scholarly journals Dynein efficiently navigates the dendritic cytoskeleton to drive the retrograde trafficking of BDNF/TrkB signaling endosomes

2017 ◽  
Vol 28 (19) ◽  
pp. 2543-2554 ◽  
Author(s):  
Swathi Ayloo ◽  
Pedro Guedes-Dias ◽  
Amy E. Ghiretti ◽  
Erika L. F. Holzbaur
Keyword(s):  
Real Time ◽  
Axonal Transport ◽  
Hippocampal Neurons ◽  
Cytoplasmic Dynein ◽  
Microtubule Dynamics ◽  
Neuronal Function ◽  
Retrograde Trafficking ◽  
Basic Understanding ◽  
Dependent Transport ◽  
Dynamic Microtubule

The efficient transport of cargoes within axons and dendrites is critical for neuronal function. Although we have a basic understanding of axonal transport, much less is known about transport in dendrites. We used an optogenetic approach to recruit motor proteins to cargo in real time within axons or dendrites in hippocampal neurons. Kinesin-1, a robust axonal motor, moves cargo less efficiently in dendrites. In contrast, cytoplasmic dynein efficiently navigates both axons and dendrites; in both compartments, dynamic microtubule plus ends enhance dynein-dependent transport. To test the predictions of the optogenetic assay, we examined the contribution of dynein to the motility of an endogenous dendritic cargo and found that dynein inhibition eliminates the retrograde bias of BDNF/TrkB trafficking. However, inhibition of microtubule dynamics has no effect on BDNF/TrkB motility, suggesting that dendritic kinesin motors may cooperate with dynein to drive the transport of signaling endosomes into the soma. Collectively our data highlight compartment-specific differences in kinesin activity that likely reflect specialized tuning for localized cytoskeletal determinants, whereas dynein activity is less compartment specific but is more responsive to changes in microtubule dynamics.

Microtubule Dynamics and Organelle Transport in Reticulomyxa

1990 ◽  
Vol 48 (3) ◽  
pp. 194-195
Author(s):  
Yih-Tai Chen ◽  
Ursula Euteneuer ◽  
Ken B. Johnson ◽  
Michael P. Koonce ◽  
Manfred Schliwa
Keyword(s):  
Plasma Membrane ◽  
Light Microscopy ◽  
Cytoplasmic Dynein ◽  
Living Cells ◽  
Microtubule Dynamics ◽  
Model Systems ◽  
Organelle Transport ◽  
Biochemical Characteristics ◽  
Dependent Transport

The application of video techniques to light microscopy and the development of motility assays in reactivated or reconstituted model systems rapidly advanced our understanding of the mechanism of organelle transport and microtubule dynamics in living cells. Two microtubule-based motors have been identified that are good candidates for motors that drive organelle transport: kinesin, a plus end-directed motor, and cytoplasmic dynein, which is minus end-directed. However, the evidence that they do in fact function as organelle motors is still indirect.We are studying microtubule-dependent transport and dynamics in the giant amoeba, Reticulomyxa. This cell extends filamentous strands backed by an extensive array of microtubules along which organelles move bidirectionally at up to 20 μm/sec (Fig. 1). Following removal of the plasma membrane with a mild detergent, organelle transport can be reactivated by the addition of ATP (1). The physiological, pharmacological and biochemical characteristics show the motor to be a cytoplasmic form of dynein (2).

scholarly journals Dynein activator Hook1 is required for trafficking of BDNF-signaling endosomes in neurons

2018 ◽  
Vol 218 (1) ◽  
pp. 220-233 ◽  
Author(s):  
Mara A. Olenick ◽  
Roberto Dominguez ◽  
Erika L.F. Holzbaur
Keyword(s):  
Axonal Transport ◽  
Hippocampal Neurons ◽  
Neuronal Development ◽  
Molecular Motor ◽  
Cytoplasmic Dynein ◽  
Differential Association ◽  
Primary Hippocampal Neurons ◽  
Bdnf Signaling ◽  
Microfluidic Chambers

Axonal transport is required for neuronal development and survival. Transport from the axon to the soma is driven by the molecular motor cytoplasmic dynein, yet it remains unclear how dynein is spatially and temporally regulated. We find that the dynein effector Hook1 mediates transport of TrkB–BDNF-signaling endosomes in primary hippocampal neurons. Hook1 comigrates with a subpopulation of Rab5 endosomes positive for TrkB and BDNF, which exhibit processive retrograde motility with faster velocities than the overall Rab5 population. Knockdown of Hook1 significantly reduced the motility of BDNF-signaling endosomes without affecting the motility of other organelles. In microfluidic chambers, Hook1 depletion resulted in a significant decrease in the flux and processivity of BDNF-Qdots along the mid-axon, an effect specific for Hook1 but not Hook3. Hook1 depletion inhibited BDNF trafficking to the soma and blocked downstream BDNF- and TrkB-dependent signaling to the nucleus. Together, these studies support a model in which differential association with cargo-specific effectors efficiently regulates dynein in neurons.

Spastin Interacts with CRMP2 to Regulate Neurite Outgrowth by Controlling Microtubule Dynamics through Phosphorylation Modifications

2020 ◽  
Vol 19 ◽  
Author(s):  
Sumei Li ◽  
Jifeng Zhang ◽  
Jiaqi Zhang ◽  
Jiong Li ◽  
Longfei Cheng ◽  
...  
Keyword(s):  
Neurite Outgrowth ◽  
Hippocampal Neurons ◽  
Neuronal Development ◽  
Phosphorylation Site ◽  
Microtubule Dynamics ◽  
Microtubule Assembly ◽  
C Terminus ◽  
Mediator Protein ◽  
Branch Formation

Aims: Our work aims to revealing the underlying microtubule mechanism of neurites outgrowth during neuronal development, and also proposes a feasible intervention pathway for reconstructing neural network connections after nerve injury. Background: Microtubule polymerization and severing are the basis for the neurite outgrowth and branch formation. Collapsin response mediator protein 2 (CRMP2) regulates axonal growth and branching as a binding partner of the tubulin heterodimer to promote microtubule assembly. And spastin participates in the growth and regeneration of neurites by severing microtubules into small segments. However, how CRMP2 and spastin cooperate to regulate neurite outgrowth by controlling the microtubule dynamics needs to be elucidated. Objective: To explore whether neurite outgrowth was mediated by coordination of CRMP2 and spastin. Method: Hippocampal neurons were cultured in vitro in 24-well culture plates for 4 days before being used to perform the transfection. Calcium phosphate was used to transfect the CRMP2 and spastin constructs and their control into the neurons. An interaction between CRMP2 and spastin was examined by using pull down, CoIP and immunofluorescence colocalization assays. And immunostaining was also performed to determine the morphology of neurites. Result: We first demonstrated that CRMP2 interacted with spastin to promote the neurite outgrowth and branch formation. Furthermore, our results identified that phosphorylation modification failed to alter the binding affinities of CRMP2 for spastin, but inhibited their binding to microtubules. CRMP2 interacted with the MTBD domain of spastin via its C-terminus, and blocking the binding sites of them inhibited the outgrowth and branch formation of neurites. In addition, we confirmed one phosphorylation site S210 at spastin in hippocampal neurons and phosphorylation spastin at site S210 promoted the neurite outgrowth but not branch formation by remodeling microtubules. Conclusion: Taken together, our data demonstrated that the interaction of CRMP2 and spastin is required for neurite outgrowth and branch formation and their interaction is not regulated by their phosphorylation.

scholarly journals Identification and developmental regulation of a neuron-specific subunit of cytoplasmic dynein.

1996 ◽  
Vol 7 (2) ◽  
pp. 331-343 ◽  
Author(s):  
K K Pfister ◽  
M W Salata ◽  
J F Dillman ◽  
E Torre ◽  
R J Lye
Keyword(s):  
Axonal Transport ◽  
Developmental Regulation ◽  
Cytoplasmic Dynein ◽  
Retrograde Axonal Transport ◽  
Protein Isoforms ◽  
Cultured Neurons ◽  
Developmentally Regulated ◽  
Time Period ◽  
Intermediate Chains

Cytoplasmic dynein is the microtubule minus-end-directed motor for the retrograde axonal transport of membranous organelles. Because of its similarity to the intermediate chains of flagellar dynein, the 74-kDa intermediate chain (IC74) subunit of dynein is thought to be involved in binding dynein to its membranous organelle cargo. Previously, we identified six isoforms of the IC74 cytoplasmic dynein subunit in the brain. We further demonstrated that cultured glia and neurons expressed different dynein IC74 isoforms and phospho-isoforms. Two isoforms were observed when dynein from glia was analyzed. When dynein from cultured neurons was analyzed, six IC74 isoforms were observed, although the relative amounts of the dynein isoforms from cultured neurons differed from those found in dynein from brain. To better understand the role of the neuronal IC74 isoforms and identify neuron-specific IC74 dynein subunits, the expression of the IC74 protein isoforms and mRNAs of various tissues were compared. As a result of this comparison, the identity of each of the isoform spots observed on two-dimensional gels was correlated with the products of each of the IC74 mRNAs. We also found that between the fifteenth day of gestation (E15) and the fifth day after birth (P5), the relative expression of the IC74 protein isoforms changes, demonstrating that the expression of IC74 isoforms is developmentally regulated in brain. During this time period, there is relatively little change in the abundance of the various IC74 mRNAs. The E15 to P5 time period is one of rapid process extension and initial pattern formation in the rat brain. This result indicates that the changes in neuronal IC74 isoforms coincide with neuronal differentiation, in particular the extension of processes. This suggests a role for the neuronal IC74 isoforms in the establishment or regulation of retrograde axonal transport.

scholarly journals Stabilization of dynamic microtubules by mDia1 drives Tau-dependent Aβ1–42 synaptotoxicity

2017 ◽  
Vol 216 (10) ◽  
pp. 3161-3178 ◽  
Author(s):  
Xiaoyi Qu ◽  
Feng Ning Yuan ◽  
Carlo Corona ◽  
Silvia Pasini ◽  
Maria Elena Pero ◽  
...  
Keyword(s):  
Alzheimer’S Disease ◽  
Alzheimer's Disease ◽  
Axonal Transport ◽  
Dendritic Spines ◽  
Posttranslational Modifications ◽  
Hippocampal Neurons ◽  
Neuronal Damage ◽  
Driving Factors ◽  
Hyperphosphorylated Tau ◽  
Tau Hyperphosphorylation

Oligomeric Amyloid β1–42 (Aβ) plays a crucial synaptotoxic role in Alzheimer’s disease, and hyperphosphorylated tau facilitates Aβ toxicity. The link between Aβ and tau, however, remains controversial. In this study, we find that in hippocampal neurons, Aβ acutely induces tubulin posttranslational modifications (PTMs) and stabilizes dynamic microtubules (MTs) by reducing their catastrophe frequency. Silencing or acute inhibition of the formin mDia1 suppresses these activities and corrects the synaptotoxicity and deficits of axonal transport induced by Aβ. We explored the mechanism of rescue and found that stabilization of dynamic MTs promotes tau-dependent loss of dendritic spines and tau hyperphosphorylation. Collectively, these results uncover a novel role for mDia1 in Aβ-mediated synaptotoxicity and demonstrate that inhibition of MT dynamics and accumulation of PTMs are driving factors for the induction of tau-mediated neuronal damage.

scholarly journals ATAT1-enriched vesicles promote microtubule acetylation via axonal transport

2019 ◽  
Author(s):  
Aviel Even ◽  
Giovanni Morelli ◽  
Chiara Scaramuzzino ◽  
Ivan Gladwyn-Ng ◽  
Romain Le Bail ◽  
...  
Keyword(s):  
Axonal Transport ◽  
Vesicular Transport ◽  
Microtubule Dynamics ◽  
Tubulin Acetylation ◽  
Cytosolic Side ◽  
Neuronal Vesicles ◽  
Β Tubulin

Microtubules are polymerized dimers of α- and β-tubulin that underlie a broad range of cellular activities. Acetylation of α-tubulin by the acetyl-transferase ATAT1 modulates microtubule dynamics and functions in neurons. However, it remains unclear how and why this enzyme acetylates microtubules over long distances in axons. Here, we show that loss of ATAT1 impairs axonal transport in neurons and cell free motility assays confirm a requirement of tubulin acetylation for proper bidirectional vesicular transport. Moreover, we demonstrate that the main cellular pool of ATAT1 is transported at the cytosolic side of neuronal vesicles that are moving along axons. Altogether, our data suggest that axonal transport of ATAT1-enriched vesicles is the predominant driver of α-tubulin acetylation in axons.

scholarly journals C9orf72-derived arginine-containing dipeptide repeats associate with axonal transport machinery and impede microtubule-based motility

2019 ◽  
Author(s):  
Laura Fumagalli ◽  
Florence L. Young ◽  
Steven Boeynaems ◽  
Mathias De Decker ◽  
Arpan R. Mehta ◽  
...  
Keyword(s):  
Axonal Transport ◽  
Motor Neurons ◽  
Single Molecule ◽  
Induced Pluripotent Stem Cell ◽  
Cytoplasmic Dynein ◽  
Inhibitory Interaction ◽  
Hexanucleotide Repeat ◽  
Repeat Expansions

ABSTRACTHexanucleotide repeat expansions in the C9orf72 gene are the most common genetic cause of amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). How this mutation leads to these neurodegenerative diseases remains unclear. Here, we use human induced pluripotent stem cell-derived motor neurons to show that C9orf72 repeat expansions impair microtubule-based transport of mitochondria, a process critical for maintenance of neuronal function. Cargo transport defects are recapitulated by treating healthy neurons with the arginine-rich dipeptide repeat proteins (DPRs) that are produced by the hexanucleotide repeat expansions. Single-molecule imaging shows that these DPRs perturb motility of purified kinesin-1 and cytoplasmic dynein-1 motors along microtubules in vitro. Additional in vitro and in vivo data indicate that the DPRs impair transport by interacting with both microtubules and the motor complexes. We also show that kinesin-1 is enriched in DPR inclusions in patient brains and that increasing the level of this motor strongly suppresses the toxic effects of arginine-rich DPR expression in a Drosophila model. Collectively, our study implicates an inhibitory interaction of arginine-rich DPRs with the axonal transport machinery in C9orf72-associated ALS/FTD and thereby points to novel potential therapeutic strategies.

Real time imaging reveals a peroxisomal reticulum in living cells

2000 ◽  
Vol 113 (20) ◽  
pp. 3663-3671 ◽  
Author(s):  
M. Schrader ◽  
S.J. King ◽  
T.A. Stroh ◽  
T.A. Schroer
Keyword(s):  
Real Time ◽  
Cytoplasmic Dynein ◽  
Living Cells ◽  
Real Time Imaging ◽  
Peroxisomal Targeting ◽  
Microtubule Motor ◽  
Targeting Signal ◽  
Peroxisomal Targeting Signal

We have directly imaged the dynamic behavior of a variety of morphologically different peroxisomal structures in HepG2 and COS-7 cells transfected with a construct encoding GFP bearing the C-terminal peroxisomal targeting signal 1. Real time imaging revealed that moving peroxisomes interacted with each other and were engaged in transient contacts, and at higher magnification, tubular peroxisomes appeared to form a peroxisomal reticulum. Local remodeling of these structures could be observed involving the formation and detachment of tubular processes that interconnected adjacent organelles. Inhibition of cytoplasmic dynein based motility by overexpression of the dynactin subunit, dynamitin (p50), inhibited the movement of peroxisomes in vivo and interfered with the reestablishment of a uniform distribution of peroxisomes after recovery from nocodazole treatment. Isolated peroxisomes moved in vitro along microtubules in the presence of a microtubule motor fraction. Our data reveal that peroxisomal behavior in vivo is significantly more dynamic and interactive than previously thought and suggest a role for the dynein/dynactin motor in peroxisome motility.

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