of multiple nuclei in hyphae of the filamentous fungus Ashbya gossypii. Three types of cytoplasmic microtubule (cMT)-dependent nuclear movements were characterized using live cell imaging: short-range oscillations (up to 4.5 μm/min), rotations (up to 180° in 30 s), and long-range nuclear bypassing (up to 9 μm/min). These movements were superimposed on a cMT-independent mode of nuclear migration, cotransport with the cytoplasmic stream. This latter mode is sufficient to support wild-type-like hyphal growth speeds. cMT-dependent nuclear movements were led by a nuclear-associated microtubule-organizing center, the spindle pole body (SPB), which is the sole site of microtubule nucleation in A. gossypii. Analysis of A. gossypii SPBs by electron microscopy revealed an overall laminar structure similar to the budding yeast SPB but with distinct differences at the cytoplasmic side. Up to six perpendicular and tangential cMTs emanated from a more spherical outer plaque. The perpendicular and tangential cMTs most likely correspond to short, often cortex-associated cMTs and to long, hyphal growth-axis–oriented cMTs, respectively, seen by in vivo imaging. Each SPB nucleates its own array of cMTs, and the lack of overlapping cMT arrays between neighboring nuclei explains the autonomous nuclear oscillations and bypassing observed in A. gossypii hyphae.

scholarly journals Centriole and Golgi microtubule nucleation are dispensable for the migration of human neutrophil-like cells

2021 ◽  
pp. mbc.E21-02-0060
Author(s):  
Lucas C. Klemm ◽  
Ryan A. Denu ◽  
Laurel E. Hind ◽  
Briana L. Rocha-Gregg ◽  
Mark E. Burkard ◽  
...  
Keyword(s):  
Cell Line ◽  
Host Defense ◽  
Human Neutrophil ◽  
Microtubule Organizing Center ◽  
Microtubule Nucleation ◽  
Directed Migration ◽  
Inhibitory Role ◽  
Organizing Center ◽  
Cytoskeletal Rearrangements

Neutrophils migrate in response to chemoattractants to mediate host defense. Chemoattractants drive rapid intracellular cytoskeletal rearrangements including the radiation of microtubules from the microtubule-organizing center (MTOC) towards the rear of polarized neutrophils. Microtubules regulate neutrophil polarity and motility, but little is known about the specific role of MTOCs. To characterize the role of MTOCs on neutrophil motility we depleted centrioles in a well-established neutrophil-like cell line. Surprisingly, both chemical and genetic centriole depletion increased neutrophil speed and chemotactic motility, suggesting an inhibitory role for centrioles during directed migration. We also found that depletion of both centrioles and GM130-mediated Golgi microtubule nucleation did not impair neutrophil directed migration. Taken together, our findings demonstrate an inhibitory role for centrioles and a resilient MTOC system in motile human neutrophil-like cells. [Media: see text] [Media: see text] [Media: see text]

The small GTP-binding protein rab6p is redistributed in the cytosol by brefeldin A

1993 ◽  
Vol 106 (3) ◽  
pp. 789-802 ◽  
Author(s):  
M. Roa ◽  
V. Cornet ◽  
C.Z. Yang ◽  
B. Goud
Keyword(s):  
Membrane Trafficking ◽  
Mammalian Cells ◽  
Brefeldin A ◽  
Atp Depletion ◽  
Cell Fractionation ◽  
Microtubule Organizing Center ◽  
Trans Golgi Network ◽  
Gtp Binding ◽  
Organizing Center ◽  
Golgi Network

Rab6 protein belongs to the Sec4/Ypt/rab subfamily of small GTP-binding proteins involved in intracellular membrane trafficking in yeast and mammalian cells. Its localization both in medial and trans-Golgi network prompted us to study the effects of brefeldin A (BFA) on rab6p redistribution. By two techniques, indirect immunofluorescence and cell fractionation, we investigated the fate of rab6p and compared it to other Golgi or trans-Golgi network markers in BHK-21 and NIH-3T3 cells. BFA, at 5 micrograms/ml, induced redistribution of rab6p according to a biphasic process: during the first 10–15 minutes, tubulo-vesicular structures--colabelled with a bona fide medial Golgi marker called CTR 433--were observed; these structures were then replaced by punctate diffuse staining, which was stable for up to 3 hours. The 110 kDa peripheral membrane protein beta-COP was released much more rapidly from the Golgi membranes, whereas the trans-Golgi network marker TGN 38 relocated to the microtubule organizing center. The kinetics of reversion of BFA action on these antigens was also followed by immunofluorescence. Consistent with these results, rab6 antigen, originally found as 40% in the cytosolic versus 60% in the particulate (P 150,000 g) fraction, became almost entirely cytosolic; moreover, it partitioned in the aqueous phase of Triton X-114 whereas the membrane fraction was detergent-soluble. Rab6p did not become part of the coatomers after its BFA-induced release from Golgi structures. Three requirements seemed to be necessary for such a release: integrity of the microtubules, presence of energy, and a hypothetical trimeric G protein, as revealed by the respective roles of nocodazole, ATP depletion, and sensitivity to aluminium fluoride. Finally, we have shown that BFA does not prevent attachment of newly synthesized rab6p to membranes.

Localization of myosin Va is dependent on the cytoskeletal organization in the cell

2001 ◽  
Vol 79 (1) ◽  
pp. 93-106 ◽  
Author(s):  
Corinne Lionne ◽  
Folma Buss ◽  
Tony Hodge ◽  
Gudrun Ihrke ◽  
John Kendrick-Jones
Keyword(s):  
Membrane Trafficking ◽  
Molecular Motors ◽  
Leading Edge ◽  
Apical Surface ◽  
Myosin V ◽  
Microtubule Organizing Center ◽  
Organizing Center ◽  
Myosin Va ◽  
Dynamic Membranes ◽  
In Cells

Myosin V plays an important role in membrane trafficking events. Its implication in the transport of pigment granules in melanocytes and synaptic vesicles in neurons is now well established. However, less is known about its function(s) in other cell types. Finding a common function is complicated by the diversity of myosin V expression in different tissues and organisms and by its association with different subcellular compartments. Here we show that myosin V is present in a variety of cells. Within the same cell type under different physiological conditions, we observed two main cellular locations for myosin V that were dependent on the dynamics of the plasma membrane: in cells with highly dynamic membranes, myosin V was specifically concentrated at the leading edge in membrane ruffles, whereas in cells with less dynamic membranes, myosin V was enriched around the microtubule-organizing center. The presence of myosin V in the leading ruffling edge of the cell was induced by growth factor stimulation and was dependent on the presence of a functional motor domain. Moreover, myosin V localization at the microtubule-organizing center was dependent on the integrity of the microtubules. In polarized epithelial cells (WIF-B), where the microtubule-organizing region is close to the actin-rich apical surface, one single pool of myosin V, sensitive to the integrity of both microtubules and actin filaments, was observed.Key words: cell motility, cytoskeleton dynamics, molecular motors, mouse brain unconventional myosin Va, ruffles.

scholarly journals Golgi Outposts Locally Regulate Microtubule Orientation in Neurons but Are Not Required for the Overall Polarity of the Dendritic Cytoskeleton

Genetics ◽  
2020 ◽  
Vol 215 (2) ◽  
pp. 435-447 ◽  
Author(s):  
Sihui Z. Yang ◽  
Jill Wildonger
Keyword(s):  
Matrix Protein ◽  
Cell Function ◽  
Microtubule Nucleation ◽  
Microtubule Organization ◽  
Microtubule Orientation ◽  
Microtubule Polarity ◽  
Ring Complex ◽  
Microtubule Growth ◽  
Microtubule Networks ◽  
Golgi Matrix

Microtubule-organizing centers often play a central role in organizing the cellular microtubule networks that underlie cell function. In neurons, microtubules in axons and dendrites have distinct polarities. Dendrite-specific Golgi “outposts,” in particular multicompartment outposts, have emerged as regulators of acentrosomal microtubule growth, raising the question of whether outposts contribute to establishing or maintaining the overall polarity of the dendritic microtubule cytoskeleton. Using a combination of genetic approaches and live imaging in a Drosophila model, we found that dendritic microtubule polarity is unaffected by eliminating known regulators of Golgi-dependent microtubule organization including the cis-Golgi matrix protein GM130, the fly AKAP450 ortholog pericentrin-like protein, and centrosomin. This indicates that Golgi outposts are not essential for the formation or maintenance of a dendrite-specific cytoskeleton. However, the overexpression of GM130, which promotes the formation of ectopic multicompartment units, is sufficient to alter dendritic microtubule polarity. Axonal microtubule polarity is similarly disrupted by the presence of ectopic multicompartment Golgi outposts. Notably, multicompartment outposts alter microtubule polarity independently of microtubule nucleation mediated by the γ-tubulin ring complex. Thus, although Golgi outposts are not essential to dendritic microtubule polarity, altering their organization correlates with changes to microtubule polarity. Based on these data, we propose that the organization of Golgi outposts is carefully regulated to ensure proper dendritic microtubule polarity.

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