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Heatmap of transcription profiles of selected gene families with multiple induced family members. The heatmap showed the log 2 counts per million (CPM) for each gene in either C17H1 gene family or DUF713 gene family. Each experimental condition had three replicates and each replicate was represented in a column
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Background
Caenorhabditis elegans is a powerful model organism for probing many biological processes including host-pathogen interactions with bacteria and fungi. The recent identification of nematode viruses that naturally infect C. elegans and Caenorhabditis briggsae provides a unique opportunity to define host-virus interactions in these model h...
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Context 1
... and DUF713 domain genes, more than 50% of the family members were differentially expressed. The C17H1 gene family has a total of 36 members in C. elegans, and of those the same 25 members (except for F22G12.7 in Orsay virus [N2] condition that was not statistically significant) were up-regulated following both Orsay virus and N. parisii infection (Fig. 2, Table 2). The DUF713 domain genes have a total of 10 members in C. elegans and have from 5-9 members of the gene family up-regulated following pathogen infection (Fig. 2, Table 2). The C17H1 family had the most DEGs repre- sented in the Orsay virus and N. parisii infections (Table 2), and some of the genes were among the highest ...
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... same 25 members (except for F22G12.7 in Orsay virus [N2] condition that was not statistically significant) were up-regulated following both Orsay virus and N. parisii infection (Fig. 2, Table 2). The DUF713 domain genes have a total of 10 members in C. elegans and have from 5-9 members of the gene family up-regulated following pathogen infection (Fig. 2, Table 2). The C17H1 family had the most DEGs repre- sented in the Orsay virus and N. parisii infections (Table 2), and some of the genes were among the highest induced with close to 1000-fold increase compared to mock infection. Given the highly distinct nature of Orsay virus from the eukaryotic microsporidium N. parisii, this shared ...
Similar publications
Natural 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...
Citations
... A genome-wide association study (GWAS) revealed an intermediate-frequency indel polymorphism in the drh-1 gene (encoding a RIG-I-like helicase) as the main locus explaining variation in ORV sensitivity [19]. This RIG-I homolog triggers viral genome degradation via small RNA silencing pathways [19,20] and a host transcriptional response [21][22][23][24][25][26]. In addition to this genome-wide association approach, a biparental cross between the reference C. elegans strain N2 and the wild isolate CB4856 recently identified a quantitative trait locus on the right of chromosome IV, which may be partially explained by a non-synonymous polymorphism in the cul-6 gene, coding for a cullin, an ubiquitin ligase cofactor [22,27]. ...
... In addition to the analysis of natural variation, forward and reverse genetic studies in C. elegans revealed a number of factors required in host defense or for the viral cycle [24,28,29,30,31,32]. Especially, C. elegans antiviral immune response involves: i) the small RNA response; ii) the ubiquitin pathway [22,23]; iii) a conserved SID-3-dependent signaling pathway involved in receptor-mediated endocytosis, similar to their mammalian orthologs [28], and in the phosphorylation of STA-1, a homolog of mammalian STAT [24]; iv) the conserved role of uridylation in destabilization of the viral RNA [29]. Viral infection in C. briggsae has not been studied on the host side beyond articles describing the intestinal site of infection [12] and the similarity of the transcriptional response compared to C. elegans [23]. ...
... Especially, C. elegans antiviral immune response involves: i) the small RNA response; ii) the ubiquitin pathway [22,23]; iii) a conserved SID-3-dependent signaling pathway involved in receptor-mediated endocytosis, similar to their mammalian orthologs [28], and in the phosphorylation of STA-1, a homolog of mammalian STAT [24]; iv) the conserved role of uridylation in destabilization of the viral RNA [29]. Viral infection in C. briggsae has not been studied on the host side beyond articles describing the intestinal site of infection [12] and the similarity of the transcriptional response compared to C. elegans [23]. ...
Antagonistic relationships such as host-virus interactions potentially lead to rapid evolution and specificity in interactions. The Orsay virus is so far the only horizontal virus naturally infecting the nematode C. elegans. In contrast, several related RNA viruses infect its congener C. briggsae, including Santeuil (SANTV) and Le Blanc (LEBV) viruses. Here we focus on the host’s intraspecific variation in sensitivity to these two intestinal viruses. Many temperate-origin C. briggsae strains, including JU1264 and JU1498, are sensitive to both, while many tropical strains, such as AF16, are resistant to both. Interestingly, some C. briggsae strains exhibit a specific resistance, such as the HK104 strain, specifically resistant to LEBV. The viral sensitivity pattern matches the strains’ geographic and genomic relationships. The heavily infected strains mount a seemingly normal small RNA response that is insufficient to suppress viral infection, while the resistant strains show no small RNA response, suggesting an early block in viral entry or replication. We use a genetic approach from the host side to map genomic regions participating in viral resistance polymorphisms. Using Advanced Intercrossed Recombinant Inbred Lines (RILs) between virus-resistant AF16 and SANTV-sensitive HK104, we detect Quantitative Trait Loci (QTLs) on chromosomes IV and III. Building RILs between virus-sensitive JU1498 and LEBV-resistant HK104 followed by bulk segregant analysis, we identify a chromosome II QTL. In both cases, further introgressions of the regions confirmed the QTLs. This diversity provides an avenue for studying virus entry, replication, and exit mechanisms, as well as host-virus specificity and the host response to a specific virus infection.
... Strikingly, C. elegans activates a similar transcriptional response when its intestinal cells are infected by virus or microsporidia, and this response is distinct from that activated by extracellular pathogens remaining in the intestinal lumen Bakowski et al., 2014;Yang et al., 2016;Chen et al., 2017a). This transcriptional response has thus been named the Intracellular Pathogen Response (IPR) (Reddy et al., 2017). ...
... Of the approximately 20,000 genes in C. elegans (The C. elegans Sequencing Consortium, 1998), the IPR encompasses a core of 80 highly upregulated genes, in addition to hundreds of other genes that are upregulated to a lesser extent and in different conditions (Reddy et al., 2017). Activation of these genes has been observed as soon as 8 h post inoculation (hpi) for N. parisii and 12 hpi for OrV (Bakowski et al., 2014;Chen et al., 2017a). A large proportion of IPR genes is formed by a subset of the pals gene family, whose biochemical activity is currently unknown. ...
... The IPR is also enriched for genes encoding components of ubiquitin ligase complexes (such as the cullin CUL-6, SKP1-related), which mediate a defense response and thermotolerance (see the section on Ubiquitin-mediated responses). The core set of genes is regulated by both microsporidia and viral infections (see dataset comparisons in Chen et al., 2017a;Lažetić et al., 2022). Beyond this canonical response, further genes are upregulated depending on host genetics, pathogen inoculum, laboratory conditions, and stage of infection Bakowski et al., 2014;Chen et al., 2017a;. ...
The model organism Caenorhabditis elegans is susceptible to infection by obligate intracellular pathogens, specifically microsporidia and viruses. These intracellular pathogens infect intestinal cells, or, for some microsporidia, epidermal cells. Strikingly, intestinal cell infections by viruses or microsporidia trigger a common transcriptional response, activated in part by the ZIP-1 transcription factor. Among the strongest activated genes in this response are ubiquitin-pathway members and members of the pals family, an intriguing gene family with cross-regulations of different members of genomic clusters. Some of the induced genes participate in host defense against the pathogens, for example through ubiquitin-mediated inhibition. Other mechanisms defend the host specifically against viral infections, including antiviral RNA interference and uridylation. These various immune responses are altered by environmental factors and by intraspecific genetic variation of the host. These pathogens were first isolated 15 years ago and much remains to be discovered using C. elegans genetics; also, other intracellular pathogens of C. elegans may yet to be discovered.
... Studies of Orsay virus infection in the past few years have uncovered several conserved antiviral defense mechanisms. These include the RNA interference pathway that degrades viral RNA and triggers antiviral gene expression [1,14,15]; an inductive transcriptional response called Intracellular Pathogen Response [16]; a uridylyltransferase that destabilizes viral RNAs by 3 0 end uridylation [17]; and ubiquitin protein modifications and turnover [18,19]. In another study, bioinformatics analyses of expressed genes during Orsay virus infection revealed the presence of uncharacterized anti-stress pathways [20]. ...
C . elegans is a free-living nematode that is widely used as a small animal model for studying fundamental biological processes and disease mechanisms. Since the discovery of the Orsay virus in 2011, C . elegans also holds the promise of dissecting virus-host interaction networks and innate antiviral immunity pathways in an intact animal. Orsay virus primarily targets the worm intestine, causing enlarged intestinal lumen as well as visible changes to infected cells such as liquefaction of cytoplasm and convoluted apical border. Previous studies of Orsay virus identified that C . elegans is able to mount antiviral responses by DRH-1/RIG-I mediated RNA interference and Intracellular Pathogen Response, a uridylyltransferase that destabilizes viral RNAs by 3′ end uridylation, and ubiquitin protein modifications and turnover. To comprehensively search for novel antiviral pathways in C . elegans , we performed genome-wide RNAi screens by bacterial feeding using existing bacterial RNAi libraries covering 94% of the entire genome. Out of the 106 potential antiviral gene hits identified, we investigated those in three new pathways: collagens, actin remodelers, and epigenetic regulators. By characterizing Orsay virus infection in RNAi and mutant worms, our results indicate that collagens likely form a physical barrier in intestine cells to inhibit viral infection by preventing Orsay virus entry. Furthermore, evidence suggests that actin remodeling proteins ( unc-34 , wve-1 and wsp-1 ) and chromatin remodelers ( nurf-1 and isw-1 ) exert their antiviral activities by regulating the intestinal actin ( act-5 ), a critical component of the terminal web which likely function as another physical barrier to prevent Orsay infection.
... Differentially expressed genes from RNA-seq expression data for pals-17(*) versus wild type and pals-17(*) pals-20(*) versus wild type were ranked based on Log2 fold changes and converted into a GSEA-compatible file. 93 previously published gene sets were used for comparison [9,10,12,19,23,54,[59][60][61][62][63][64][65][66][67][68][69][70][71][72][73][74][75][76][77]. For both analyses, a signal-to-noise metric of 1,000 permutations was used with "no collapse" as gene symbols were pre-defined in the ranked gene list. ...
The immune system continually battles against pathogen-induced pressures, which often leads to the evolutionary expansion of immune gene families in a species-specific manner. For example, the pals gene family expanded to 39 members in the Caenorhabditis elegans genome, in comparison to a single mammalian pals ortholog. Our previous studies have revealed that two members of this family, pals-22 and pals-25, act as antagonistic paralogs to control the Intracellular Pathogen Response (IPR). The IPR is a protective transcriptional response, which is activated upon infection by two molecularly distinct natural intracellular pathogens of C. elegans-the Orsay virus and the fungus Nematocida parisii from the microsporidia phylum. In this study, we identify a previously uncharacterized member of the pals family, pals-17, as a newly described negative regulator of the IPR. pals-17 mutants show constitutive upregulation of IPR gene expression, increased immunity against intracellular pathogens, as well as impaired development and reproduction. We also find that two other previously uncharacterized pals genes, pals-20 and pals-16, are positive regulators of the IPR, acting downstream of pals-17. These positive regulators reverse the effects caused by the loss of pals-17 on IPR gene expression, immunity, and development. We show that the negative IPR regulator protein PALS-17 and the positive IPR regulator protein PALS-20 colocalize inside and at the apical side of intestinal epithelial cells, which are the sites of infection for IPR-inducing pathogens. In summary, our study demonstrates that several pals genes from the expanded pals gene family act as ON/OFF switch modules to regulate a balance between organismal development and immunity against natural intracellular pathogens in C. elegans.
... RNAi knockdown. Another study reported downregulation of pud-1.2 and pud-4 following viral infection of C. elegans [76]. Other overlapping genes such as hsp-17 encoding a heat shock protein, exhibited inverted regulation in either strain. ...
Parasitic nematodes pose a significant threat to human and animal health, as well as cause economic losses in the agricultural sector. The use of anthelmintic drugs, such as Ivermectin (IVM), to control these parasites has led to widespread drug resistance. Identifying genetic markers of resistance in parasitic nematodes can be challenging, but the free-living nematode Caenorhabditis elegans provides a suitable model. In this study, we aimed to analyze the transcriptomes of adult C. elegans worms of the N2 strain exposed to the anthelmintic drug Ivermectin (IVM), and compare them to those of the resistant strain DA1316 and the recently identified Abamectin Quantitative Trait Loci (QTL) on chromosome V. We exposed pools of 300 adult N2 worms to IVM (10-7 and 10-8 M) for 4 hours at 20°C, extracted total RNA and sequenced it on the Illumina NovaSeq6000 platform. Differentially expressed genes (DEGs) were determined using an in-house pipeline. The DEGs were compared to genes from a previous microarray study on IVM-resistant C. elegans and Abamectin-QTL. Our results revealed 615 DEGs (183 up-regulated and 432 down-regulated genes) from diverse gene families in the N2 C. elegans strain. Of these DEGs, 31 overlapped with genes from IVM-exposed adult worms of the DA1316 strain. We identified 19 genes, including the folate transporter (folt-2) and the transmembrane transporter (T22F3.11), which exhibited an opposite expression in N2 and the DA1316 strain and were deemed potential candidates. Additionally, we compiled a list of potential candidates for further research including T-type calcium channel (cca-1), potassium chloride cotransporter (kcc-2), as well as other genes such as glutamate-gated channel (glc-1) that mapped to the Abamectin-QTL.
... Studies of Orsay infection in the past few years have uncovered several conserved antiviral defense mechanisms. These include the RNA interference pathway that degrades viral RNA and triggers antiviral gene expression (1,13,14); an inductive transcriptional response called Intracelluar Pathogen Response (15); a uridylyltransferase that destabilizes viral RNAs by 3′ end uridylation (16); and ubiquitin protein modifications and turnover (17,18). In another study, bioinformatics analyses of expressed genes during Orsay infection revealed the presence of uncharacterized anti-stress pathways (19). ...
C. elegans is a free-living nematode that is widely used as a small animal model for studying fundamental biological processes and disease mechanisms. Since the discovery of the Orsay virus in 2011, C. elegans also holds the promise of dissecting virus-host interaction networks and innate antiviral immunity pathways in an intact animal. Orsay primarily targets the worm intestine, causing enlarged intestinal lumen as well as visible changes to infected cells such as liquefaction of cytoplasm and rearrangement of the terminal web. Previous studies of Orsay identified that C. elegans is able to mount antiviral responses by DRH-1/RIG-I mediated RNA interference and Intracellular Pathogen Response, a uridylyltransferase that destabilizes viral RNAs by 3′ end uridylation, and ubiquitin protein modifications and turnover. To comprehensively search for novel antiviral pathways in C. elegans , we performed genome-wide RNAi screens by bacterial feeding using existing bacterial RNAi libraries covering 94% of the entire genome. Out of the 106 antiviral genes identified, we investigated those in three new pathways: collagens, actin remodelers, and epigenetic regulators. By characterizing Orsay infection in RNAi and mutant worms, our results indicate that collagens likely form a physical barrier in intestine cells to inhibit viral infection by preventing Orsay entry. Furthermore, evidence suggests that the intestinal actin ( act-5 ), which is regulated by actin remodeling proteins ( unc-34 , wve-1 and wsp-1 ), a Rho GTPase ( cdc-42 ) and chromatin remodelers ( nurf-1 and isw-1 ), also provides antiviral immunity against Orsay possibly through another physical barrier presented as the terminal web.
... We chose to look at F-box, MATH/BATH, PALS, DUF684 and DUF713 domains containing genes, implicated from differentially regulated genes of N. parisii infected C. elegans in previous publications [54,85]. Lists of gene names from respective gene classes were downloaded separately from Wormbase (http://wormbase.org/) and the corresponding protein-coding sequences were extracted from Wormbase ParaSite (https://parasite.wormbase.org/). ...
Animals are under constant selective pressure from a myriad of diverse pathogens. Microsporidia are ubiquitous animal parasites, but the influence they exert on shaping animal genomes is mostly unknown. Using multiplexed competition assays, we measured the impact of four different species of microsporidia on 22 wild isolates of Caenorhabditis elegans . This resulted in the identification and confirmation of 13 strains with significantly altered population fitness profiles under infection conditions. One of these identified strains, JU1400, is sensitive to an epidermal-infecting species by lacking tolerance to infection. JU1400 is also resistant to an intestinal-infecting species and can specifically recognize and destroy this pathogen. Genetic mapping of JU1400 demonstrates that these two opposing phenotypes are caused by separate loci. Transcriptional analysis reveals the JU1400 sensitivity to epidermal microsporidia infection results in a response pattern that shares similarity to toxin-induced responses. In contrast, we do not observe JU1400 intestinal resistance being regulated at the transcriptional level. The transcriptional response to these four microsporidia species is conserved, with C . elegans strain-specific differences in potential immune genes. Together, our results show that phenotypic differences to microsporidia infection amongst C . elegans are common and that animals can evolve species-specific genetic interactions.
... Similarities of responses between different host species have also been observed. For example, the response to nodavirus infection is conserved between C. elegans and C. briggsae [38]. An intergenerational transcriptional response to Pseudomonas vranovensis was conserved between some, but not all species of Caenorhabditis [39]. ...
... The type of gene classes and domains investigated for the enrichment analyses are listed in S6 Table. Among these, we actively chose to look at F-box, MATH/BATH, PALS, DUF684, DUF713, C-type lectins, and skr domains containing genes, implicated from differentially regulated genes of N. parisii infected C. elegans in previous publications [27,38]. We also examined additional types of genes and domains enriched in our datasets using DAVID Bioinformatics Resources (2021) [56,57] for C. elegans and the WormBase simple Ggne queries tool for C. briggsae. ...
... We designed our infection experiment to compare transcriptional differences between related microsporidia and host species (Fig 1B). Unlike previous transcriptional profiling experiments of Nematocida infection, which were done as continuous infections at 25˚C [27,38], we infected the worms at 21˚C for a short period of time to synchronize the infections. We pulse-infected C. elegans and C. briggsae with either N. parisii, N. ausubeli, or a mock treatment for 2.5 hours, washed to remove spores from outside the worms, and then replated the animals for a total of 10, 20, or 28 hours of infection at 21˚C. ...
Microsporidia are obligate intracellular parasites that are known to infect most types of animals. Many species of microsporidia can infect multiple related hosts, but it is not known if microsporidia express different genes depending upon which host species is infected or if the host response to infection is specific to each microsporidia species. To address these questions, we took advantage of two species of Nematocida microsporidia, N. parisii and N. ausubeli, that infect two species of Caenorhabditis nematodes, C. elegans and C. briggsae. We performed RNA-seq at several time points for each host infected with either microsporidia species. We observed that Nematocida transcription was largely independent of its host. We also observed that the host transcriptional response was similar when infected with either microsporidia species. Finally, we analyzed if the host response to microsporidia infection was conserved across host species. We observed that although many of the genes upregulated in response to infection are not direct orthologs, the same expanded gene families are upregulated in both Caenorhabditis hosts. Together our results describe the transcriptional interactions of Nematocida infection in Caenorhabditis hosts and demonstrate that these responses are evolutionarily conserved.
... We determined the concentration of the filtrate to be 428.1 (95% CI: 173.4, 972.3) × the median tissue culture infectious dose (TCID50) per µl (electronic supplementary material, Information A) [41]. We maintained populations at 20°C until freshly starved (i.e. ...
A lack of tractable experimental systems in which to test hypotheses about the ecological and evolutionary drivers of disease spillover and emergence has limited our understanding of these processes. Here we introduce a promising system: Caenorhabditis hosts and Orsay virus, a positive-sense single-stranded RNA virus that naturally infects C. elegans. We assayed species across the Caenorhabditis tree and found Orsay virus susceptibility in 21 of 84 wild strains belonging to 14 of 44 species. Confirming patterns documented in other systems, we detected effects of host phylogeny on susceptibility. We then tested whether susceptible strains were capable of transmitting Orsay virus by transplanting exposed hosts and determining whether they transmitted infection to conspecifics during serial passage. We found no evidence of transmission in 10 strains (virus undetectable after passaging in all replicates), evidence of low-level transmission in 5 strains (virus lost between passage 1 and 5 in at least one replicate) and evidence of sustained transmission in 6 strains (including all three experimental C. elegans strains) in at least one replicate. Transmission was strongly associated with viral amplification in exposed populations. Variation in Orsay virus susceptibility and transmission among Caenorhabditis strains suggests that the system could be powerful for studying spillover and emergence.
... Because of the minor effect of OrV infection on transcriptional activity, a relaxed false discovery rate (FDR < 0.1) was used to analyze the data. As all genes discovered using this threshold were IPR genes that are previously described by others, these were probably true positive hits Chen et al., 2017). ...
... (B) Viral loads (log 2 ) as determined by RT-qPCR for the strains N2, CB4856 and JU1580 after exposure to 20, 50 or 100µL OrV/500µL infection solution (student t-test; *p < 0.05). Figure S5; Supplementary Table S7B) Chen et al., 2017). Gene expression analysis by a linear model showed that 27 genes (represented by 57 spots) were differentially expressed upon infection by OrV (FDR < 0.1) (Supplementary Figure S7A) and 18 genes (represented by 44 spots) were differentially expressed by a combination of both treatment and genotype (FDR < 0.1) (Supplementary Figure S7B). ...
... Yet, after a longer period of viral exposure on the plate, CB4856 accumulates as much virus in the population as N2 and subsequently some IPR genes were also upregulated in CB4856. Therefore, plate infection assays may evoke stronger transcriptional responses which could explain why previous studies found more differentially expressed genes than this study Chen et al., 2017). Pathogen immunity could also link to the different life stage of the nematodes, as stage-dependent immunity differences were described before (Sterken et al., 2014;Balla et al., 2015). ...
Genetic variation in host populations may lead to differential viral susceptibilities. Here, we investigate the role of natural genetic variation in the Intracellular Pathogen Response (IPR), an important antiviral pathway in the model organism Caenorhabditis elegans against Orsay virus (OrV). The IPR involves transcriptional activity of 80 genes including the pals-genes. We examine the genetic variation in the pals-family for traces of selection and explore the molecular and phenotypic effects of having distinct pals-gene alleles. Genetic analysis of 330 global C. elegans strains reveals that genetic diversity within the IPR-related pals-genes can be categorized in a few haplotypes worldwide. Importantly, two key IPR regulators, pals-22 and pals-25, are in a genomic region carrying signatures of balancing selection, suggesting that different evolutionary strategies exist in IPR regulation. We infected eleven C. elegans strains that represent three distinct pals-22 pals-25 haplotypes with Orsay virus to determine their susceptibility. For two of these strains, N2 and CB4856, the transcriptional response to infection was also measured. The results indicate that pals-22 pals-25 haplotype shapes the defense against OrV and host genetic variation can result in constitutive activation of IPR genes. Our work presents evidence for balancing genetic selection of immunity genes in C. elegans and provides a novel perspective on the functional diversity that can develop within a main antiviral response in natural host populations.
