Patterson, Joseph R et al. (2011) International Worm Meeting "RNP granules: insights into their regulation and function."

Wood, Megan P, Patterson, Joseph R, Schisa, Jennifer A

[International Worm Meeting, 2011]

As sperm is depleted in Caenorhabditis elegans hermaphrodites, ovulation arrests and ribonucleoprotein (RNP) granules assemble in the arrested oocytes. RNP granules are also induced in non-arrested oocytes by environmental stresses such as heat shock, osmotic shock, and anoxia. The large RNP granules in oocytes are hypothesized to maintain mRNA stability and prevent precocious translation when fertilization is delayed or a stress is present. The assembly of RNP granules in response to extended meiotic arrest and stress is conserved in a related species, Caenorhabditis remanei. We have focused on two broad questions: 1) how is the assembly of RNP granules regulated? and 2) what is the consequence to an arrested oocyte if RNP granules fail to assemble normally? To gain insights into the mechanism for RNP granule assembly we first performed an ultrastructural analysis of the C. remanei germline. We were very surprised to see dramatic nuclear blebbing along the nuclear envelope of oocytes of unmated and heat stressed females. A combination of TEM, confocal analyses, and live imaging studies indicate the blebs appear to detach from the nucleus and may traffick to the cell cortex and assemble into annulate lamellae, in close proximity to RNP granules. Using RNAi, we further show that several nucleoporins are required for the assembly of RNP granules, and a disruption in RNP granule assembly coupled with a low frequency of nuclear blebbing in arrested oocytes negatively impacts embryonic viability. Our observations support a model where nuclear membrane blebbing is required to increase the trafficking of nucleoporins to the cell cortex in stressed or meiotically-arrested cells and to facilitate the recruitment of RNA and protein components of RNPs into large complexes. To further address the question of the function of RNP granules, we are performing additional fertility tests after preventing normal RNP granule assembly. We are using RNAi to knockdown genes that have recently been identified in our lab as required for normal RNP granule assembly, and performing mating assays to assess any consequences to embryonic viability. We expect these experiments to address part of our hypothesis for the function of RNP granules in meiotically arrested oocytes.

Arena JP (1994) Parasitol Today "Expression of Caenorahbditis elegans messenger RNA in Xenopus oocytes: A model systems to study the mechanism of action of avermectins."

Arena JP

[Parasitol Today, 1994]

It has recently been shown that Xenopus oocytes injected with mRNA from the free-living nematode Caenorhabditis elegans express avermectin-sensitive chloride channels(1). Joseph Arena here reviews whet is known about the mechanism of action of avermectin and how these recent results relate to the mechanism in nematodes.

Wood, Megan et al. (2013) International Worm Meeting "Assembly of RNP granules in C. elegans oocytes promotes oocyte quality and is regulated by the cytoskeleton."

Gorman, Kevin, Schisa, Jennifer, Boag, Peter, Severance, Ashley, Davis, Gregory, Patterson, Joseph, Hollis, Angela, Wood, Megan

[International Worm Meeting, 2013]

In many animal species, oocytes arrest in meiosis until they are fertilized. It is well established that fertility diminishes as oocytes age. Our goal is to better understand the regulation and function of large ribonucleoprotein (RNP) granules that assemble in the germ lines of Caenorhabditis nematodes that are either stressed or in which ovulation is arrested. The RNP granules are hypothesized to maintain oocyte quality by regulating mRNA stability or translation in arrested or stressed oocytes (Jud et al., 2008). Their assembly is influenced by nuclear pore proteins, and we have hypothesized that nuclear blebs trafficking from the nuclear envelope to the cortex may promote the formation of the cortical RNP granules (Patterson et al., 2011). We have performed a targeted, functional RNAi screen to identify genes that are required for the assembly of RNP granules in arrested oocytes and identified 143 genes that are necessary. Among the gene classes of our positives are several cytoskeleton proteins including KCA-1 (kinesin cargo adaptor), several beta-tubulins, and WSP-1 (involved in actin polymerization). To gain insight into the mechanism of action of RNP granule regulators we are dissecting defined protein complexes; e.g. we are determining if KLC-1 (kinesin light chain) and UNC-116 (kinesin 1 heavy chain) which function with KCA-1 to position the meiotic spindle (Yang et al., 2005), also contribute to RNP granule assembly. The discovery of these novel regulators of RNP granule assembly allows for direct testing of our hypothesis for their function. When the normal assembly of RNP granules is prevented, we observe fertility is decreased, supporting the hypothesis that RNP granules maintain the quality of oocytes when fertilization is delayed. On-going studies are testing if RNA stability is diminished or translation of maternal mRNAs is de-repressed when RNP granule assembly is defective. These results have provided insight into novel regulators of RNP dynamics that likely apply to RNPs important for fertility and stress responses in many species.

Teixeira-Castro A et al. (2015) Brain "Serotonergic signalling suppresses ataxin 3 aggregation and neurotoxicity in animal models of Machado-Joseph disease."

Bessa C, Duarte-Silva S, Maciel P, Bessa J, Silverman RB, Miranda A, Kang S, Summavielle T, Oliveira S, da Silva Santos L, Neto MF, Esteves S, Brielmann RM, Neves-Carvalho A, Teixeira-Castro A, Oliveira P, Morimoto RI, Silva-Fernandes A, Jalles A

[Brain, 2015]

Polyglutamine diseases are a class of dominantly inherited neurodegenerative disorders for which there is no effective treatment. Here we provide evidence that activation of serotonergic signalling is beneficial in animal models of Machado-Joseph disease. We identified citalopram, a selective serotonin reuptake inhibitor, in a small molecule screen of FDA-approved drugs that rescued neuronal dysfunction and reduced aggregation using a Caenorhabditis elegans model of mutant ataxin 3-induced neurotoxicity. MOD-5, the C. elegans orthologue of the serotonin transporter and cellular target of citalopram, and the serotonin receptors SER-1 and SER-4 were strong genetic modifiers of ataxin 3 neurotoxicity and necessary for therapeutic efficacy. Moreover, chronic treatment of CMVMJD135 mice with citalopram significantly reduced ataxin 3 neuronal inclusions and astrogliosis, rescued diminished body weight and strikingly ameliorated motor symptoms. These results suggest that small molecule modulation of serotonergic signalling represents a promising therapeutic target for Machado-Joseph disease.

Kawaguchi Y et al. (1994) Nat Genet "CAG expansions in a novel gene for Machado-Joseph disease at chromosome 14q32.1."

Kawaguchi Y, Inoue M, Nishimura M, Kawakami H, Okamoto T, Katayama S, Nakamura S, Aizawa M, Akiguchi I, Taniwaki M

[Nat Genet, 1994]

We have identified a novel gene containing CAG repeats and mapped it to chromosome 14q32.1, the genetic locus for Machado-Joseph disease (MJD). In normal individuals the gene contains between 13 and 36 CAG repeats, whereas most of the clinically diagnosed patients and all of the affected members of a family with the clinical and pathological diagnosis of MJD show expansion of the repeat-number (from 68-79). Southern blot analyses and genomic cloning demonstrates the existence of related genes. These results raise the possibility that similar abnormalities in related genes may give rise to diseases similar to MJD.

Rodrigues AJ et al. (2007) FASEB J "Functional genomics and biochemical characterization of the C. elegans orthologue of the Machado-Joseph disease protein ataxin-3."

Ailion M, Rodrigues AJ, Maciel P, Geschwind DH, Coppola G, Santos C, Costa MD, Sequeiros J

[FASEB J, 2007]

Machado-Joseph disease (MJD) is the most common dominant spinocerebellar ataxia. MJD is caused by a CAG trinucleotide expansion in the ATXN3 gene, which encodes a protein named ataxin-3. Ataxin-3 has been proposed to act as a deubiquitinating enzyme in the ubiquitin-proteasome pathway and to be involved in transcriptional repression; nevertheless, its precise biological function(s) remains unknown. To gain further insight into the function of ataxin-3, we have identified the Caenorhabditis elegans orthologue of the ATXN3 gene and characterized its pattern of expression, developmental regulation, and subcellular localization. We demonstrate that, analogous to its human orthologue, C. elegans ataxin-3 has deubiquitinating activity in vitro against polyubiquitin chains with four or more ubiquitins, the minimum ubiquitin length for proteasomal targeting. To further evaluate C. elegans ataxin-3, we characterized the first known knockout animal models both phenotypically and biochemically, and found that the two C. elegans strains were viable and displayed no gross phenotype. To identify a molecular phenotype, we performed a large-scale microarray analysis of gene expression in both knockout strains. The data revealed a significant deregulation of core sets of genes involved in the ubiquitin-proteasome pathway, structure/motility, and signal transduction. This gene identification provides important clues that can help elucidate the specific biological role of ataxin-3 and unveil some of the physiological effects caused by its absence or diminished function.--Rodrigues, A-J., Coppola, G., Santos, C., do Carmo Costa, M., Ailion, M., Sequeiros, J., Geschwind, D. H., Maciel, P. Functional genomics and biochemical characterization of the C. elegans orthologue of the Machado-Joseph disease protein ataxin-3.

Pohl F et al. (2024) MicroPubl Biol "Pharmacological inhibition of acetylcholinesterase improves the locomotion defective phenotype of a SCA3 C. elegans model."

Maciel P, Pohl F, Teixeira-Castro A, Lindsay-McGee V, Kong Thoo Lin P

[MicroPubl Biol, 2024]

Inhibition of acetylcholinesterase (AChE) is a common used treatment option for Alzheimer's disease. However, there has been limited research on the potential use of AChE inhibitors for the treatment of Machado-Joseph disease (MJD)/Spinocerebellar Ataxia 3 (SCA3), in spite of the positive results using AChE inhibitors in patients with other inherited ataxias. MJD/SCA3, the most common form of dominant Spinocerebellar Ataxia worldwide, is caused by an expansion of the polyglutamine tract within the ataxin-3 protein, and is characterized by motor impairments. Our study shows that administration of the AChE inhibitor neostigmine is beneficial in treating the locomotion defective phenotype of a SCA3/MJD model of <i>C. elegans</i> and highlights the potential contribution of AChE enzymes to mutant ataxin-3-mediated toxicity.

Teixeira-Castro A et al. (2011) Hum Mol Genet "Neuron-specific proteotoxicity of mutant ataxin-3 in C. elegans: rescue by the DAF-16 and HSF-1 pathways."

Oliveira JF, Vilaca JL, Brignull HR, Ailion M, Teixeira-Castro A, Dias N, Neves-Carvalho A, Maciel P, Morimoto RI, Jalles A, Rodrigues P

[Hum Mol Genet, 2011]

The risk of developing neurodegenerative diseases increases with age. Although many of the molecular pathways regulating proteotoxic stress and longevity are well characterized, their contribution to disease susceptibility remains unclear. In this study, we describe a new Caenorhabditis elegans model of Machado-Joseph disease pathogenesis. Pan-neuronal expression of mutant ATXN3 leads to a polyQ-length dependent, neuron subtype-specific aggregation and neuronal dysfunction. Analysis of different neurons revealed a pattern of dorsal nerve cord and sensory neuron susceptibility to mutant ataxin-3 that was distinct from the aggregation and toxicity profiles of polyQ-alone proteins. This reveals that the sequences flanking the polyQ-stretch in ATXN3 have a dominant influence on cell-intrinsic neuronal factors that modulate polyQ-mediated pathogenesis. Aging influences the ATXN3 phenotypes which can be suppressed by the downregulation of the insulin/insulin growth factor-1-like signaling pathway and activation of heat shock factor-1.

Worrall L et al. (2006) Acta Crystallogr Sect F Struct Biol Cryst Commun "Crystallization and X-ray data analysis of the 10 kDa C-terminal lid subdomain from Caenorhabditis elegans Hsp70."

Worrall L, Walkinshaw MD

[Acta Crystallogr Sect F Struct Biol Cryst Commun, 2006]

Hsp70 is an important molecular chaperone involved in the regulation of protein folding. Crystals of the C-terminal 10 kDa helical lid domain (residues 542-640) from a Caenorhabditis elegans Hsp70 homologue have been produced that diffract X-rays to approximately 3.4 A. Crystals belong to space group I2(1)2(1)2(1), with unit-cell parameters a = b = 197, c = 200 A. The Matthews coefficient, self-rotation function and Patterson map indicate 24 monomers in the asymmetric unit, showing non-crystallographic 432 symmetry. Molecular-replacement studies using the corresponding domain from rat, the only eukaryotic homologue with a known structure, failed and a mercury derivative was obtained. Preliminary MAD phasing using SHELXD and SHARP for location and refinement of the heavy-atom substructure and SOLOMON for density modification produced interpretable maps with a clear protein-solvent boundary. Further density-modification, model-building and refinement are currently under way.

Wu Y et al. (2023) Int J Mol Sci "Towards Understanding Neurodegenerative Diseases: Insights from Caenorhabditis elegans."

Yu X, Wu Y, Zhang M, Li Z, Chen Y

[Int J Mol Sci, 2023]

The elevated occurrence of debilitating neurodegenerative disorders, such as amyotrophic lateral sclerosis (ALS), Huntington's disease (HD), Alzheimer's disease (AD), Parkinson's disease (PD) and Machado-Joseph disease (MJD), demands urgent disease-modifying therapeutics. Owing to the evolutionarily conserved molecular signalling pathways with mammalian species and facile genetic manipulation, the nematode <i>Caenorhabditis elegans</i> (<i>C. elegans</i>) emerges as a powerful and manipulative model system for mechanistic insights into neurodegenerative diseases. Herein, we review several representative <i>C. elegans</i> models established for five common neurodegenerative diseases, which closely simulate disease phenotypes specifically in the gain-of-function aspect. We exemplify applications of high-throughput genetic and drug screenings to illustrate the potential of <i>C. elegans</i> to probe novel therapeutic targets. This review highlights the utility of <i>C. elegans</i> as a comprehensive and versatile platform for the dissection of neurodegenerative diseases at the molecular level.