scholarly journals Novel exc Genes Involved in Formation of the Tubular Excretory Canals of C. elegans

2018 ◽  
Author(s):  
Hikmat Al-Hashimi ◽  
Travis Chiarelli ◽  
Erik A. Lundquist ◽  
Matthew Buechner
Keyword(s):  
Genetic Model ◽  
Luminal Diameter ◽  
Recycling Endosomes ◽  
C Elegans ◽  
Novel Proteins ◽  
Cytoplasmic Structure ◽  
Cytoskeletal Regulation ◽  
Conserved Genes ◽  
Excretory Canals ◽  
Vesicle Movement

ABSTRACTRegulation of luminal diameter is critical to the function of small single-celled tubes, of which the seamless tubular excretory canals of C. elegans provide a tractable genetic model. Mutations in several sets of genes exhibit the Exc phenotype, in which canal luminal growth is visibly altered. Here, a focused reverse genomic screen of genes highly expressed in the canals found 24 genes that significantly affect luminal outgrowth or diameter. These genes encode novel proteins as well as highly conserved proteins involved in processes including gene expression, cytoskeletal regulation, vesicular movement, and transmembrane transport. In addition, two genes act as suppressors on a pathway of conserved genes whose products mediate vesicle movement from early to recycling endosomes. The results provide new tools for understanding the integration of cytoplasmic structure and physiology in forming and maintaining the narrow diameter of single-cell tubules.

scholarly journals Caenorhabditis elegans as an in vivo Model to Assess Amphetamine Tolerance

2021 ◽  
pp. 1-9
Author(s):  
Dayana Torres Valladares ◽  
Sirisha Kudumala ◽  
Murad Hossain ◽  
Lucia Carvelli
Keyword(s):  
Caenorhabditis Elegans ◽  
Molecular Mechanisms ◽  
Drugs Of Abuse ◽  
Genetic Model ◽  
In Vivo Model ◽  
C Elegans ◽  
Hyperactivity Disorder ◽  
In Vitro Data

Amphetamine is a potent psychostimulant also used to treat attention deficit/hyperactivity disorder and narcolepsy. In vivo and in vitro data have demonstrated that amphetamine increases the amount of extra synaptic dopamine by both inhibiting reuptake and promoting efflux of dopamine through the dopamine transporter. Previous studies have shown that chronic use of amphetamine causes tolerance to the drug. Thus, since the molecular mechanisms underlying tolerance to amphetamine are still unknown, an animal model to identify the neurochemical mechanisms associated with drug tolerance is greatly needed. Here we took advantage of a unique behavior caused by amphetamine in <i>Caenorhabditis elegans</i> to investigate whether this simple, but powerful, genetic model develops tolerance following repeated exposure to amphetamine. We found that at least 3 treatments with 0.5 mM amphetamine were necessary to see a reduction in the amphetamine-induced behavior and, thus, to promote tolerance. Moreover, we found that, after intervals of 60/90 minutes between treatments, animals were more likely to exhibit tolerance than animals that underwent 10-minute intervals between treatments. Taken together, our results show that <i>C. elegans</i> is a suitable system to study tolerance to drugs of abuse such as amphetamines.

20. Discovery of Novel Proteins Involved the Insulin/IGF-1 Signalling Pathway

2018 ◽  
Author(s):  
David (Wen Xiao) Wei
Keyword(s):  
Protein Interactions ◽  
Mutant Form ◽  
Yeast Two Hybrid ◽  
C Elegans ◽  
Insulin Growth Factor ◽  
Novel Proteins ◽  
Similar Phenotype ◽  
Two Hybrid ◽  
Hsp90 Protein ◽  
Novel Protein

The insulin/insulin growth factor-1 (IGF-1) signalling (IIS) pathway plays a key role in metabolism, growth and development. Though research has elucidated aspects of this pathway, it is not fully characterized or understood. A better understanding of the pathway will give insight into related diseases such as cancer. To discover novel proteins involved in the IIS pathway, the C. elegans worm was used due to the homology its insulin/IGF-1 receptor shares with that of humans.  To identify novel protein interactions with the insulin/IGF-1 receptor, we performed a yeast two-hybrid screen using a library of worm proteins. We found several separate interactions with the worm homolog of the HSP90 protein. To support the involvement of HSP90 in the IIS pathway, we studied the phenotypes of worm strains with a mutant form of HSP90. They showed a similar phenotype to those that have a mutant form of the insulin/IGF-1 receptor, inappropriately entering a developmental stage known as dauer. This strongly suggests the involvement of HSP90 in the IIS pathway. Based on previous research, we hypothesized the interaction between HSP90 and the insulin/IGF-1 receptor may allow it to bind other proteins. Thus, we performed a modified yeast two-hybrid screen to identify proteins which interact with the receptor in the presence of HSP90. The screen identified 15 interactions, many more than with the insulin/IGF-1 receptor alone, supporting this hypothesis. Overall, we provide evidence of a novel interaction with insulin/IGF-1 receptor, suggesting HSP90 may be a potential target for developing therapies for IIS pathway related diseases.

scholarly journals An Evolutionarily Conserved Pathway Essential for Orsay Virus Infection of Caenorhabditis elegans

mBio ◽  
2017 ◽  
Vol 8 (5) ◽  
Author(s):  
Hongbing Jiang ◽  
Kevin Chen ◽  
Luis E. Sandoval ◽  
Christian Leung ◽  
David Wang
Keyword(s):  
Caenorhabditis Elegans ◽  
Virus Infection ◽  
Tyrosine Kinase ◽  
Model Organism ◽  
Wiskott Aldrich Syndrome ◽  
C Elegans ◽  
Evolutionarily Conserved ◽  
Conserved Genes ◽  
Orsay Virus ◽  
Syndrome Protein

ABSTRACT Many fundamental biological discoveries have been made in Caenorhabditis elegans. The discovery of Orsay virus has enabled studies of host-virus interactions in this model organism. To identify host factors critical for Orsay virus infection, we designed a forward genetic screen that utilizes a virally induced green fluorescent protein (GFP) reporter. Following chemical mutagenesis, two Viro (virus induced reporter off) mutants that failed to express GFP were mapped to sid-3, a nonreceptor tyrosine kinase, and B0280.13 (renamed viro-2), an ortholog of human Wiskott-Aldrich syndrome protein (WASP). Both mutants yielded Orsay virus RNA levels comparable to that of the residual input virus, suggesting that they are not permissive for Orsay virus replication. In addition, we demonstrated that both genes affect an early prereplication stage of Orsay virus infection. Furthermore, it is known that the human ortholog of SID-3, activated CDC42-associated kinase (ACK1/TNK2), is capable of phosphorylating human WASP, suggesting that VIRO-2 may be a substrate for SID-3 in C. elegans. A targeted RNA interference (RNAi) knockdown screen further identified the C. elegans gene nck-1, which has a human ortholog that interacts with TNK2 and WASP, as required for Orsay virus infection. Thus, genetic screening in C. elegans identified critical roles in virus infection for evolutionarily conserved genes in a known human pathway. IMPORTANCE Orsay virus is the only known virus capable of naturally infecting the model organism Caenorhabditis elegans, which shares many evolutionarily conserved genes with humans. We exploited the robust genetic tractability of C. elegans to identify three host genes, sid-3, viro-2, and nck-1, which are essential for Orsay virus infection. Mutant animals that lack these three genes are highly defective in viral replication. Strikingly, the human orthologs of these three genes, activated CDC42-associated kinase (TNK2), Wiskott-Aldrich syndrome protein (WASP), and noncatalytic region of tyrosine kinase adaptor protein 1 (NCK1) are part of a known signaling pathway in mammals. These results suggest that TNK2, WASP, and NCK1 may play important roles in mammalian virus infection. IMPORTANCE Orsay virus is the only known virus capable of naturally infecting the model organism Caenorhabditis elegans, which shares many evolutionarily conserved genes with humans. We exploited the robust genetic tractability of C. elegans to identify three host genes, sid-3, viro-2, and nck-1, which are essential for Orsay virus infection. Mutant animals that lack these three genes are highly defective in viral replication. Strikingly, the human orthologs of these three genes, activated CDC42-associated kinase (TNK2), Wiskott-Aldrich syndrome protein (WASP), and noncatalytic region of tyrosine kinase adaptor protein 1 (NCK1) are part of a known signaling pathway in mammals. These results suggest that TNK2, WASP, and NCK1 may play important roles in mammalian virus infection.

scholarly journals Alzheimer’s disease-relevant tau modifications selectively impact neurodegeneration and mitophagy in a novel C. elegans single-copy transgenic model

2020 ◽  
Author(s):  
Sanjib Guha ◽  
Sarah Fischer ◽  
Gail VW Johnson ◽  
Keith Nehrke
Keyword(s):  
Alzheimer’S Disease ◽  
Alzheimer's Disease ◽  
Genetic Model ◽  
Neurodegenerative Disorder ◽  
Model Organism ◽  
Single Copy ◽  
Mitochondrial Stress ◽  
C Elegans ◽  
Touch Receptor Neurons ◽  
New Perspective

ABSTRACTBackgroundA defining pathological hallmark of the progressive neurodegenerative disorder Alzheimer’s disease (AD) is the accumulation of misfolded tau with abnormal post-translational modifications (PTMs). These include phosphorylation at Threonine 231 (T231) and acetylation at Lysine 274 (K274) and at Lysine 281 (K281). Although tau is recognized to play a central role in pathogenesis of AD, the precise mechanisms by which these abnormal PTMs contribute to the neural toxicity of tau is unclear.MethodsHuman 0N4R tau (wild type) was expressed in touch receptor neurons of the genetic model organism C. elegans through single-copy gene insertion. Defined mutations were then introduced into the single-copy tau transgene through CRISPR-Cas9 genome editing. These mutations included T231E and T231A, to mimic phosphorylation and phospho-ablation of a commonly observed pathological epitope, respectively, and K274/281Q, to mimic disease-associated lysine acetylation. Stereotypical touch response assays were used to assess behavioral defects in the transgenic strains as a function of age, and genetically-encoded fluorescent biosensors were used to measure the morphological dynamics and turnover of touch neuron mitochondria.ResultsUnlike existing tau overexpression models, C. elegans single-copy expression of tau did not elicit overt pathological phenotypes at baseline. However, strains expressing disease associated PTM-mimetics (T231E and K274/281Q) exhibited reduced touch sensation and morphological abnormalities that increased with age. In addition, the PTM-mimetic mutants lacked the ability to engage mitophagy in response to mitochondrial stress.ConclusionsLimiting the expression of tau results in a genetic model where pathological modifications and age result in evolving phenotypes, which may more closely resemble the normal progression of AD. The finding that disease-associated PTMs suppress compensatory responses to mitochondrial stress provides a new perspective into the pathogenic mechanisms underlying AD.

Caenorhabditis elegans lin-25: cellular focus, protein expression and requirement for sur-2 during induction of vulval fates

Development ◽  
1998 ◽  
Vol 125 (23) ◽  
pp. 4809-4819 ◽  
Author(s):  
L. Nilsson ◽  
X. Li ◽  
T. Tiensuu ◽  
R. Auty ◽  
I. Greenwald ◽  
...  
Keyword(s):  
Protein Expression ◽  
Map Kinase ◽  
Signal Transduction Pathway ◽  
Precursor Cells ◽  
C Elegans ◽  
Novel Proteins ◽  
Genetic Mosaic ◽  
Mosaic Analysis ◽  
Map Kinase Pathway ◽  
Cell Fusions

Induction of vulval fates in the C. elegans hermaphrodite is mediated by a signal transduction pathway involving Ras and MAP kinase. Previous genetic analysis has suggested that two potential targets of this pathway in the vulva precursor cells are two novel proteins, LIN-25 and SUR-2. In this report, we describe further studies of lin-25. The results of a genetic mosaic analysis together with those of experiments in which lin-25 was expressed under the control of an heterologous promoter suggest that the major focus of lin-25 during vulva induction is the vulva precursor cells themselves. We have generated antisera to LIN-25 and used these to analyse the pattern of protein expression. LIN-25 is present in all six precursor cells prior to and during vulva induction but later becomes restricted to cells of the vulval lineages. Mutations in genes in the Ras/MAP kinase pathway do not affect the pattern of expression but the accumulation of LIN-25 is reduced in the absence of sur-2. Overexpression of LIN-25 does not rescue sur-2 mutant defects suggesting that LIN-25 and SUR-2 may function together. LIN-25 is also expressed in the lateral hypodermis. Overexpression of LIN-25 disrupts lateral hypodermal cell fusion, suggesting that lin-25 may play a role in regulating cell fusions in C. elegans.

scholarly journals Combining Auxin-Induced Degradation and RNAi Screening Identifies Novel Genes Involved in Lipid Bilayer Stress Sensing in Caenorhabditis elegans

2020 ◽  
Vol 10 (11) ◽  
pp. 3921-3928 ◽  
Author(s):  
Richard Venz ◽  
Anastasiia Korosteleva ◽  
Elisabeth Jongsma ◽  
Collin Y. Ewald
Keyword(s):  
Transcription Factors ◽  
Stress Response ◽  
Lipid Bilayer ◽  
Screening Strategy ◽  
Suppressor Genes ◽  
C Elegans ◽  
Link Type ◽  
Conserved Genes ◽  
Stress Sensing ◽  
The Unfolded Protein Response

Alteration of the lipid composition of biological membranes interferes with their function and can cause tissue damage by triggering apoptosis. Upon lipid bilayer stress, the endoplasmic reticulum mounts a stress response similar to the unfolded protein response. However, only a few genes are known to regulate lipid bilayer stress. We performed a suppressor screen that combined the auxin-inducible degradation (AID) system with conventional RNAi in C. elegans to identify members of the lipid bilayer stress response. AID-mediated degradation of the mediator MDT-15, a protein required for the upregulation of fatty acid desaturases, induced the activation of lipid bilayer stress-sensitive reporters. We screened through most C. elegans kinases and transcription factors by feeding RNAi. We discovered nine genes that suppressed the lipid bilayer stress response in C. elegans. These suppressor genes included drl-1/MAP3K3, gsk-3/GSK3, let-607/CREB3, ire-1/IRE1, and skn-1/NRF1,2,3. Our candidate suppressor genes suggest a network of transcription factors and the integration of multiple tissues for a centralized lipotoxicity response in the intestine. Thus, we demonstrated proof-of-concept for combining AID and RNAi as a new screening strategy and identified eight conserved genes that had not previously been implicated in the lipid bilayer stress response.

scholarly journals Calcineurin homologous proteins regulate the membrane localization and activity of sodium/proton exchangers in C. elegans

2016 ◽  
Vol 310 (3) ◽  
pp. C233-C242 ◽  
Author(s):  
Erik Allman ◽  
Qian Wang ◽  
Rachel L. Walker ◽  
Molly Austen ◽  
Maureen A. Peters ◽  
...  
Keyword(s):  
Genetic Model ◽  
Expression Patterns ◽  
Critical Role ◽  
Model Organism ◽  
Loss Of Function ◽  
Homologous Proteins ◽  
Relative Significance ◽  
C Elegans ◽  
Fluorescent Tags

Calcineurin B homologous proteins (CHP) are N-myristoylated, EF-hand Ca2+-binding proteins that bind to and regulate Na+/H+ exchangers, which occurs through a variety of mechanisms whose relative significance is incompletely understood. Like mammals, Caenorhabditis elegans has three CHP paralogs, but unlike mammals, worms can survive CHP loss-of-function. However, mutants for the CHP ortholog PBO-1 are unfit, and PBO-1 has been shown to be required for proton signaling by the basolateral Na+/H+ exchanger NHX-7 and for proton-coupled intestinal nutrient uptake by the apical Na+/H+ exchanger NHX-2. Here, we have used this genetic model organism to interrogate PBO-1's mechanism of action. Using fluorescent tags to monitor Na+/H+ exchanger trafficking and localization, we found that loss of either PBO-1 binding or activity caused NHX-7 to accumulate in late endosomes/lysosomes. In contrast, NHX-2 was stabilized at the apical membrane by a nonfunctional PBO-1 protein and was only internalized following its complete loss. Additionally, two pbo-1 paralogs were identified, and their expression patterns were analyzed. One of these contributed to the function of the excretory cell, which acts like a kidney in worms, establishing an alternative model for testing the role of this protein in membrane transporter trafficking and regulation. These results lead us to conclude that the role of CHP in Na+/H+ exchanger regulation differs between apical and basolateral transporters. This further emphasizes the importance of proper targeting of Na+/H+ exchangers and the critical role of CHP family proteins in this process.

scholarly journals Regulation of Membrane Trafficking by a Novel Cdc42-related Protein in Caenorhabditis elegans Epithelial Cells

2005 ◽  
Vol 16 (4) ◽  
pp. 1629-1639 ◽  
Author(s):  
S. Jenna ◽  
M.-E. Caruso ◽  
A. Emadali ◽  
D. T. Nguyên ◽  
M. Dominguez ◽  
...  
Keyword(s):  
Caenorhabditis Elegans ◽  
Epithelial Cells ◽  
Membrane Trafficking ◽  
Rho Gtpases ◽  
Related Protein ◽  
Recycling Endosomes ◽  
C Elegans ◽  
Apical Trafficking ◽  
Golgi Network

Rho GTPases are mainly known for their implication in cytoskeleton remodeling. They have also been recently shown to regulate various aspects of membrane trafficking. Here, we report the identification and the characterization of a novel Caenorhabditis elegans Cdc42-related protein, CRP-1, that shows atypical enzymatic characteristics in vitro. Expression in mouse fibroblasts revealed that, in contrast with CDC-42, CRP-1 was unable to reorganize the actin cytoskeleton and mainly localized to trans-Golgi network and recycling endosomes. This subcellular localization, as well as its expression profile restricted to a subset of epithelial-like cells in C. elegans, suggested a potential function for this protein in polarized membrane trafficking. Consistent with this hypothesis, alteration of CRP-1 expression affected the apical trafficking of CHE-14 in vulval and rectal epithelial cells and sphingolipids (C6-NBD-ceramide) uptake and/or trafficking in intestinal cells. However, it did not affect basolateral trafficking of myotactin in the pharynx and the targeting of IFB-2 and AJM-1, two cytosolic apical markers of intestine epithelial cells. Hence, our data demonstrate a function for CRP-1 in the regulation of membrane trafficking in a subset of cells with epithelial characteristics.

scholarly journals The FDA-approved drugs ticlopidine, sertaconazole, and dexlansoprazole can cause morphological changes in C. elegans

2020 ◽  
Author(s):  
Kyle F Galford ◽  
Antony M Jose
Keyword(s):  
Proton Pump Inhibitor ◽  
Genetic Model ◽  
Morphological Changes ◽  
Model System ◽  
Model Organisms ◽  
Regulatory Agencies ◽  
C Elegans ◽  
Therapeutic Drugs ◽  
Approved Drugs ◽  
Fda Approved Drugs

AbstractUrgent need for treatments limit studies of therapeutic drugs before approval by regulatory agencies. Analyses of drugs after approval can therefore improve our understanding of their mechanism of action and enable better therapies. We screened a library of 1443 Food and Drug Administration (FDA)-approved drugs using a simple assay in the nematode C. elegans and found three compounds that caused morphological changes. While the anticoagulant ticlopidine and the antifungal sertaconazole caused morphologically distinct pharyngeal defects upon acute exposure, the proton-pump inhibitor dexlansoprazole caused molting defects and required exposure during larval development. Such easily detectable defects in a powerful genetic model system advocate the continued exploration of current medicines using a variety of model organisms to better understand drugs already prescribed to millions of patients.

scholarly journals Hybrid assembly of the genome of the entomopathogenic nematode Steinernema carpocapsae identifies the X-chromosome

2019 ◽  
Author(s):  
Lorrayne Serra ◽  
Marissa Macchietto ◽  
Aide Macias-Muñoz ◽  
Cassandra Joan McGill ◽  
Isaryhia Maya Rodriguez ◽  
...  
Keyword(s):  
Gene Expression ◽  
X Chromosome ◽  
Genetic Model ◽  
Biological Control Agent ◽  
Gc Content ◽  
Steinernema Carpocapsae ◽  
Comparative Genomic ◽  
Chromosome X ◽  
C Elegans ◽  
Long Reads

AbstractEntomopathogenic nematodes from the genus Steinernema are lethal insect parasites that quickly kill their insect hosts with the help of their symbiotic bacteria. Steinernema carpocapsae is one of the most studied entomopathogens due to its broad lethality to diverse insect species and its effective commercial use as a biological control agent for insect pests, as well as a genetic model for studying parasitism, pathogenesis, and symbiosis. In this study, we used long-reads from the Pacific Biosciences platform and BioNano Genomics Irys system to assemble the best genome of S. carpocapsae ALL strain to date, comprising 84.5 Mb in 16 scaffolds, with an N50 of 7.36Mb. The largest scaffold, with 20.9Mb, was identified as chromosome X based on sex-specific genome sequencing. The high level of contiguity allowed us to characterize gene density, repeat content, and GC content. RNA-seq data from 17 developmental stages, spanning from embryo to adult, were used to predict 30,957 gene models. Using this new genome, we performed a macrosyntenic analysis to Caenorhabditis elegans and Pristionchus pacificus and found S. carpocapsae’s chromosome X to be primarily orthologous to C. elegans’ and P. pacificus’ chromosome II and IV. We also investigated the expansion of protein families and gene expression differences between male and female stage nematodes. This new genome and more accurate set of annotations provide a foundation for new comparative genomic and gene expression studies within the Steinernema clade and across the Nematoda phylum.Article SummaryThe insect killing worms Steinernema carpocapsae is a model organism for parasitism and symbiosis. The authors have used long reads and optical mapping to generate substantially contiguous assembly and a new set of gene annotations. They have identified the X chromosome as well as expansions in specific family proteases found in the venom of this worm. A macrosyntenic analysis with C. elegans shows a broad conservation of ancestral chromosomes with the exception of chromosome X. This new assembly will be useful to the Steinernema community and the broader nematode genomics community.

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