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Wang, Zhiping, Wu, Zilu, Ghosh-Roy, Anindya, Bowerman, Bruce, O'Rourke, Sean, Hubert, Thomas, Yan, Dong, Jin, Yishi, Chen, Lizhen, Chisholm, Andrew
[
International Worm Meeting,
2011]
The mechanisms underlying the ability of axons to regrow after injury remain little explored at the molecular genetic level. We use a recently established laser injury model in Caenorhabditis elegans mechanosensory neurons to screen 654 conserved genes for novel regulators of axonal regrowth. We uncover several unexpected functional clusters of genes that promote or repress regrowth, including genes classically known to affect membrane excitability, neurotransmission, and synaptic vesicle endocytosis. We find that the conserved Arf Guanine nucleotide Exchange Factor (GEF), EFA-6, acts as an intrinsic inhibitor of regrowth. By combining genetics and in vivo imaging we show that EFA-6 inhibits regrowth via microtubule dynamics, independent of its Arf GEF activity. Among the newly identified regrowth inhibitors, only loss of function in EFA-6 partially bypasses the requirement for DLK-1. Identification of these pathways significantly expands our understanding of the genetic basis of axonal injury responses and repair.
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Chuang, Marian, Chisholm, Andrew, Chen, Lizhen, Koorman, Thijs, Boxem, Mike, Jin, Yishi
[
International Worm Meeting,
2015]
Axon injury triggers a complex sequence of changes in the axonal cytoskeleton that are prerequisites for effective axon regeneration. We previously identified EFA-6 as a potent intrinsic inhibitor of axon regrowth in C. elegans (Chen et al. 2011, Neuron). We report that axon injury triggers a rapid and transient relocalization of EFA-6 from the plasma membrane to the cytoskeleton, concomitant with a local downregulation of axonal MT dynamics after injury. Relocalization is modulated by axonal Ca2+ levels and correlates with EFA-6 protein function in microtubule (MT) regulation and axon regeneration. The N-terminus of EFA-6 is predicted to be an intrinsically disordered domain, and mediates the abilioty of EFA-6 to modulate MT dynamics and axon growth and regeneration. A conserved 18-aa motif in the N terminus is required for its injury-induced relocalization and for inhibition of axon regeneration. We show that the EFA-6 N-terminal domain directly interacts with MT associated proteins TAC-1, a member of the TACC (Transforming-Acidic-Coiled-Coil) family, and ZYG-8, an ortholog of Doublecortin-Like Kinase (DCLK). Using conditional alleles and tissue-specific knockout strategies we find that TAC-1 and ZYG-8 are required for initiation of axon regeneration, and that their overexpression can promote regrowth. Furthermore, injury triggers relocalization of EFA-6 and TAC-1 to sites overlapping with the MT minus end binding protein Patronin/PTRN-1. We propose that EFA-6 is a bifunctional injury-responsive regulator of MT dynamics, acting at the cell cortex in the steady state and at MT minus ends after axon injury.
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[
International Worm Meeting,
2017]
Aging impacts the function of the nervous system and is the major risk factor for neurodegenerative diseases and is a fundamental problem in basic neuroscience and in human health. On the other hand, the nervous system corporate the organism's overall metabolism and affect homeostasis and longevity. MTs are essential cytoskeleton involved in cell division, shaping the cell and intracellular transport. Microtubule (MT) regulation is involved on several levels in neuronal function and maintenance of neuronal structure, and also appears to be a general downstream indicator and effector in age-dependent neurodegeneration. Here we hypothesized that neuronal MT stability regulates neuronal aging and organismal longevity. Consistent with previous reports, we found that MT-regulating genes played a role in maintaining neuronal integrity during aging. We then tested the lifespan of genetic mutants of MT-regulating genes in C. elegans and found that loss of MT stabilizing genes shortened lifespan, while loss of MT destabilizing genes enhanced lifespan. Decline in mobility is often associated with aging. Our data suggested that stabilizing neuronal MT enhance health span by improving mobility, and destabilization of MT accelerated age-dependent mobility decline. We further tested the effect of aging on MT-based intracellular transport by examining the localization of synaptic vesicles. We found MT-regulating genes and MT drugs could modulate the age-associated changes in vesicle localization. Finally we tested the genetic interaction between MT genes and aging regulators. Our data suggested that neuronal MT regulation might be involved in stress response to modulate longevity.
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[
Neuron,
2012]
The adult mammalian central nervous system exhibits restricted regenerative potential. Chen etal. (2011) and El Bejjani and Hammarlund (2012) used Caenorhabditis elegans to uncover intrinsic factors that inhibit regeneration of axotomized mature neurons, opening avenues for potential therapeutics.
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[
International Worm Meeting,
2019]
Autophagy is involved in normal physiology and various diseases including cancers, heart diseases and neurodegeneration. Genetically inhibiting autophagy has been shown to cause neurodegeneration in various brain regions. On the other hand, pharmacologically inhibiting autophagy is sufficient to suppress acute neurodegeneration of retinal ganglion cell axons. These results suggest a key role of autophagy in maintaining neuronal homeostasis. More recently the role of autophagy in axon regeneration post injury has been assessed. Expression of LC3, which is critical for autophagosome formation and a marker of autophagy, is up regulated in dorsal root ganglia (DRGs) after acute spinal cord injury. Activating autophagy using autophagy-inducing peptide can promote axon regeneration through limiting SCG10, a microtubule destabilizing protein. We investigated the regulation of autophagy in axon regeneration after injury using C. elegans as a model. We found that genetic mutants of genes involved in different steps in the autophagy process displayed impaired axon regeneration. Taking advantage of a tandem reporter, we were able to monitor the dynamics of autophagosomes (AP) and autolysosomes (AL) in injured neurons. We found that axotomy induced an elevation of both AP and AL in an age-dependent manner. Pharmacologically blocking autophagy flux inhibited axon regrowth, while activating autophagy with rapamycin promoted axon regeneration, but the regrowth-promoting effect was only seen in aged animals. Axon injury failed to activate autophagy in animals lacking DLK-1, a conserved regulator of axon regeneration that promotes retrograde injury signaling. Activating autophagy was able to partially bypass the requirement of DLK-1 in axon regeneration. We hypothesize that DLK-mediated injury signaling can activate autophagy, which promotes axon regeneration by degrading regeneration-inhibiting proteins. We are currently investigating the functional targets of autophagy in axon regeneration.
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[
International Worm Meeting,
2021]
The concept of phase separation provides a new framework to understand the role of proteins with intrinsically disordered regions (IDRs). We have previously identified the signaling protein Exchange Factor for ARF-6 (EFA-6) as a potent inhibitor of axon regrowth. We have also shown that axon injury triggers a rapid redistribution of EFA-6 protein from a generally even plasma membrane localization to more discrete puncta within the cytosol. Both the redistribution of EFA-6 and inhibition of axon regrowth are mediated by its intrinsically disordered N-terminal domain and requires a conserved 18aa motif at the N terminus. We further demonstrated that EFA-6 inhibited axonal microtubule growth and interacted with the MT-associated proteins TAC-1 and ZYG-8, both of which are required for axon regeneration. EFA-6 is known to limit microtubule growth at the cell cortex. The D. melanogaster ortholog of EFA-6 has also been reported to inhibit microtubule polymerization at the cortex. However, how EFA-6 plays its role in regulating microtubules remains unclear. We recently purified recombinant EFA-6 proteins and observed that EFA-6 formed liquid-like droplets and phase separated with TAC-1. Interestingly, we found that EFA-6 could phase transition from liquid-like droplets to gel-like aggregates, and the transition was dependent on the conserved 18aa motif. We are currently investigating whether and how EFA-6 phase transition regulates its function.
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[
Immunity,
2017]
IL-17 is a cytokine known primarily for its role in inflammation. In a recent issue of Nature, Chen etal. (2017) demonstrate that IL-17 plays a neuromodulatory role in Caenorhabditis elegans by acting directly on neurons to amplify neuronal responses to stimuli and produce changes in animal behavior.
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[
International Worm Meeting,
2021]
Autophagy plays a conserved role in maintaining cellular homeostasis by degrading dysfunctional proteins, lipids, and organelles. Autophagic dysfunction has been reported in various neurodegenerative diseases. It can occur at any step of the autophagic process and can contribute to the formation of protein aggregates and ultimately to neuronal death, although it remains unclear whether autophagy impairment is a contributor or a consequence of neurodegeneration. Axonal injury is acute neuronal stress that triggers autophagic responses in an age-dependent manner. In this study, we investigate the injury-triggered autophagy response in a C. elegans model of tauopathy. We found that the pro-aggregant F3deltaK280 Tau abolished injury-induced autophagic activity, thereby reducing axon regeneration capacity. Besides, axon trafficking of autophagic vesicles is significantly reduced in animals expressing the pro-aggregant F3deltaK280 Tau, showing that Tau aggregation impairs autophagy regulation. Notably, in the tauopathy model, the reduced number of total or trafficking autophagic vesicles was not restored by the autophagy activator rapamycin. Loss of PTL-1, the sole Tau homologue in C. elegans, also led to impaired injury-induced autophagy activation and decreased axon trafficking of autophagic vesicles, but with an increased basal level of autophagic vesicles. Therefore, we have demonstrated that Tau aggregation as well as Tau depletion both lead to disruption of injury-induced autophagy responses, suggesting that aberrant protein aggregation or microtubule dysfunction can modulate autophagy regulation in injured neurons.
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[
Cell,
2014]
Surface receptors can link binding of ligands to changes in the actin-based cell cytoskeleton. Chia etal. and Chen etal. provide evidence for direct binding between the cytoplasmic tails ofreceptorsand the WAVE complex, a regulator of the actin nucleator Arp2/3 complex, which mighthelp to explain how environmental signals are translated into changes in morphology andmotility.
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[
Dev Cell,
2016]
Temperature-sensing neurons in C.elegans reduce the life-shortening effects of high temperatures via steroid signaling. In this issue of Developmental Cell, Chen etal. (2016) elucidate the underlying mechanisms by which the transcription factor CREB induces the neuropeptide FLP-6 in the temperature-sensing neurons to counteract the life-shortening effects of high temperature.