Punctuated loci on chromosome IV determine natural variation in Orsay virus susceptibility of Caenorhabditis elegans strains Bristol N2 and Hawaiian CB4856

AbstractHost-pathogen interactions play a major role in evolutionary selection and shape natural genetic variation. The genetically distinct Caenorhabditis elegans strains, Bristol N2 and Hawaiian CB4856, are differentially susceptible to the Orsay virus (OrV). Here we report the dissection of the genetic architecture of susceptibility to OrV infection. We compare OrV infection in the relatively resistant wild-type CB4856 strain to the more susceptible canonical N2 strain. To gain insight into the genetic architecture of viral susceptibility, 52 fully sequenced recombinant inbred lines (CB4856 x N2 RILs) were exposed to OrV. This led to the identification of two loci on chromosome IV associated with OrV resistance. To verify the two loci and gain additional insight into the genetic architecture controlling virus infection, introgression lines (ILs) that together cover chromosome IV, were exposed to OrV. Of the 27 ILs used, 17 had an CB4856 introgression in an N2 background and 10 had an N2 introgression in a CB4856 background. Infection of the ILs confirmed and fine-mapped the locus underlying variation in OrV susceptibility and we found that a single nucleotide polymorphism in cul-6 contributes to the difference in OrV susceptibility between N2 and CB4856. An allele swap experiment showed the strain CB4856 became more susceptible by having an N2 cul-6 allele, although having the CB4856 cul-6 allele did not increase resistance in N2. Additionally, we found that multiple strains with non-overlapping introgressions showed a distinct infection phenotype from the parental strain, indicating that there are punctuated locations on chromosome IV determining OrV susceptibility. Thus, our findings reveal the genetic complexity of OrV susceptibility in C. elegans and suggest that viral susceptibility is governed by multiple genes.

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Punctuated loci on chromosome IV determine natural variation in Orsay virus susceptibility of Caenorhabditis elegans strains Bristol N2 and Hawaiian CB4856

Journal of Virology ◽

10.1128/jvi.02430-20 ◽

2021 ◽

Author(s):

Mark G. Sterken ◽

Lisa van Sluijs ◽

Yiru A. Wang ◽

Wannisa Ritmahan ◽

Mitra L. Gultom ◽

...

Keyword(s):

Genetic Variation ◽

Caenorhabditis Elegans ◽

Virus Infection ◽

Genetic Architecture ◽

Recombinant Inbred Lines ◽

Model Organism ◽

C Elegans ◽

Viral Susceptibility ◽

Orsay Virus ◽

Insight Into

Host-pathogen interactions play a major role in evolutionary selection and shape natural genetic variation. The genetically distinct Caenorhabditis elegans strains, Bristol N2 and Hawaiian CB4856, are differentially susceptible to the Orsay virus (OrV). Here we report the dissection of the genetic architecture of susceptibility to OrV infection. We compare OrV infection in the relatively resistant wild-type CB4856 strain to the more susceptible canonical N2 strain. To gain insight into the genetic architecture of viral susceptibility, 52 fully sequenced recombinant inbred lines (CB4856 x N2 RILs) were exposed to OrV. This led to the identification of two loci on chromosome IV associated with OrV resistance. To verify the two loci and gain additional insight into the genetic architecture controlling virus infection, introgression lines (ILs) that together cover chromosome IV, were exposed to OrV. Of the 27 ILs used, 17 had an CB4856 introgression in an N2 background and 10 had an N2 introgression in a CB4856 background. Infection of the ILs confirmed and fine-mapped the locus underlying variation in OrV susceptibility and we found that a single nucleotide polymorphism in cul-6 may contribute to the difference in OrV susceptibility between N2 and CB4856. An allele swap experiment showed the strain CB4856 became as susceptible as the N2 strain by having an N2 cul-6 allele, although having the CB4856 cul-6 allele did not increase resistance in N2. Additionally, we found that multiple strains with non-overlapping introgressions showed a distinct infection phenotype from the parental strain, indicating that there are punctuated locations on chromosome IV determining OrV susceptibility. Thus, our findings reveal the genetic complexity of OrV susceptibility in C. elegans and suggest that viral susceptibility is governed by multiple genes. Importance Genetic variation determines the viral susceptibility of hosts. Yet, pinpointing which genetic variants determine viral susceptibility remains challenging. Here, we have exploited the genetic tractability of the model organism C. elegans to dissect the genetic architecture of Orsay virus infection. Our results provide novel insight into natural determinants of Orsay virus infection.

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Heat Stress Reduces the Susceptibility of Caenorhabditis elegans to Orsay Virus Infection

Genes ◽

10.3390/genes12081161 ◽

2021 ◽

Vol 12 (8) ◽

pp. 1161

Author(s):

Yuqing Huang ◽

Mark G. Sterken ◽

Koen van Zwet ◽

Lisa van Sluijs ◽

Gorben P. Pijlman ◽

...

Keyword(s):

Heat Stress ◽

Caenorhabditis Elegans ◽

Virus Infection ◽

Intracellular Pathogen ◽

Potential Candidate ◽

Molecular Responses ◽

C Elegans ◽

Pathogen Response ◽

Orsay Virus ◽

Natural Virus

The nematode Caenorhabditis elegans has been a versatile model for understanding the molecular responses to abiotic stress and pathogens. In particular, the response to heat stress and virus infection has been studied in detail. The Orsay virus (OrV) is a natural virus of C. elegans and infection leads to intracellular infection and proteostatic stress, which activates the intracellular pathogen response (IPR). IPR related gene expression is regulated by the genes pals-22 and pals-25, which also control thermotolerance and immunity against other natural pathogens. So far, we have a limited understanding of the molecular responses upon the combined exposure to heat stress and virus infection. We test the hypothesis that the response of C. elegans to OrV infection and heat stress are co-regulated and may affect each other. We conducted a combined heat-stress-virus infection assay and found that after applying heat stress, the susceptibility of C. elegans to OrV was decreased. This difference was found across different wild types of C. elegans. Transcriptome analysis revealed a list of potential candidate genes associated with heat stress and OrV infection. Subsequent mutant screens suggest that pals-22 provides a link between viral response and heat stress, leading to enhanced OrV tolerance of C. elegans after heat stress.

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

mBio ◽

10.1128/mbio.00940-17 ◽

2017 ◽

Vol 8 (5) ◽

Cited By ~ 13

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.

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The Caenorhabditis elegans DEG-3/DES-2 Channel Is a Betaine-Gated Receptor Insensitive to Monepantel

Molecules ◽

10.3390/molecules27010312 ◽

2022 ◽

Vol 27 (1) ◽

pp. 312

Author(s):

Tina V. A. Hansen ◽

Heinz Sager ◽

Céline E. Toutain ◽

Elise Courtot ◽

Cédric Neveu ◽

...

Keyword(s):

Caenorhabditis Elegans ◽

Nicotinic Acetylcholine Receptors ◽

Acetylcholine Receptors ◽

Parasitic Nematode ◽

Nematode Species ◽

Xenopus Laevis Oocytes ◽

Allosteric Modulators ◽

C Elegans ◽

Electrode Voltage ◽

Insight Into

Natural plant compounds, such as betaine, are described to have nematocidal properties. Betaine also acts as a neurotransmitter in the free-living model nematode Caenorhabditis elegans, where it is required for normal motility. Worm motility is mediated by nicotinic acetylcholine receptors (nAChRs), including subunits from the nematode-specific DEG-3 group. Not all types of nAChRs in this group are associated with motility, and one of these is the DEG-3/DES-2 channel from C. elegans, which is involved in nociception and possibly chemotaxis. Interestingly, the activity of DEG-3/DES-2 channel from the parasitic nematode of ruminants, Haemonchus contortus, is modulated by monepantel and its sulfone metabolite, which belong to the amino-acetonitrile derivative anthelmintic drug class. Here, our aim was to advance the pharmacological knowledge of the DEG-3/DES-2 channel from C. elegans by functionally expressing the DEG-3/DES-2 channel in Xenopus laevis oocytes and using two-electrode voltage-clamp electrophysiology. We found that the DEG-3/DES-2 channel was more sensitive to betaine than ACh and choline, but insensitive to monepantel and monepantel sulfone when used as direct agonists and as allosteric modulators in co-application with betaine. These findings provide important insight into the pharmacology of DEG-3/DES-2 from C. elegans and highlight the pharmacological differences between non-parasitic and parasitic nematode species.

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New strains for tissue-specific RNAi studies in Caenorhabditis elegans

10.1101/283325 ◽

2018 ◽

Author(s):

Jason S. Watts ◽

Henry F. Harrison ◽

Shizue Omi ◽

Quentin Guenthers ◽

James Dalelio ◽

...

Keyword(s):

Fatty Acid ◽

Caenorhabditis Elegans ◽

Gene Function ◽

Target Genes ◽

Resistant Mutant ◽

Knockout Mutant ◽

Tissue Specific ◽

Double Stranded Rna ◽

C Elegans ◽

Insight Into

AbstractRNA interference is a powerful tool for dissecting gene function. In Caenorhabditis elegans, ingestion of double stranded RNA causes strong, systemic knockdown of target genes. Further insight into gene function can be revealed by tissue-specific RNAi techniques. Currently available tissue-specific C. elegans strains rely on rescue of RNAi function in a desired tissue or cell in an otherwise RNAi deficient genetic background. We attempted to assess the contribution of specific tissues to polyunsaturated fatty acid (PUFA) synthesis using currently available tissue-specific RNAi strains. We discovered that rde-1 (ne219), a commonly used RNAi-resistant mutant strain, retains considerable RNAi capacity against RNAi directed at PUFA synthesis genes. By measuring changes in the fatty acid products of the desaturase enzymes that synthesize PUFAs, we found that the before mentioned strain, rde-1 (ne219) and the reported germline only RNAi strain, rrf-1 (pk1417) are not appropriate genetic backgrounds for tissue-specific RNAi experiments. However, the knockout mutant rde-1 (ne300) was strongly resistant to dsRNA induced RNAi, and thus is more appropriate for construction of a robust tissue-specific RNAi strains. Using newly constructed strains in the rde-1(null) background, we found considerable desaturase activity in intestinal, epidermal, and germline tissues, but not in muscle. The RNAi-specific strains reported in this study will be useful tools for C. elegans researchers studying a variety of biological processes.

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Feeding recombinant Royalactin: Measures of improved lifespan and mitochondrial function in Caenorhabditis elegans

10.1101/2020.09.14.296053 ◽

2020 ◽

Author(s):

Xia Li ◽

Thomas L. Ingram ◽

Ying Wang ◽

Kamila Derecka ◽

Nathan Courtier ◽

...

Keyword(s):

Caenorhabditis Elegans ◽

Mitochondrial Function ◽

Royal Jelly ◽

Biological Functions ◽

C Elegans ◽

Beneficial Effects ◽

Physiological Processes ◽

Royal Jelly Protein ◽

Insight Into ◽

Over Time

AbstractAgeing, the decline of biological functions over time, is inherent to eukaryotes. Female honeybees attain a long-lived queen phenotype upon continuous consumption of royal jelly, whereas restricted supply of this nutritional substance promotes the development of worker bees, which are short-lived. An abundant protein found within royal jelly is major royal jelly protein 1 (MRJP1), also known as ‘Royalactin’. Health- and lifespan promoting effects have been attributed to Royalactin in species from diverse animal taxa, suggesting it acts on phylogenetically conserved physiological processes. Here, we explore the effects of feeding the nematode Caenorhabditis elegans with Escherichia coli that express a recombinant form of Royalactin (RArec). We confirm that consumption of RArec increases body size, improves locomotion and extends lifespan. We discover a link between Royalactin and mitochondria, organelles which play a key part in the ageing process: both spare respiratory capacity and morphology indicate improved mitochondrial function in RArec fed C. elegans. These results demonstrate the feasibility of using recombinant Royalactin to gain further insight into processes of healthy ageing in many species.RArec production allows insight into potential beneficial effects across species.

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Natural genetic variation drives microbiome selection in the Caenorhabditis elegans gut

10.1101/2021.03.12.435148 ◽

2021 ◽

Author(s):

Fan Zhang ◽

Jessica L. Weckhorst ◽

Adrien Assié ◽

Ciara Hosea ◽

Christopher A. Ayoub ◽

...

Keyword(s):

Genetic Variation ◽

Caenorhabditis Elegans ◽

Signaling Pathways ◽

Insulin Signaling ◽

Natural Genetic Variation ◽

Microbiome Composition ◽

C Elegans ◽

Free Living Nematode ◽

Wild Strains ◽

The Stability

Host genetic landscapes can shape microbiome assembly in the animal gut by contributing to the establishment of distinct physiological environments. However, the genetic determinants contributing to the stability and variation of these microbiome types remain largely undefined. Here, we use the free-living nematode Caenorhabditis elegans to identify natural genetic variation among wild strains of C. elegans strains that drives assembly of distinct microbiomes. To achieve this, we first established a diverse model microbiome that represents the phylogenetic and functional diversity naturally found in the C. elegans microbiome. Using this community, we show that C. elegans utilizes immune, xenobiotic and metabolic signaling pathways to favor the assembly of different microbiome types. Variations in these pathways were associated with the enrichment for specific commensals, including the Alphaproteobacteria Ochrobactrum. Using RNAi and mutant strains, we showed that host selection for Ochrobactrum is mediated specifically by host insulin signaling pathways. Ochrobactrum recruitment is blunted in the absence of daf-2/IGFR and requires the insulin signaling transcription factors daf-16/FOXO and pqm-1/SALL2. Further, the ability of C. elegans to enrich for Ochrobactrum is correlated positively with host outcomes, as animals that develop faster are larger and have higher gut Ochrobactrum colonization as adults. These results highlight a new role for the highly conserved insulin signaling pathways in the regulation of microbiome composition in C. elegans.

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The genetic architecture underlying body-size traits plasticity over different temperatures and developmental stages in Caenorhabditis elegans

10.1101/2021.04.21.440777 ◽

2021 ◽

Author(s):

Muhammad I. Maulana ◽

Joost A.G. Riksen ◽

Basten L. Snoek ◽

Jan E. Kammenga ◽

Mark G. Sterken

Keyword(s):

Body Size ◽

Caenorhabditis Elegans ◽

Genetic Architecture ◽

Developmental Stages ◽

Recombinant Inbred Lines ◽

The Body ◽

Body Volume ◽

Chromosome X ◽

Different Temperatures ◽

Length Width

Most ectotherms obey the temperature-size rule, meaning they grow larger in a colder environment. This raises the question of how the interplay between genes and temperature affect the body size of ectotherms. Despite the growing body of literature on the physiological life-history and molecular genetic mechanism underlying the temperature-size rule, the overall genetic architecture orchestrating this complex phenotype is not yet fully understood. One approach to identify genetic regulators of complex phenotypes is Quantitative Trait Locus (QTL) mapping. Here, we explore the genetic architecture of body size phenotypes, in different temperatures using Caenorhabditis elegans as a model ectotherm. We used 40 recombinant inbred lines (RILs) derived from N2 and CB4856, which were reared at four different temperatures (16oC, 20oC, 24oC, and 26oC) and measured at two developmental stages (L4 and adult). The animals were measured for body length, width at vulva, body volume, length/width ratio, and seven other body-size traits. The genetically diverse RILs varied in their body-size phenotypes with heritabilities ranging from 0.0 to 0.99. We detected 18 QTL underlying the body-size phenotypes across all treatment combinations, with the majority clustering on chromosome X. We hypothesize that the chromosome X QTL could result from a known pleiotropic regulator - npr-1 - known to affect the body size of C. elegans through behavioural changes. In conclusion, our findings shed more light on multiple loci affecting body size plasticity and allow for a more refined analysis of the temperature-size rule.

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The ancestral C. elegans cuticle suppresses rol-1

10.1101/2020.02.07.938696 ◽

2020 ◽

Author(s):

Luke M. Noble ◽

Asif Miah ◽

Taniya Kaur ◽

Matthew V. Rockman

Keyword(s):

Genetic Variation ◽

Caenorhabditis Elegans ◽

Comparative Genomics ◽

Linkage Mapping ◽

Genetic Background ◽

Gene Editing ◽

Wild Strain ◽

Collagen Gene ◽

Natural Genetic Variation ◽

C Elegans

ABSTRACTGenetic background commonly modifies the effects of mutations. We discovered that worms mutant for the canonical rol-1 gene, identified by Brenner in 1974, do not roll in the genetic background of the wild strain CB4856. Using linkage mapping, association analysis and gene editing, we determined that N2 carries an insertion in the collagen gene col-182 that acts as a recessive enhancer of rol-1 rolling. From population and comparative genomics, we infer the insertion is derived in N2 and related laboratory lines, likely arising during the domestication of Caenorhabditis elegans, and breaking a conserved protein. The ancestral version of col-182 also modifies the phenotypes of four other classical cuticle mutant alleles, and the effects of natural genetic variation on worm shape and locomotion. These results underscore the importance of genetic background and the serendipity of Brenner’s choice of strain.

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Natural variation in the roles of C. elegans autophagy components during microsporidia infection

10.1101/418749 ◽

2018 ◽

Author(s):

Keir M. Balla ◽

Vladimir Lažetić ◽

Emily Troemel

Keyword(s):

Genetic Variation ◽

Caenorhabditis Elegans ◽

Natural Variation ◽

Early Stage ◽

Laboratory Strain ◽

Host Strain ◽

Standard Laboratory ◽

Natural Genetic Variation ◽

C Elegans ◽

Natural Variant

AbstractNatural genetic variation can determine the outcome of an infection, and often reflects the co-evolutionary battle between hosts and pathogens. We previously found that a natural variant of the nematode Caenorhabditis elegans from Hawaii (HW) has increased resistance against natural microsporidian pathogens in the Nematocida genus, when compared to the standard laboratory strain of N2. In particular, HW animals can clear infection, while N2 animals cannot. In addition, HW animals have lower levels of intracellular colonization of Nematocida compared to N2. Here we investigate how this natural variation in resistance relates to autophagy. We found that there is much better targeting of autophagy-related machinery to parasites under conditions where they are cleared. In particular, ubiquitin targeting to Nematocida cells correlates very well with their subsequent clearance in terms of timing, host strain and age, as well as Nematocida species. Furthermore, clearance correlates with targeting of the LGG-2/LC3 autophagy protein to parasite cells, with HW animals having much more efficient targeting of LGG-2 to parasite cells than N2 animals. Surprisingly, however, we found that lgg-2 is not required to clear infection. Instead we found that loss of lgg-2 leads to increased intracellular colonization in the HW background, although interestingly, it does not affect colonization in the N2 background. Altogether our results suggest that there is natural genetic variation in an lgg-2-dependent process that regulates intracellular levels of microsporidia at a very early stage of infection prior to clearance.

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