Zdraljevic, S. et al. (2017) International Worm Meeting "Multiple interacting loci contribute to topoisomerase II poison responses in Caenorhabditis elegans."

Kim, T.W., Cook, D.E., Andersen, E.C., Brady, S.C., Kameny, L.P., Hippleheuser, S.W., Zdraljevic, S.

[International Worm Meeting, 2017]

Interacting genetic and environmental factors influence responses to anti-neoplastic treatments causing large differences in efficacy among individuals in the human population. Understanding the causes of these population-wide differences is a critical step towards the elucidation of markers that are predictive of patient responses in the clinic to personalize treatments and improve existing medications. In an attempt to disentangle this complexity, we used an unbiased linkage mapping approach in Caenorhabditis elegans to identify genetic determinants underlying response to a broadly administered class of anti-neoplastic compounds that poison the activity of topoisomerase II (TOP-2) enzymes. We identified numerous quantitative trait loci (QTL) that explain phenotypic variation among a panel of recombinant inbred advanced intercross lines generated between the Bristol and Hawaiian strains in response to amsacrine, dactinomycin, and etoposide. Interestingly, multiple overlapping QTL located on the center of chromosome V explain phenotypic variation in response to these three poisons, suggesting a common mechanism underlying the responses to these compounds. We constructed reciprocal Bristol and Hawaiian near-isogenic lines (NILs) that encompass all six chrV QTL. Interestingly, these NILs only recapitulated the etoposide and dactinomycin QTL effects, but failed to recapitulate the amsacrine QTL effects. We used these NILs to generate 80 recombinant NILs with smaller introgressed regions to separate a possible amsacrine-specific multi-locus interaction within the NIL region and to narrow the QTL confidence intervals associated with responses to all three drugs. Phenotypic analysis of these sub-interval NILs isolated the two etoposide and dactinomycin-specific QTL within the original NIL region and uncoupled an amsacrine-specific incompatibility locus from an amsacrine-specific QTL. These results highlight the complexity of genetic factors that influence metazoan responses to anti-neoplastic compounds.

Hyojin Lee et al. (2008) East Asia Worm Meeting "A C. elegans AMP-activated Protein Kinase Is Phosphorylated by an LKB1 Homologue and Influences the Sensitivity to Oxidative Stress and Motility of Worms."

Jeong Soo Cho, Hyeon-Sook Koo, Anton Gartner, Nils Lambacher, Hyojin Lee, Jieun Lee

[East Asia Worm Meeting, 2008]

AAK-2 is one of two alpha isoforms of the AMP-activated protein kinase in C. elegans, and is involved in life span maintenance and stress responses. In this study, we found that AAK-2 was phosphorylated in response to paraquat treatment and the phosphorylation was attenuated by mutants of PAR-4, an LKB1 homologue. Both aak-2 and par-4 mutations increased the sensitivity of C. elegans worms to paraquat, and the double deficiency did not further increase sensitivity, indicating that they affect the same pathway. Both mutations also slowed body bending during locomotion and resulted in abnormalities of head oscillation. Consistent with this abnormal motility, expression of AAK-2::GFP fusion protein was observed in the ventral cord and body wall muscle, as well as in other neurons, pharynx, vulva, and excretory cells. Our study suggests that AMPK can influence the behavior of C. elegans worms in addition to its well-known function in metabolic control.

Bernstein, Max et al. (2013) International Worm Meeting "The hunt for quantitative trait nucleotides: a near-isogenic line based approach in C. elegans."

Rockman, Matthew, Bernstein, Max

[International Worm Meeting, 2013]

The individual nucleotides responsible for phenotypic diversity have been very difficult to isolate. The majority of known causal variants that affect traits in wild populations of a species occur within protein-coding regions. These variants are typically of very large effect and may not be typical of the sorts of alleles that drive evolution. There are many confounding effects that can explain the relative lack of causal variants, or quantitative trait nucleotides (QTNs), particularly alleles of small effect, epistasis, and gene-environment interactions. One way to elucidate quantitative trait loci (QTLs) is through the creation of near-isogenic lines (NILs), where the majority of the genome (>95%) is of one genomic background. NILs have previously been created for the nematode C. elegans and identified QTLs across the genome for several phenotypes. C. elegans is a naturally inbred species, which makes it ideal for isolating QTNs. Starting with a NIL that harbors a ~1.5 Mb segment on Chromosome X originating from the Hawaiian isolate CB4856, we have created a series of more than 1000 sub-NILs that break up this segment into smaller regions by recombination. These sub-NILs will be genotyped at 278 SNPs within the 1.5 Mb segment. We are currently piloting high-throughput fitness assays similar to previous work by Elvin et al (2011) and Ramani et al (2012). Additionally, we plan to do gene expression analysis on a subset of the sub-NIL panel to characterize expression QTNs (eQTNs). QTL mapping on simulated sub-NIL datasets shows that for a single QTL model, we can accurately map small effect (h2<0.1) causal variants. Due to the linkage disequilibrium pattern in the sub-NIL panel, traditional QTL mapping methods are not applicable. We show that pairwise comparisons among groups of similar sub-NIL genotypes provide a systematic method for identifying QTLs. We have also modeled more complex QTL scenarios, including multiple QTLs and epistatic interactions. These simulations have informed us of our potential statistical power for future QTL mapping experiments.

Montgomery, Jordan et al. (2015) International Worm Meeting "Genetic mapping of hybrid developmental delay with novel near-isogenic lines of Caenorhabditis briggsae."

Montgomery, Jordan, Ross, Joseph

[International Worm Meeting, 2015]

When two organisms are able to reproduce, but their hybrid offspring exhibit reduced fitness, this is often evidence of the onset of speciation. Such hybrid incompatibility is a result of genetic differences between the parental populations. C. briggsae is a useful system for studying the onset of speciation because of its unique and distinct phylogeographic population structure. C. briggsae is also well suited for speciation studies because of empirical evidence suggesting that genetic differences between populations result in reduced hybrid fitness. For example, previous studies have shown that approximately 20% of F2 offspring from an AF16-HK104 cross exhibit a developmental delay phenotype. This phenotype is associated with homozygosity of AF16 alleles near the middle of chromosome III. Identifying the genetic basis of this hybrid dysfunction phenotype will aid in the identification of the genetic basis of a putative speciation event. To achieve this goal, we created a panel of replicate near-isogenic lines (NILs), with AF16 alleles on chromosome III introgressed onto the HK104 background.By comparing genotypes of the NILs able to produce developmentally delayed offspring with those that cannot, we propose that the locus on chromosome III partly responsible for the epistatic interaction producing the delay phenotype exists between 4.271 and 5.649 Mbp. Future studies will focus on high-resolution genotyping of the NILs, followed by fine-mapping and candidate gene approaches to identifying the molecular basis of this trait. Ultimately, these results assist in better understanding the evolutionary forces and genetic mechanisms driving speciation events.

S.C. Harvey et al. (2006) European Worm Meeting "The Genetics of Natural Variation in the Phenotypic Plasticity of Dauer Larvae Development"

A. Shorto, S.C. Harvey, M.E. Viney

[European Worm Meeting, 2006]

S.C. Harvey, A. Shorto and M.E. Viney . Natural isolates (including N2 and DR1350) of the free-living nematode Caenorhabditis elegans vary in their phenotypic plasticity of dauer larvae development. For example, some lines appear to be highly sensitive to dauer inducing conditions while others are less so. We have sought to investigate the causes and consequences of this plasticity.. To determine the genetic basis of this plasticity of dauer development we have undertaken a quantitative trait loci (QTL) mapping based analysis of recombinant inbred lines (RILs) produced from crosses between N2 x DR1350. This has identified several regions containing candidate QTLs that affect the plasticity of dauer development. Nearly isogenic lines (NILs) have been constructed for a candidate QTL on chromosome II, with analysis of these NILs confirming that the region contains genes that affect the plasticity of dauer larvae development. This nearly isogenic region contains a maximum of 441 genes, none of which have previously been identified as being part of the genetic pathway regulating dauer development. Overall, these data have developed a clearer picture of the genetics underlying natural variation in a complex trait and demonstrate that the analysis of natural variation can reveal genes not identified by other genetic approaches.

Gaertner B E et al. (2010) Neuronal Development, Synaptic Function and Behavior, Madison, WI "The Role of Natural Genetic Variation in Shaping Thermosensory Behavior"

Parmenter M, Gaertner B E, Lockery S, KRUGLYAK L, Phillips P, Noll L, Rockman M

[Neuronal Development, Synaptic Function and Behavior, Madison, WI, 2010]

Genetic screens and ablation studies have been hugely informative in studying the genetic and neural basis to sensory behavior. However, these types of approaches are insufficient concerning questions about the role of natural genetic variation in shaping behavioral networks. Additionally, mutations in genes of small effect or epistatic effects could be missed using traditional approaches. A quantitative genetics approach compliments traditional methods by explicitly analyzing natural genetic variation and permitting statistical analysis of multiple loci, thus identifying genes that are undetectable in traditional screens. Using a dual quantitative genetics / systems biology approach, we have identified five novel loci contributing to natural variation in thermal preference as assayed on a stable linear gradient. Three of these loci have an allele-dependent epistatic relationship that appears to be rooted in differing "motivation" to move. Another two loci interact with each other seemingly to disrupt thermosensation, as strains that are heterogeneous between these two loci do not avoid physiologically damaging temperatures. The nature of these interactions are being tested directly using nearly isogenic lines (NILs) and behavioral analysis of individuals as they klinotax in the controlled environment of a microfluidic chip. Preliminary results indicate that between-strain individual differences in klinotaxis in this chip are consistent with between-strain differences in thermal preferences at the population level. Additionally, we are examining the roles that neural activity and klinokinesis play in varying thermosensory behavior among natural isolates and using NILs to identify the precise genetic variants that cause differences in these navigation strategies.

Alkan , Cigdem et al. (2017) International Worm Meeting "Interactions between Caenorhabditis nematodes and viruses infecting them naturally."

Ruaud, Albane, Alkan , Cigdem, BRESARD, Gautier, Felix, Marie-Anne

[International Worm Meeting, 2017]

The discovery of RNA viruses that naturally infect C. elegans and C. briggsae serves as an ideal model system to study antiviral immunity and host-pathogen co-evolution. The Orsay virus only infects C. elegans whereas Santeuil and Le Blanc viruses only infect C. briggsae. Intraspecifically, within both species we found a wide variation in viral sensitivity, as well as a positive correlation among wild isolates in sensitivity to both viruses in C. briggsae. An exception to this correlation is the C. briggsae strain HK104, which is specifically resistant to Le Blanc virus but sensitive to Santeuil virus. Taking advantage of this natural variation in the host, we use a genetic approach from the host side and use Recombinant Inbred Lines (RILs) to first map the recombinant genomic regions participating to the resistance/sensitivity in a general and/or specific manner. The RILs were phenotyped for the sensitivity to the relevant viruses using Fluorescent In Situ Hybridization (FISH). The genotype (SNP markers from pool sequencing) and phenotype (resistance/sensitivity from FISH) data were used to perform QTL analysis. Several Near Isogenic Lines (NILs) were created by introgressing the candidate regions. C. briggsae AF16 is resistant to both Santeuil and Le Blanc viruses while C. briggsae HK104 is specifically sensitive to the Santeuil virus. Using AF16xHK104 Advanced Intercrossed RILs (AIRILs) (Ross et al. 2011), two QTLs were detected on chromosomes III and IV for Santeuil virus sensitivity. The NILs in the AF16 background confirm both candidate regions. C. briggsae JU1498 is sensitive to both Santeuil and Le Blanc viruses. Using JU1498xHK104 RILs, a QTL on chromosome II was detected and is being introgressed. Once candidate polymorphisms associated with the virus sensitivity/resistance are identified, we will test them by RNAi knockdown, transformation rescue and/or CRISPR-mediated gene replacement.

Kleemann, Gunnar et al. (2013) International Worm Meeting "Natural genetic polymorphisms in set-15 and a casein kinase 1a homolog shape the mortality schedule of worms."

Cox, Edward, Murphy, Coleen, Kuhlman, Thomas, Kleemann, Gunnar, Bloom, Joshua, Yang, Alina, Kruglyak, Leonid, Cherng, Hua-Jay

[International Worm Meeting, 2013]

Large-scale loss-of-function C. elegans screens have demonstrated the profound flexibility of the aging schedule and identified major, conserved longevity pathways. A new challenge is to understand how longevity pathways work in the context of normal physiology, which will give us insight into the natural function of pathways controlling aging schedules and the genetic basis of naturally occurring disease states, and should lead to mechanistic insights that will allow us better control over disease and aging processes. In order to identify stable genetic polymorphisms in longevity pathways, we performed Quantitative Trait Loci (QTL) analysis using our rapid lifespan profiling platform (Chronos) to estimate lifespans across a panel of Bristol (N2) X Hawaiian (CB4856) Recombinant Inbred Lines (RILs) (Rockman et al, 2009). We identified a region on the left arm of Chromosome IV associated with variation in longevity. This region was further refined using a panel of Near Isogenic Lines (NILs) that contain short genomic fragments from the Hawaiian strain on the genetic background of the Bristol strain. Finally, using transgenic lines that rescue the longevity defect in our short-lived NILs, we identified two polymorphic genes, set-15 and a close homolog to human casein kinase 1a, which shape the mortality schedule of C. elegans. Since set-15 is a histone methyl transferase homolog, the polymorphism in set-15 may influence longevity by regulating epigenetic modification of the genome. This is particularly interesting since the polymorphisms in the longevity-linked region have been linked to gene expression differences (expressionQTLs; eQTLs) across the genome. The regulation of these eQTLs provides a potential mechanism by which natural allelic variation can lead to large changes in gene expression and subsequently, longevity. Such natural variation likely contributes to differences in longevity observed in human populations, as well.

Line Elnif Thomsen et al. (2006) European Worm Meeting "Caenorhabditis elegans is a model host for Listeria monocytogenes"

Bente M. Jensen, Line Elnif Thomsen, Birgitte H. Kallipolitis, Nils J. Fargeman, Hanne Ingmer

[European Worm Meeting, 2006]

Line Elnif Thomsen1, Bente M. Jensen2, Nils J. Frgeman2, Birgitte H. Kallipolitis2 and Hanne Ingmer1. Several bacterial pathogens, both Gram-positive and Gram-negative, kill the nematode Caenorhabditis elegans when supplied as a food source, and a variety of bacterial virulence factors, known to play a role in mouse-models, have been shown also to play a role in nematode pathogenesis. The gram-positive facultative intracellular food-borne pathogen Listeria monocytogenes is associated with serious invasive infections in humans and animals. We have investigated the possibility of using C. elegans as a model to analyse the virulence of various L. monocytogenes strains. We found that L. monocytogenes kill C. elegans over the course of several days, as a consequence of an accumulation of bacteria in the worm intestine, and that there is an overlap between L. monocytogenes virulence factors required for mammalian and nematode pathogenesis. C. elegans is therefore an attractive model to study the virulence of this pathogen and potentially to identify new virulence factors and further work might reveal the genetic repertoire required for infection of the nematode by L. monocytogenes.

Bernstein MR et al. (2019) Evol Lett "Tightly linked antagonistic-effect loci underlie polygenic phenotypic variation in C. elegans."

Andersen EC, Rockman MV, Zdraljevic S, Bernstein MR

[Evol Lett, 2019]

Recent work has provided strong empirical support for the classic polygenic model for trait variation. Population-based findings suggest that most regions of genome harbor variation affecting most traits. Here, we use the approach of experimental genetics to show that, indeed, most genomic regions carry variants with detectable effects on growth and reproduction in <i>Caenorhabditis elegans</i> populations sensitized by nickel stress. Nine of 15 adjacent intervals on the X chromosome, each encompassing 0.001 of the genome, have significant effects when tested individually in near-isogenic lines (NILs). These intervals have effects that are similar in magnitude to those of genome-wide significant loci that we mapped in a panel of recombinant inbred advanced intercross lines (RIAILs). If NIL-like effects were randomly distributed across the genome, the RIAILs would exhibit phenotypic variance that far exceeds the observed variance. However, the NIL intervals are arranged in a pattern that significantly reduces phenotypic variance relative to a random arrangement; adjacent intervals antagonize one another, cancelling each other's effects. Contrary to the expectation of small additive effects, our findings point to large-effect variants whose effects are masked by epistasis or linkage disequilibrium between alleles of opposing effect.