Detecting changes in the Caenorhabditis elegans intestinal environment using an engineered bacterial biosensor

AbstractCaenorhabditis elegans has become a key model organism within biology. In particular, the transparent gut, rapid growing time and ability to create a defined gut microbiota make it an ideal candidate organism for understanding and engineering the host microbiota. Here we present the development of an experimental model which can be used to characterise whole-cell bacterial biosensors in vivo. A dual-plasmid sensor system responding to isopropyl β-D-1-thiogalactopyranoside was developed and fully characterised in vitro. Subsequently, we show the sensor was capable of detecting and reporting on changes in the intestinal environment of C. elegans after introducing exogenous inducer into the environment. The protocols presented here may be used for aiding the rational design of engineered bacterial circuits, primarily for diagnostic applications. In addition, the model system may serve to reduce the use of current animal models and aid in the exploration of complex questions within general nematode and host-microbe biology.

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A fast and reliable method for monitoring genomic instability in the model organism Caenorhabditis elegans

Archives of Toxicology ◽

10.1007/s00204-021-03144-7 ◽

2021 ◽

Author(s):

Merle Marie Nicolai ◽

Barbara Witt ◽

Andrea Hartwig ◽

Tanja Schwerdtle ◽

Julia Bornhorst

Keyword(s):

Caenorhabditis Elegans ◽

Animal Experiments ◽

Model Organism ◽

Animal Testing ◽

C Elegans ◽

Testing Strategies ◽

Genotoxic Agents ◽

Genotoxicity Testing

AbstractThe identification of genotoxic agents and their potential for genotoxic alterations in an organism is crucial for risk assessment and approval procedures of the chemical and pharmaceutical industry. Classically, testing strategies for DNA or chromosomal damage focus on in vitro and in vivo (mainly rodent) investigations. In cell culture systems, the alkaline unwinding (AU) assay is one of the well-established methods for detecting the percentage of double-stranded DNA (dsDNA). By establishing a reliable lysis protocol, and further optimization of the AU assay for the model organism Caenorhabditis elegans (C. elegans), we provided a new tool for genotoxicity testing in the niche between in vitro and rodent experiments. The method is intended to complement existing testing strategies by a multicellular organism, which allows higher predictability of genotoxic potential compared to in vitro cell line or bacterial investigations, before utilizing in vivo (rodent) investigations. This also allows working within the 3R concept (reduction, refinement, and replacement of animal experiments), by reducing and possibly replacing animal testing. Validation with known genotoxic agents (bleomycin (BLM) and tert-butyl hydroperoxide (tBOOH)) proved the method to be meaningful, reproducible, and feasible for high-throughput genotoxicity testing, and especially preliminary screening.

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Caenorhabditis elegans: A promising contrivance to study neurotoxicity and oxidative stress

Research Journal of Biotechnology ◽

10.25303/1610rjbt198206 ◽

2021 ◽

Vol 16 (10) ◽

pp. 198-206

Author(s):

Kiran Singh ◽

Shweta Yadav

Keyword(s):

Oxidative Stress ◽

Caenorhabditis Elegans ◽

Genetic Manipulation ◽

Model Organism ◽

Mechanisms Of Toxicity ◽

C Elegans ◽

Toxicological Studies ◽

And Oxidative Stress

Owing to ubiquitous distribution, high abundances and ecological relevance, Caenorhabditis elegans has strong potential interest as barometer of environment and human health. Ecotoxicological methods are used to evaluate the effect of various anthropogenic contaminants on the ecosystems that circumscribe both in-vivo and in-vitro toxicities to explore the pathways and mechanisms of toxicity and to set precise toxicity thresholds. The interest in C. elegans, as a model organism in toxicological studies, has increased over the past few decades. The enticement of C. elegans comes from the ease of metabolically active digestive, sensory, endocrine, neuromuscular, reproductive systems and genetic manipulation along with the ability to ﬂuorescently label neuronal subtypes. The study reviews the competence of Caenorhabditis elegans as a potential model organism in various toxicity assays specifically neurotoxicity and oxidative stress.

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Identification, functional expression and enzymic analysis of two distinct CaaX proteases from Caenorhabditis elegans

Biochemical Journal ◽

10.1042/bj20021514 ◽

2003 ◽

Vol 370 (3) ◽

pp. 1047-1054 ◽

Cited By ~ 21

Author(s):

Juan CADIÑANOS ◽

Walter K. SCHMIDT ◽

Antonio FUEYO ◽

Ignacio VARELA ◽

Carlos LÓPEZ-OTÍN ◽

...

Keyword(s):

Caenorhabditis Elegans ◽

Model Organism ◽

Functional Expression ◽

Cysteine Residue ◽

C Elegans ◽

Ras Gtpases ◽

Acid Protein ◽

Bona Fide

Post-translational processing of proteins such as the Ras GTPases, which contain a C-terminal CaaX motif (where C stands for cysteine, a for aliphatic and X is one of several amino acids), includes prenylation, proteolytic removal of the C-terminal tripeptide and carboxy-methylation of the isoprenyl-cysteine residue. In the present study, we report the presence of two distinct CaaX-proteolytic activities in membrane extracts from Caenorhabditis elegans, which are sensitive to EDTA and Tos-Phe-CH2Cl (tosylphenylalanylchloromethane; ‘TPCK') respectively. A protein similar to the mammalian and yeast farnesylated-proteins converting enzyme-1 (FACE-1)/Ste24p CaaX metalloprotease, encoded by a hypothetical gene (CeFACE-1/C04F12.10) found in C. elegans chromosome I, probably accounts for the EDTA-sensitive activity. An orthologue of FACE-2/Rce1p, the enzyme responsible for the proteolytic maturation of Ras oncoproteins and other prenylated substrates, probably accounts for the Tos-Phe-CH2Cl-sensitive activity, even though the gene for FACE-2/Rce1 has not been previously identified in this model organism. We have identified a previously overlooked gene in C. elegans chromosome V, which codes for a 266-amino-acid protein (CeFACE-2) with 30% sequence identity to human FACE-2/Rce1. We show that both CeFACE-1 and CeFACE-2 have the ability to promote production of the farnesylated yeast pheromone a-factor in vivo and to cleave a farnesylated peptide in vitro. These results indicate that CeFACE-1 and CeFACE-2 are bona fide CaaX proteases and support the evolutionary conservation of this proteolytic system in eukaryotes.

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Promoters of the dhs-21 gene encoding dicarbonyl/l-xylulose reductase in Caenorhabditis elegans

Vietnam Journal of Biotechnology ◽

10.15625/1811-4989/15/2/12353 ◽

2018 ◽

Vol 15 (2) ◽

pp. 359-365

Author(s):

Lê Thọ Sơn ◽

Joohong Ahnn ◽

Jeong Hoon Cho ◽

Nguyễn Huy Hoàng

Keyword(s):

Caenorhabditis Elegans ◽

Model Organism ◽

Biological Research ◽

Loss Of Function ◽

Wild Type ◽

C Elegans ◽

Larval Stages ◽

Vital Functions

Dicarbonyl/L-xylulose (DCXR) was identified as a dehydrogenase. This type of enzyme was presented in various forms of lives including bacteria, fungi, plants and animals. Generally, it converts L-xylulose to xylitol in the presence of either cofactor NADH or NADPH in vitro. Previous studies reported the biochemistry properties and crystal structure but largely uncovered biological roles of DCXRs. It was impossible to dissect the functions in mice or human cells that had many DCXR homologs in their genomes. Interestingly, the wild-type Caenorhabditis elegans, a well-known model organism in biological research, has only nuclear genomic dhs-21 that encodes a unique homologous DCXR. Thus Ce.dhs-21 and the host C. elegans were relevant for investigation of the physiologically-vital functions of the DCXR. This research aimed to the expression of dhs-21 in vivo. We defined three promoters , manipulated three relative reporter-constructs that conjugated the dhs-21 gene and Green Flouresent Protein (known as GFP) one. The construct vectors were transferred into wild-type C. elegans N2 and as well as the hermaphroditic loss of function dhs-21(jh129) by microinjection. In the results, we found that the expression pattern of dhs-21 under the only p2-promoter construct was stable and similar to immunogold Electric Microscopy (EM) images. The dhs-21 gene was expressed in both sexes of at all larval stages till the deaths of worms. DHS-21 was expressed in the cytosol of the intestinal, gonad sheath and uterous seam cell (utse).

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Caenorhabditis elegans as an in vivo Model to Assess Amphetamine Tolerance

Brain Behavior and Evolution ◽

10.1159/000514858 ◽

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.

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Identification of gmhA, a Yersinia pestis Gene Required for Flea Blockage, by Using a Caenorhabditis elegans Biofilm System

Infection and Immunity ◽

10.1128/iai.73.11.7236-7242.2005 ◽

2005 ◽

Vol 73 (11) ◽

pp. 7236-7242 ◽

Cited By ~ 55

Author(s):

Creg Darby ◽

Sandya L. Ananth ◽

Li Tan ◽

B. Joseph Hinnebusch

Keyword(s):

Caenorhabditis Elegans ◽

Yersinia Pestis ◽

Biofilm Formation ◽

Yersinia Pseudotuberculosis ◽

C Elegans ◽

Transposon Insertion ◽

Insertion Mutants ◽

Random Screening

ABSTRACT Yersinia pestis, the cause of bubonic plague, blocks feeding by its vector, the flea. Recent evidence indicates that blockage is mediated by an in vivo biofilm. Y. pestis and the closely related Yersinia pseudotuberculosis also make biofilms on the cuticle of the nematode Caenorhabditis elegans, which block this laboratory animal's feeding. Random screening of Y. pseudotuberculosis transposon insertion mutants with a C. elegans biofilm assay identified gmhA as a gene required for normal biofilms. gmhA encodes phosphoheptose isomerase, an enzyme required for synthesis of heptose, a conserved component of lipopolysaccharide and lipooligosaccharide. A Y. pestis gmhA mutant was constructed and was severely defective for C. elegans biofilm formation and for flea blockage but only moderately defective in an in vitro biofilm assay. These results validate use of the C. elegans biofilm system to identify genes and pathways involved in Y. pestis flea blockage.

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Rosemary Flowers as Edible Plant Foods: Phenolic Composition and Antioxidant Properties in Caenorhabditis elegans

Antioxidants ◽

10.3390/antiox9090811 ◽

2020 ◽

Vol 9 (9) ◽

pp. 811

Author(s):

Cristina Moliner ◽

Víctor López ◽

Lillian Barros ◽

Maria Inês Dias ◽

Isabel C. F. R. Ferreira ◽

...

Keyword(s):

Caenorhabditis Elegans ◽

Antioxidant Properties ◽

Ethanolic Extract ◽

Food Ingredient ◽

Edible Plant ◽

Phenolic Profile ◽

C Elegans ◽

Rosmarinus Officinalis L

Rosmarinus officinalis L., commonly known as rosemary, has been largely studied for its wide use as food ingredient and medicinal plant; less attention has been given to its edible flowers, being necessary to evaluate their potential as functional foods or nutraceuticals. To achieve that, the phenolic profile of the ethanolic extract of R. officinalis flowers was determined using LC-DAD-ESI/MSn and then its antioxidant and anti-ageing potential was studied through in vitro and in vivo assays using Caenorhabditis elegans. The phenolic content was 14.3 ± 0.1 mg/g extract, trans rosmarinic acid being the predominant compound in the extract, which also exhibited a strong antioxidant capacity in vitro and increased the survival rate of C. elegans exposed to lethal oxidative stress. Moreover, R. officinalis flowers extended C. elegans lifespan up to 18%. Therefore, these findings support the potential use of R. officinalis flowers as ingredients to develop products with pharmaceutical and/or nutraceutical potential.

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Mulberry Anthocyanin Extract Ameliorates Oxidative Damage in HepG2 Cells and Prolongs the Lifespan of Caenorhabditis elegans through MAPK and Nrf2 Pathways

Oxidative Medicine and Cellular Longevity ◽

10.1155/2017/7956158 ◽

2017 ◽

Vol 2017 ◽

pp. 1-12 ◽

Cited By ~ 19

Author(s):

Fujie Yan ◽

Yushu Chen ◽

Ramila Azat ◽

Xiaodong Zheng

Keyword(s):

Oxidative Stress ◽

Caenorhabditis Elegans ◽

Hepg2 Cells ◽

Insulin Signalling ◽

Glucose Consumption ◽

Redox Status ◽

C Elegans ◽

Beneficial Effects

Mulberry anthocyanins possess many pharmacological effects including liver protection, anti-inflammation, and anticancer. The aim of this study was to evaluate whether mulberry anthocyanin extract (MAE) exerts beneficial effects against oxidative stress damage in HepG2 cells and Caenorhabditis elegans. In vitro, MAE prevented cytotoxicity, increased glucose consumption and uptake, and eliminated excessive intracellular free radicals in H2O2-induced cells. Moreover, MAE pretreatment maintained Nrf2, HO-1, and p38 MAPK stimulation and abolished upregulation of p-JNK, FOXO1, and PGC-1α that were involved in oxidative stress and insulin signalling modulation. In vivo, extended lifespan was observed in C. elegans damaged by paraquat in the presence of MAE, while these beneficial effects were disappeared in pmk-1 and daf-16 mutants. PMK-1 and SKN-1 were activated after exposure to paraquat and MAE suppressed PMK-1 activation but enhanced SKN-1 stimulation. Our findings suggested that MAE recovered redox status in HepG2 cells and C. elegans that suffered from oxidative stress, which might be by targeting MAPKs and Nrf2.

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Identification of phenolic secondary metabolites from Schotia brachypetala Sond. (Fabaceae) and demonstration of their antioxidant activities in Caenorhabditis elegans

10.7287/peerj.preprints.1768v1 ◽

2016 ◽

Author(s):

Mansour Sobeh ◽

Esraa A ElHawary ◽

Herbenya Peixoto ◽

Rola M Labib ◽

Heba Handoussa ◽

...

Keyword(s):

Antioxidant Activity ◽

Caenorhabditis Elegans ◽

Nuclear Translocation ◽

Antioxidant Activities ◽

In Vitro Assays ◽

Flavonoid Glycosides ◽

Alcoholic Extract ◽

C Elegans

Background: Schotia brachypetala Sond. (Fabaceae) is an endemic tree of Southern Africa whose phytochemistry and pharmacology were slightly studied.The present work aimed at profiling the major phenolics compounds present in the hydro-alcoholic extract from S. brachypetala leaves (SBE) using LC/HRESI/MS/MS and NMR and prove their antioxidant capabilities using novel methods. Methods: In vitro assays; DPPH, TEAC persulfate decolorizing kinetic and FRAP assays, and in vivo assays: Caenorhabditis elegans strains maintenance, Intracellular ROS in C. elegans, Survival assay, GFP expression and Subcellular DAF-16 localization were employed to evaluate the antioxidant activity. Results: More than forty polyphenols ,including flavonoid glycosides, galloylated flavonoid glycosides, isoflavones, dihydrochalcones, procyanidins, anthocyanins, hydroxybenzoic acid derivatives, hydrolysable tannins, and traces of methylated and acetylated flavonoid derivatives were identified. Three compounds were isolated and identified from the genus Schotia for the first time, namely gallic acid, myricetin-3-O-α-L-1C4-rhamnoside and quercetin-3-O-L-1C4-rhamnoside.The tested extract was able to protect the worms against juglone induced oxidative stress and attenuate the reactive oxygen species (ROS) accumulation. SBE was also able to attenuate the levels of heat shock protein (HSP) expression. Discussion: A pronounced antioxidant activity in vivo, which can be attributed to its ability to promote the nuclear translocation of DAF-16/FOXO, the main transcription factor regulating the expression of stress response genes. The remarkable antioxidant activity in vitro and in vivo correlates to SBE rich phenolic profile.

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Polyphenols from Blumea laciniata Extended the Lifespan and Enhanced Resistance to Stress in Caenorhabditis elegans via the Insulin Signaling Pathway

Antioxidants ◽

10.3390/antiox10111744 ◽

2021 ◽

Vol 10 (11) ◽

pp. 1744

Author(s):

Tao Chen ◽

Siyuan Luo ◽

Xiaoju Wang ◽

Yiling Zhou ◽

Yali Dai ◽

...

Keyword(s):

Caenorhabditis Elegans ◽

Signaling Pathway ◽

Relevant Literature ◽

Folk Medicine ◽

Ageing Effect ◽

C Elegans ◽

Enhanced Resistance ◽

Resistance To Stress

Blumea laciniata is widely used as a folk medicine in Asia, but relevant literature on it is rarely reported. We confirmed that polyphenol extract (containing chlorogenic acid, rutin, and luteolin-4-O-glucoside) from B. laciniata (EBL) showed strong antioxidant ability in vitro. Hence, in this work, we applied Caenorhabditis elegans to further investigate the antioxidant and anti-ageing abilities of EBL in vivo. The results showed that EBL enhanced the survival of C. elegans under thermal stress by 12.62% and sharply reduced the reactive oxygen species level as well as the content of malonaldehyde. Moreover, EBL increased the activities of antioxidant enzymes such as catalase and superoxide dismutase. Additionally, EBL promoted DAF-16, a transcription factor, into the nucleus. Besides, EBL extended the lifespan of C. elegans by 17.39%, showing an anti-ageing effect. Different mutants indicated that the insulin/IGF-1 signaling pathway participated in the antioxidant and anti-ageing effect of EBL on C. elegans.

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