Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 59
Filtrar
1.
PLoS Genet ; 20(8): e1011388, 2024 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-39186815

RESUMO

Most neurons are not replaced after injury and thus possess robust intrinsic mechanisms for repair after damage. Axon injury triggers a calcium wave, and calcium and cAMP can augment axon regeneration. In comparison to axon regeneration, dendrite regeneration is poorly understood. To test whether calcium and cAMP might also be involved in dendrite injury signaling, we tracked the responses of Drosophila dendritic arborization neurons to laser severing of axons and dendrites. We found that calcium and subsequently cAMP accumulate in the cell body after both dendrite and axon injury. Two voltage-gated calcium channels (VGCCs), L-Type and T-Type, are required for the calcium influx in response to dendrite injury and play a role in rapid initiation of dendrite regeneration. The AC8 family adenylyl cyclase, Ac78C, is required for cAMP production after dendrite injury and timely initiation of regeneration. Injury-induced cAMP production is sensitive to VGCC reduction, placing calcium upstream of cAMP generation. We propose that two VGCCs initiate global calcium influx in response to dendrite injury followed by production of cAMP by Ac78C. This signaling pathway promotes timely initiation of dendrite regrowth several hours after dendrite damage.


Assuntos
Adenilil Ciclases , Canais de Cálcio Tipo L , Cálcio , AMP Cíclico , Dendritos , Animais , Adenilil Ciclases/metabolismo , Adenilil Ciclases/genética , Axônios/metabolismo , Axônios/fisiologia , Cálcio/metabolismo , Canais de Cálcio/metabolismo , Canais de Cálcio/genética , Canais de Cálcio Tipo L/metabolismo , Canais de Cálcio Tipo L/genética , Canais de Cálcio Tipo T/metabolismo , Canais de Cálcio Tipo T/genética , Sinalização do Cálcio/genética , AMP Cíclico/metabolismo , Dendritos/metabolismo , Drosophila/genética , Drosophila melanogaster/genética , Proteínas de Drosophila/metabolismo , Proteínas de Drosophila/genética , Regeneração Nervosa/fisiologia , Regeneração Nervosa/genética , Neurônios/metabolismo , Regeneração/genética , Regeneração/fisiologia , Transdução de Sinais
2.
Dev Biol ; 497: 18-25, 2023 05.
Artigo em Inglês | MEDLINE | ID: mdl-36870669

RESUMO

Unlike many cell types, neurons are not typically replaced if damaged. Therefore, regeneration of damaged cellular domains is critical for maintenance of neuronal function. While axon regeneration has been documented for several hundred years, it has only recently become possible to determine whether neurons respond to dendrite removal with regeneration. Regrowth of dendrite arbors has been documented in invertebrate and vertebrate model systems, but whether it leads to functional restoration of a circuit remains unknown. To test whether dendrite regeneration restores function, we used larval Drosophila nociceptive neurons. Their dendrites detect noxious stimuli to initiate escape behavior. Previous studies of Drosophila sensory neurons have shown that dendrites of single neurons regrow after laser severing. We removed dendrites from 16 neurons per animal to clear most of the dorsal surface of nociceptive innervation. As expected, this reduced aversive responses to noxious touch. Surprisingly, behavior was completely restored 24 â€‹h after injury, at the stage when dendrite regeneration has begun, but the new arbor has only covered a small portion of its former territory. This behavioral recovery required regenerative outgrowth as it was eliminated in a genetic background in which new growth is blocked. We conclude that dendrite regeneration can restore behavior.


Assuntos
Axônios , Proteínas de Drosophila , Animais , Axônios/metabolismo , Dendritos/metabolismo , Regeneração Nervosa/fisiologia , Drosophila/metabolismo , Neurônios/metabolismo , Proteínas de Drosophila/genética , Proteínas de Drosophila/metabolismo
3.
Bull Math Biol ; 86(5): 58, 2024 Apr 16.
Artigo em Inglês | MEDLINE | ID: mdl-38627264

RESUMO

The microtubule cytoskeleton is responsible for sustained, long-range intracellular transport of mRNAs, proteins, and organelles in neurons. Neuronal microtubules must be stable enough to ensure reliable transport, but they also undergo dynamic instability, as their plus and minus ends continuously switch between growth and shrinking. This process allows for continuous rebuilding of the cytoskeleton and for flexibility in injury settings. Motivated by in vivo experimental data on microtubule behavior in Drosophila neurons, we propose a mathematical model of dendritic microtubule dynamics, with a focus on understanding microtubule length, velocity, and state-duration distributions. We find that limitations on microtubule growth phases are needed for realistic dynamics, but the type of limiting mechanism leads to qualitatively different responses to plausible experimental perturbations. We therefore propose and investigate two minimally-complex length-limiting factors: limitation due to resource (tubulin) constraints and limitation due to catastrophe of large-length microtubules. We combine simulations of a detailed stochastic model with steady-state analysis of a mean-field ordinary differential equations model to map out qualitatively distinct parameter regimes. This provides a basis for predicting changes in microtubule dynamics, tubulin allocation, and the turnover rate of tubulin within microtubules in different experimental environments.


Assuntos
Modelos Biológicos , Tubulina (Proteína) , Tubulina (Proteína)/metabolismo , Conceitos Matemáticos , Microtúbulos/metabolismo , Citoesqueleto
4.
Proc Natl Acad Sci U S A ; 118(4)2021 01 26.
Artigo em Inglês | MEDLINE | ID: mdl-33468672

RESUMO

The pathogenesis of chemotherapy-induced peripheral neuropathy (CIPN) is poorly understood. Here, we report that the CIPN-causing drug bortezomib (Bort) promotes delta 2 tubulin (D2) accumulation while affecting microtubule stability and dynamics in sensory neurons in vitro and in vivo and that the accumulation of D2 is predominant in unmyelinated fibers and a hallmark of bortezomib-induced peripheral neuropathy (BIPN) in humans. Furthermore, while D2 overexpression was sufficient to cause axonopathy and inhibit mitochondria motility, reduction of D2 levels alleviated both axonal degeneration and the loss of mitochondria motility induced by Bort. Together, our data demonstrate that Bort, a compound structurally unrelated to tubulin poisons, affects the tubulin cytoskeleton in sensory neurons in vitro, in vivo, and in human tissue, indicating that the pathogenic mechanisms of seemingly unrelated CIPN drugs may converge on tubulin damage. The results reveal a previously unrecognized pathogenic role for D2 in BIPN that may occur through altered regulation of mitochondria motility.


Assuntos
Bortezomib/efeitos adversos , Neoplasias/tratamento farmacológico , Doenças do Sistema Nervoso Periférico/genética , Tubulina (Proteína)/genética , Animais , Antineoplásicos/efeitos adversos , Axônios/efeitos dos fármacos , Axônios/patologia , Modelos Animais de Doenças , Drosophila melanogaster/genética , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Células HEK293 , Humanos , Larva/efeitos dos fármacos , Larva/genética , Microtúbulos/efeitos dos fármacos , Microtúbulos/genética , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/genética , Dinâmica Mitocondrial/efeitos dos fármacos , Dinâmica Mitocondrial/genética , Neoplasias/genética , Neoplasias/patologia , Doenças do Sistema Nervoso Periférico/induzido quimicamente , Doenças do Sistema Nervoso Periférico/patologia , Células Receptoras Sensoriais/efeitos dos fármacos , Células Receptoras Sensoriais/patologia , Peixe-Zebra/genética
5.
PLoS Genet ; 17(4): e1009112, 2021 04.
Artigo em Inglês | MEDLINE | ID: mdl-33819264

RESUMO

We previously identified a deletion on chromosome 16p12.1 that is mostly inherited and associated with multiple neurodevelopmental outcomes, where severely affected probands carried an excess of rare pathogenic variants compared to mildly affected carrier parents. We hypothesized that the 16p12.1 deletion sensitizes the genome for disease, while "second-hits" in the genetic background modulate the phenotypic trajectory. To test this model, we examined how neurodevelopmental defects conferred by knockdown of individual 16p12.1 homologs are modulated by simultaneous knockdown of homologs of "second-hit" genes in Drosophila melanogaster and Xenopus laevis. We observed that knockdown of 16p12.1 homologs affect multiple phenotypic domains, leading to delayed developmental timing, seizure susceptibility, brain alterations, abnormal dendrite and axonal morphology, and cellular proliferation defects. Compared to genes within the 16p11.2 deletion, which has higher de novo occurrence, 16p12.1 homologs were less likely to interact with each other in Drosophila models or a human brain-specific interaction network, suggesting that interactions with "second-hit" genes may confer higher impact towards neurodevelopmental phenotypes. Assessment of 212 pairwise interactions in Drosophila between 16p12.1 homologs and 76 homologs of patient-specific "second-hit" genes (such as ARID1B and CACNA1A), genes within neurodevelopmental pathways (such as PTEN and UBE3A), and transcriptomic targets (such as DSCAM and TRRAP) identified genetic interactions in 63% of the tested pairs. In 11 out of 15 families, patient-specific "second-hits" enhanced or suppressed the phenotypic effects of one or many 16p12.1 homologs in 32/96 pairwise combinations tested. In fact, homologs of SETD5 synergistically interacted with homologs of MOSMO in both Drosophila and X. laevis, leading to modified cellular and brain phenotypes, as well as axon outgrowth defects that were not observed with knockdown of either individual homolog. Our results suggest that several 16p12.1 genes sensitize the genome towards neurodevelopmental defects, and complex interactions with "second-hit" genes determine the ultimate phenotypic manifestation.


Assuntos
Encéfalo/metabolismo , Deleção Cromossômica , Cromossomos Humanos Par 16/genética , Transtornos do Neurodesenvolvimento/genética , Proteínas Adaptadoras de Transdução de Sinal/genética , Animais , Encéfalo/patologia , Canais de Cálcio/genética , Moléculas de Adesão Celular/genética , Proteínas de Ligação a DNA/genética , Modelos Animais de Doenças , Proteínas de Drosophila/genética , Drosophila melanogaster/genética , Epistasia Genética/genética , Regulação da Expressão Gênica no Desenvolvimento , Humanos , Metiltransferases/genética , Transtornos do Neurodesenvolvimento/patologia , Proteínas Nucleares/genética , PTEN Fosfo-Hidrolase/genética , Fatores de Transcrição/genética , Ubiquitina-Proteína Ligases/genética , Proteínas de Xenopus/genética , Xenopus laevis/genética
6.
J Neurosci ; 42(32): 6195-6210, 2022 08 10.
Artigo em Inglês | MEDLINE | ID: mdl-35840323

RESUMO

Mitogen-activated protein kinase kinase kinases (MAP3Ks) dual leucine kinase (DLK) and leucine zipper kinase (LZK) are essential mediators of axon damage responses, but their responses are varied, complex, and incompletely understood. To characterize their functions in axon injury, we generated zebrafish mutants of each gene, labeled motor neurons (MNs) and touch-sensing neurons in live zebrafish, precisely cut their axons with a laser, and assessed the ability of mutant axons to regenerate in larvae, before sex is apparent in zebrafish. DLK and LZK were required redundantly and cell autonomously for axon regeneration in MNs but not in larval Rohon-Beard (RB) or adult dorsal root ganglion (DRG) sensory neurons. Surprisingly, in dlk lzk double mutants, the spared branches of wounded RB axons grew excessively, suggesting that these kinases inhibit regenerative sprouting in damaged axons. Uninjured trigeminal sensory axons also grew excessively in mutants when neighboring neurons were ablated, indicating that these MAP3Ks are general inhibitors of sensory axon growth. These results demonstrate that zebrafish DLK and LZK promote diverse injury responses, depending on the neuronal cell identity and type of axonal injury.SIGNIFICANCE STATEMENT The MAP3Ks DLK and LZK are damage sensors that promote diverse outcomes to neuronal injury, including axon regeneration. Understanding their context-specific functions is a prerequisite to considering these kinases as therapeutic targets. To investigate DLK and LZK cell-type-specific functions, we created zebrafish mutants in each gene. Using mosaic cell labeling and precise laser injury we found that both proteins were required for axon regeneration in motor neurons but, unexpectedly, were not required for axon regeneration in Rohon-Beard or DRG sensory neurons and negatively regulated sprouting in the spared axons of touch-sensing neurons. These findings emphasize that animals have evolved distinct mechanisms to regulate injury site regeneration and collateral sprouting, and identify differential roles for DLK and LZK in these processes.


Assuntos
Axônios , Peixe-Zebra , Animais , Axônios/fisiologia , Leucina/metabolismo , Zíper de Leucina , MAP Quinase Quinase Quinases/genética , MAP Quinase Quinase Quinases/metabolismo , Neurônios Motores/metabolismo , Regeneração Nervosa/genética
7.
Dev Biol ; 483: 112-117, 2022 03.
Artigo em Inglês | MEDLINE | ID: mdl-35016908

RESUMO

The microtubule cytoskeleton is critical for maintenance of long and long-lived neurons. The overlapping array of microtubules extends from the major site of synthesis in the cell body to the far reaches of axons and dendrites. New materials are transported from the cell body along these neuronal roads by motor proteins, and building blocks and information about the state of affairs in other parts of the cell are returned by motors moving in the opposite direction. As motor proteins walk only in one direction along microtubules, the combination of correct motor and correctly oriented microtubules is essential for moving cargoes in the right direction. In this review, we focus on how microtubule polarity is established and maintained in neurons. At first thought, it seems that figuring out how microtubules are organized in neurons should be simple. After all, microtubules are essentially sticks with a slow-growing minus end and faster-growing plus end, and arranging sticks within the constrained narrow tubes of axons and dendrites should be straightforward. It is therefore quite surprising how many mechanisms contribute to making sure they are arranged in the correct polarity. Some of these mechanisms operate to generate plus-end-out polarity of axons, and others control mixed or minus-end-out dendrites.


Assuntos
Axônios/metabolismo , Polaridade Celular/fisiologia , Dendritos/metabolismo , Microtúbulos/metabolismo , Transdução de Sinais/fisiologia , Animais , Caenorhabditis elegans/crescimento & desenvolvimento , Caenorhabditis elegans/metabolismo , Citoesqueleto/metabolismo , Drosophila/crescimento & desenvolvimento , Drosophila/metabolismo , Cinesinas/metabolismo
8.
Dev Biol ; 488: 114-119, 2022 08.
Artigo em Inglês | MEDLINE | ID: mdl-35644253

RESUMO

Axon regeneration in response to injury has been documented in many animals over several hundred years. In contrast, how neurons respond to dendrite injury has been examined only in the last decade. So far, dendrite regeneration after injury has been documented in invertebrate model systems, but has not been assayed in a vertebrate. In this study, we use zebrafish motor neurons to track neurons after dendrite injury. We address two major gaps in our knowledge of dendrite regeneration: 1) whether post-synaptic dendrites can regenerate and 2) whether vertebrate dendrites can regenerate. We find that motor neurons survive laser microsurgery to remove one or all dendrites. Outgrowth of new dendrites typically initiated one to three days after injury, and a new, stable dendrite arbor was in place by five days after injury. We conclude that zebrafish motor neurons have the capacity to regenerate a new dendrite arbor.


Assuntos
Dendritos , Regeneração da Medula Espinal , Animais , Axônios , Dendritos/fisiologia , Neurônios Motores , Regeneração Nervosa/fisiologia , Medula Espinal , Peixe-Zebra
9.
Dev Biol ; 486: 56-70, 2022 06.
Artigo em Inglês | MEDLINE | ID: mdl-35341730

RESUMO

Many neurons in bilaterian animals are polarized with functionally distinct axons and dendrites. Microtubule polarity, microtubule stability, and the axon initial segment (AIS) have all been shown to influence polarized transport in neurons. Each of these cytoskeletal cues could act independently to control axon and dendrite identity, or there could be a hierarchy in which one acts upstream of the others. Here we test the hypothesis that microtubule polarity acts as a master regulator of neuronal polarity by using a Drosophila genetic background in which some dendrites have normal minus-end-out microtubule polarity and others have the axonal plus-end-out polarity. In these mosaic dendrite arbors, we found that ribosomes, which are more abundant in dendrites than axons, were reduced in plus-end-out dendrites, while an axonal cargo was increased. In addition, we determined that microtubule stability was different in plus-end-out and minus-end-out dendrites, with plus-end-out ones having more stable microtubules like axons. Similarly, we found that ectopic diffusion barriers, like those at the AIS, formed at the base of dendrites with plus-end-out regions. Thus, changes in microtubule polarity were sufficient to rearrange other cytoskeletal features associated with neuronal polarization. However, overall neuron shape was maintained with only subtle changes in branching in mosaic arbors. We conclude that microtubule polarity can act upstream of many aspects of intracellular neuronal polarization, but shape is relatively resilient to changes in microtubule polarity in vivo.


Assuntos
Polaridade Celular , Dendritos , Animais , Axônios/fisiologia , Polaridade Celular/fisiologia , Dendritos/fisiologia , Drosophila , Microtúbulos/fisiologia , Neurônios/fisiologia
10.
Dev Biol ; 492: 1-13, 2022 12.
Artigo em Inglês | MEDLINE | ID: mdl-36162553

RESUMO

The exocyst complex is an important regulator of intracellular trafficking and tethers secretory vesicles to the plasma membrane. Understanding of its role in neuron outgrowth remains incomplete, and previous studies have come to different conclusions about its importance for axon and dendrite growth, particularly in vivo. To investigate exocyst function in vivo we used Drosophila sensory neurons as a model system. To bypass early developmental requirements in other cell types, we used neuron-specific RNAi to target seven exocyst subunits. Initial neuronal development proceeded normally in these backgrounds, however, we considered this could be due to residual exocyst function. To probe neuronal growth capacity at later times after RNAi initiation, we used laser microsurgery to remove axons or dendrites and prompt regrowth. Exocyst subunit RNAi reduced axon regeneration, although new axons could be specified. In control neurons, a vesicle trafficking marker often concentrated in the new axon, but this pattern was disrupted in Sec6 RNAi neurons. Dendrite regeneration was also severely reduced by exocyst RNAi, even though the trafficking marker did not accumulate in a strongly polarized manner during normal dendrite regeneration. The requirement for the exocyst was not limited to injury contexts as exocyst subunit RNAi eliminated dendrite regrowth after developmental pruning. We conclude that the exocyst is required for injury-induced and developmental neurite outgrowth, but that residual protein function can easily mask this requirement.


Assuntos
Axônios , Exocitose , Exocitose/fisiologia , Neuritos , Regeneração Nervosa , Membrana Celular/metabolismo
11.
J Cell Sci ; 134(11)2021 06 01.
Artigo em Inglês | MEDLINE | ID: mdl-34096607

RESUMO

Axons and dendrites are distinguished by microtubule polarity. In Drosophila, dendrites are dominated by minus-end-out microtubules, whereas axons contain plus-end-out microtubules. Local nucleation in dendrites generates microtubules in both orientations. To understand why dendritic nucleation does not disrupt polarity, we used live imaging to analyze the fate of microtubules generated at branch points. We found that they had different rates of success exiting the branch based on orientation: correctly oriented minus-end-out microtubules succeeded in leaving about twice as often as incorrectly oriented microtubules. Increased success relied on other microtubules in a parallel orientation. From a candidate screen, we identified Trim9 and kinesin-5 (Klp61F) as machinery that promoted growth of new microtubules. In S2 cells, Eb1 recruited Trim9 to microtubules. Klp61F promoted microtubule growth in vitro and in vivo, and could recruit Trim9 in S2 cells. In summary, the data argue that Trim9 and kinesin-5 act together at microtubule plus ends to help polymerizing microtubules parallel to pre-existing ones resist catastrophe.


Assuntos
Polaridade Celular , Dendritos , Cinesinas/genética , Proteínas Associadas aos Microtúbulos/genética , Microtúbulos , Polimerização
12.
PLoS Biol ; 18(3): e3000657, 2020 03.
Artigo em Inglês | MEDLINE | ID: mdl-32163406

RESUMO

While many regulators of axon regeneration have been identified, very little is known about mechanisms that allow dendrites to regenerate after injury. Using a Drosophila model of dendrite regeneration, we performed a candidate screen of receptor tyrosine kinases (RTKs) and found a requirement for RTK-like orphan receptor (Ror). We confirmed that Ror was required for regeneration in two different neuron types using RNA interference (RNAi) and mutants. Ror was not required for axon regeneration or normal dendrite development, suggesting a specific role in dendrite regeneration. Ror can act as a Wnt coreceptor with frizzleds (fzs) in other contexts, so we tested the involvement of Wnt signaling proteins in dendrite regeneration. We found that knockdown of fz, dishevelled (dsh), Axin, and gilgamesh (gish) also reduced dendrite regeneration. Moreover, Ror was required to position dsh and Axin in dendrites. We recently found that Wnt signaling proteins, including dsh and Axin, localize microtubule nucleation machinery in dendrites. We therefore hypothesized that Ror may act by regulating microtubule nucleation at baseline and during dendrite regeneration. Consistent with this hypothesis, localization of the core nucleation protein γTubulin was reduced in Ror RNAi neurons, and this effect was strongest during dendrite regeneration. In addition, dendrite regeneration was sensitive to partial reduction of γTubulin. We conclude that Ror promotes dendrite regeneration as part of a Wnt signaling pathway that regulates dendritic microtubule nucleation.


Assuntos
Dendritos/fisiologia , Proteínas de Drosophila/metabolismo , Regeneração Nervosa/fisiologia , Receptores Órfãos Semelhantes a Receptor Tirosina Quinase/metabolismo , Animais , Drosophila , Proteínas de Drosophila/genética , Microtúbulos/genética , Microtúbulos/metabolismo , Mutação , Neurônios/fisiologia , Interferência de RNA , Receptores Órfãos Semelhantes a Receptor Tirosina Quinase/genética , Receptores Wnt/genética , Receptores Wnt/metabolismo , Via de Sinalização Wnt
13.
PLoS Biol ; 18(3): e3000647, 2020 03.
Artigo em Inglês | MEDLINE | ID: mdl-32163403

RESUMO

Dendrite microtubules are polarized with minus-end-out orientation in Drosophila neurons. Nucleation sites concentrate at dendrite branch points, but how they localize is not known. Using Drosophila, we found that canonical Wnt signaling proteins regulate localization of the core nucleation protein γTubulin (γTub). Reduction of frizzleds (fz), arrow (low-density lipoprotein receptor-related protein [LRP] 5/6), dishevelled (dsh), casein kinase Iγ, G proteins, and Axin reduced γTub-green fluorescent protein (GFP) at branch points, and two functional readouts of dendritic nucleation confirmed a role for Wnt signaling proteins. Both dsh and Axin localized to branch points, with dsh upstream of Axin. Moreover, tethering Axin to mitochondria was sufficient to recruit ectopic γTub-GFP and increase microtubule dynamics in dendrites. At dendrite branch points, Axin and dsh colocalized with early endosomal marker Rab5, and new microtubule growth initiated at puncta marked with fz, dsh, Axin, and Rab5. We propose that in dendrites, canonical Wnt signaling proteins are housed on early endosomes and recruit nucleation sites to branch points.


Assuntos
Dendritos/metabolismo , Proteínas de Drosophila/metabolismo , Endossomos/metabolismo , Microtúbulos/metabolismo , Proteínas Wnt/metabolismo , Animais , Complexo de Sinalização da Axina/genética , Complexo de Sinalização da Axina/metabolismo , Axônios/metabolismo , Polaridade Celular , Dendritos/genética , Drosophila , Proteínas de Drosophila/genética , Endossomos/genética , Microtúbulos/genética , Mutação , Receptores Wnt/genética , Receptores Wnt/metabolismo , Tubulina (Proteína)/genética , Tubulina (Proteína)/metabolismo , Proteínas Wnt/genética , Via de Sinalização Wnt/genética , Proteínas rab5 de Ligação ao GTP/genética , Proteínas rab5 de Ligação ao GTP/metabolismo
14.
J Exp Biol ; 226(12)2023 06 15.
Artigo em Inglês | MEDLINE | ID: mdl-37212026

RESUMO

Axon regeneration helps maintain lifelong function of neurons in many animals. Depending on the site of injury, new axons can grow either from the axon stump (after distal injury) or from the tip of a dendrite (after proximal injury). However, some neuron types do not have dendrites to be converted to a regenerating axon after proximal injury. For example, many sensory neurons receive information from a specialized sensory cilium rather than a branched dendrite arbor. We hypothesized that the lack of traditional dendrites would limit the ability of ciliated sensory neurons to respond to proximal axon injury. We tested this hypothesis by performing laser microsurgery on ciliated lch1 neurons in Drosophila larvae and tracking cells over time. These cells survived proximal axon injury as well as distal axon injury, and, like many other neurons, initiated growth from the axon stump after distal injury. After proximal injury, neurites regrew in a surprisingly flexible manner. Most cells initiated outgrowth directly from the cell body, but neurite growth could also emerge from the short axon stump or base of the cilium. New neurites were often branched. Although outgrowth after proximal axotomy was variable, it depended on the core DLK axon injury signaling pathway. Moreover, each cell had at least one new neurite specified as an axon based on microtubule polarity and accumulation of the endoplasmic reticulum. We conclude that ciliated sensory neurons are not intrinsically limited in their ability to grow a new axon after proximal axon removal.


Assuntos
Axônios , Regeneração Nervosa , Animais , Axônios/fisiologia , Regeneração Nervosa/fisiologia , Drosophila/metabolismo , Células Receptoras Sensoriais , Transdução de Sinais
15.
Dev Biol ; 478: 1-12, 2021 10.
Artigo em Inglês | MEDLINE | ID: mdl-34147472

RESUMO

Dorsal root ganglion (DRG) neurons are the predominant cell type that innervates the vertebrate skin. They are typically described as pseudounipolar cells that have central and peripheral axons branching from a single root exiting the cell body. The peripheral axon travels within a nerve to the skin, where free sensory endings can emerge and branch into an arbor that receives and integrates information. In some immature vertebrates, DRG neurons are preceded by Rohon-Beard (RB) neurons. While the sensory endings of RB and DRG neurons function like dendrites, we use live imaging in zebrafish to show that they have axonal plus-end-out microtubule polarity at all stages of maturity. Moreover, we show both cell types have central and peripheral axons with plus-end-out polarity. Surprisingly, in DRG neurons these emerge separately from the cell body, and most cells never acquire the signature pseudounipolar morphology. Like another recently characterized cell type that has multiple plus-end-out neurites, ganglion cells in Nematostella, RB and DRG neurons maintain a somatic microtubule organizing center even when mature. In summary, we characterize key cellular and subcellular features of vertebrate sensory neurons as a foundation for understanding their function and maintenance.


Assuntos
Gânglios Espinais/ultraestrutura , Microtúbulos/ultraestrutura , Células Receptoras Sensoriais/ultraestrutura , Pele/inervação , Animais , Animais Geneticamente Modificados , Axônios/fisiologia , Axônios/ultraestrutura , Corpo Celular/ultraestrutura , Polaridade Celular , Dendritos/fisiologia , Drosophila/citologia , Drosophila/crescimento & desenvolvimento , Gânglios Espinais/fisiologia , Centro Organizador dos Microtúbulos/ultraestrutura , Anêmonas-do-Mar/citologia , Anêmonas-do-Mar/crescimento & desenvolvimento , Anêmonas-do-Mar/ultraestrutura , Células Receptoras Sensoriais/fisiologia , Peixe-Zebra
16.
Dev Biol ; 465(2): 108-118, 2020 09 15.
Artigo em Inglês | MEDLINE | ID: mdl-32687893

RESUMO

Neurons extend dendrites and axons to receive and send signals. If either type of process is removed, the cell cannot function. Rather than undergoing cell death, some neurons can regrow axons and dendrites. Axon and dendrite regeneration have been examined separately and require sensing the injury and reinitiating the correct growth program. Whether neurons in vivo can sense and respond to simultaneous axon and dendrite injury with polarized regeneration has not been explored. To investigate the outcome of simultaneous axon and dendrite damage, we used a Drosophila model system in which neuronal polarity, axon regeneration, and dendrite regeneration have been characterized. After removal of the axon and all but one dendrite, the remaining dendrite was converted to a process that had a long unbranched region that extended over long distances and a region where shorter branched processes were added. These observations suggested axons and dendrites could regrow at the same time. To further test the capacity of neurons to implement polarized regeneration after axon and dendrite damage, we removed all neurites from mature neurons. In this case a long unbranched neurite and short branched neurites were regrown from the stripped cell body. Moreover, the long neurite had axonal plus-end-out microtubule polarity and the shorter neurites had mixed polarity consistent with dendrite identity. The long process also accumulated endoplasmic reticulum at its tip like regenerating axons. We conclude that neurons in vivo can respond to simultaneous axon and dendrite injury by initiating growth of a new axon and new dendrites.


Assuntos
Axônios/metabolismo , Dendritos/metabolismo , Microtúbulos/metabolismo , Animais , Axônios/patologia , Dendritos/patologia , Drosophila melanogaster , Feminino , Masculino
17.
J Neurosci ; 39(43): 8457-8470, 2019 10 23.
Artigo em Inglês | MEDLINE | ID: mdl-31492772

RESUMO

The degeneration of injured axons involves a self-destruction pathway whose components and mechanism are not fully understood. Here, we report a new regulator of axonal resilience. The transmembrane protein Raw is cell autonomously required for the degeneration of injured axons, dendrites, and synapses in Drosophila melanogaster In both male and female raw hypomorphic mutant or knock-down larvae, the degeneration of injured axons, dendrites, and synapses from motoneurons and sensory neurons is strongly inhibited. This protection is insensitive to reduction in the levels of the NAD+ synthesis enzyme Nmnat (nicotinamide mononucleotide adenylyl transferase), but requires the c-Jun N-terminal kinase (JNK) mitogen-activated protein (MAP) kinase and the transcription factors Fos and Jun (AP-1). Although these factors were previously known to function in axonal injury signaling and regeneration, Raw's function can be genetically separated from other axonal injury responses: Raw does not modulate JNK-dependent axonal injury signaling and regenerative responses, but instead restrains a protective pathway that inhibits the degeneration of axons, dendrites, and synapses. Although protection in raw mutants requires JNK, Fos, and Jun, JNK also promotes axonal degeneration. These findings suggest the existence of multiple independent pathways that share modulation by JNK, Fos, and Jun that influence how axons respond to stress and injury.SIGNIFICANCE STATEMENT Axonal degeneration is a major feature of neuropathies and nerve injuries and occurs via a cell autonomous self-destruction pathway whose mechanism is poorly understood. This study reports the identification of a new regulator of axonal degeneration: the transmembrane protein Raw. Raw regulates a cell autonomous nuclear signaling pathway whose yet unknown downstream effectors protect injured axons, dendrites, and synapses from degenerating. These findings imply that the susceptibility of axons to degeneration is strongly regulated in neurons. Future understanding of the cellular pathway regulated by Raw, which engages the c-Jun N-terminal kinase (JNK) mitogen-activated protein (MAP) kinase and Fos and Jun transcription factors, may suggest new strategies to increase the resiliency of axons in debilitating neuropathies.


Assuntos
Axônios/metabolismo , Proteínas do Citoesqueleto/metabolismo , Dendritos/metabolismo , Proteínas de Drosophila/metabolismo , Sistema de Sinalização das MAP Quinases/fisiologia , Degeneração Neural/metabolismo , Animais , Animais Geneticamente Modificados , Axônios/patologia , Proteínas do Citoesqueleto/genética , Dendritos/patologia , Proteínas de Drosophila/genética , Drosophila melanogaster , Feminino , Proteínas Quinases JNK Ativadas por Mitógeno/metabolismo , Masculino , Neurônios Motores/metabolismo , Neurônios Motores/patologia , Degeneração Neural/patologia , Sinapses/metabolismo
18.
J Exp Biol ; 223(Pt 21)2020 11 10.
Artigo em Inglês | MEDLINE | ID: mdl-32968001

RESUMO

The centralized nervous systems of bilaterian animals rely on directional signaling facilitated by polarized neurons with specialized axons and dendrites. It is not known whether axo-dendritic polarity is exclusive to bilaterians or was already present in early metazoans. We therefore examined neurite polarity in the starlet sea anemone Nematostella vectensis (Cnidaria). Cnidarians form a sister clade to bilaterians and share many neuronal building blocks characteristic of bilaterians, including channels, receptors and synaptic proteins, but their nervous systems comprise a comparatively simple net distributed throughout the body. We developed a tool kit of fluorescent polarity markers for live imaging analysis of polarity in an identified neuron type, large ganglion cells of the body column nerve net that express the LWamide-like neuropeptide. Microtubule polarity differs in bilaterian axons and dendrites, and this in part underlies polarized distribution of cargo to the two types of processes. However, in LWamide-like+ neurons, all neurites had axon-like microtubule polarity suggesting that they may have similar contents. Indeed, presynaptic and postsynaptic markers trafficked to all neurites and accumulated at varicosities where neurites from different neurons often crossed, suggesting the presence of bidirectional synaptic contacts. Furthermore, we could not identify a diffusion barrier in the plasma membrane of any of the neurites like the axon initial segment barrier that separates the axonal and somatodendritic compartments in bilaterian neurons. We conclude that at least one type of neuron in Nematostella vectensis lacks the axo-dendritic polarity characteristic of bilaterian neurons.


Assuntos
Anêmonas-do-Mar , Animais , Axônios , Citoesqueleto , Microtúbulos , Neurônios
19.
PLoS Genet ; 12(12): e1006503, 2016 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-27923046

RESUMO

Axon injury can lead to several cell survival responses including increased stability and axon regeneration. Using an accessible Drosophila model system, we investigated the regulation of injury responses and their relationship. Axon injury stabilizes the rest of the cell, including the entire dendrite arbor. After axon injury we found mitochondrial fission in dendrites was upregulated, and that reducing fission increased stabilization or neuroprotection (NP). Thus axon injury seems to both turn on NP, but also dampen it by activating mitochondrial fission. We also identified caspases as negative regulators of axon injury-mediated NP, so mitochondrial fission could control NP through caspase activation. In addition to negative regulators of NP, we found that nicotinamide mononucleotide adenylyltransferase (Nmnat) is absolutely required for this type of NP. Increased microtubule dynamics, which has previously been associated with NP, required Nmnat. Indeed Nmnat overexpression was sufficient to induce NP and increase microtubule dynamics in the absence of axon injury. DLK, JNK and fos were also required for NP. Because NP occurs before axon regeneration, and NP seems to be actively downregulated, we tested whether excessive NP might inhibit regeneration. Indeed both Nmnat overexpression and caspase reduction reduced regeneration. In addition, overexpression of fos or JNK extended the timecourse of NP and dampened regeneration in a Nmnat-dependent manner. These data suggest that NP and regeneration are conflicting responses to axon injury, and that therapeutic strategies that boost NP may reduce regeneration.


Assuntos
Axônios/metabolismo , Drosophila melanogaster/genética , Nicotinamida-Nucleotídeo Adenililtransferase/genética , Degeneração Walleriana/genética , Animais , Axônios/patologia , Caspases/biossíntese , Caspases/genética , Dendritos/metabolismo , Dendritos/patologia , Proteínas de Drosophila/biossíntese , Proteínas de Drosophila/genética , Drosophila melanogaster/crescimento & desenvolvimento , Humanos , MAP Quinase Quinase 4/biossíntese , MAP Quinase Quinase 4/genética , Microtúbulos/genética , Microtúbulos/patologia , Dinâmica Mitocondrial/genética , Neurônios/metabolismo , Neurônios/patologia , Fármacos Neuroprotetores/metabolismo , Nicotinamida-Nucleotídeo Adenililtransferase/antagonistas & inibidores , Nicotinamida-Nucleotídeo Adenililtransferase/biossíntese , RNA Interferente Pequeno/genética , Degeneração Walleriana/patologia
20.
PLoS Genet ; 12(12): e1006457, 2016 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-27911898

RESUMO

In vertebrate neurons, the axon initial segment (AIS) is specialized for action potential initiation. It is organized by a giant 480 Kd variant of ankyrin G (AnkG) that serves as an anchor for ion channels and is required for a plasma membrane diffusion barrier that excludes somatodendritic proteins from the axon. An unusually long exon required to encode this 480Kd variant is thought to have been inserted only recently during vertebrate evolution, so the giant ankyrin-based AIS scaffold has been viewed as a vertebrate adaptation for fast, precise signaling. We re-examined AIS evolution through phylogenomic analysis of ankyrins and by testing the role of ankyrins in proximal axon organization in a model multipolar Drosophila neuron (ddaE). We find giant isoforms of ankyrin in all major bilaterian phyla, and present evidence in favor of a single common origin for giant ankyrins and the corresponding long exon in a bilaterian ancestor. This finding raises the question of whether giant ankyrin isoforms play a conserved role in AIS organization throughout the Bilateria. We examined this possibility by looking for conserved ankyrin-dependent AIS features in Drosophila ddaE neurons via live imaging. We found that ddaE neurons have an axonal diffusion barrier proximal to the cell body that requires a giant isoform of the neuronal ankyrin Ank2. Furthermore, the potassium channel shal concentrates in the proximal axon in an Ank2-dependent manner. Our results indicate that the giant ankyrin-based cytoskeleton of the AIS may have evolved prior to the radiation of extant bilaterian lineages, much earlier than previously thought.


Assuntos
Anquirinas/genética , Segmento Inicial do Axônio/metabolismo , Proteínas de Drosophila/genética , Filogenia , Canais de Potássio Shal/genética , Potenciais de Ação/genética , Animais , Anquirinas/biossíntese , Membrana Celular/genética , Proteínas de Drosophila/biossíntese , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/genética , Drosophila melanogaster/crescimento & desenvolvimento , Neurônios/metabolismo , Canais de Potássio Shal/metabolismo
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA