Crombie, T.A. et al. (2019) International Worm Meeting "Massive sampling of Caenorhabditis elegans across the Hawaiian Islands reveals remarkable genetic diversity on the islands and admixture with European populations."

Andersen, E.C., Evans, K.S., Zdraljevic, S., Cook, D.E., Hahnel, S.R., Rodriguez, B.C., Zhang, G., Crombie, T.A., Tanny, R.E., van der Zwaag, J., Brady, S.C., Lee, D., Wang, Y.

[International Worm Meeting, 2019]

Global surveys of Caenorhabditis elegans genetic diversity suggest that large-scale selective sweeps on chromosomes I, IV, V, and X are associated with reduced diversity in some regions of the world. These sweeps are thought to be caused by adaptation to human-associated environments, so swept strains might not represent the natural diversity present before human influence. Interestingly, strains isolated from the Hawaiian Islands are generally highly diverged from the rest of the global population and contain little evidence of these sweeps. Therefore, we sought to better characterize the natural diversity of the species by collecting C. elegans from the Hawaiian islands. At two different times, we used a collection protocol to sample nematodes from a total of 3,264 sites across five Hawaiian islands. Among the 4,558 nematodes we isolated, we identified five Caenorhabditis species including C. elegans, C. briggsae, C. tropicalis, C. kamaaina, and a new species, C. oiwi. On the first collection trip, we discovered that C. elegans was generally found in cooler environments at higher elevations than other members of the genus. These data enabled improvements to our collection protocols that increased our success rate for isolating C. elegans from 1.8% to 4.1% on the second collection trip. We identified 26 distinct C. elegans strains from our first collection trip, increasing the total number of Hawaiian strains to 43. The mean genome-wide nucleotide diversity of these 43 Hawaiian strains is three-fold higher than the 233 non-Hawaiian strains from around the globe. We used ADMIXTURE to investigate the ancestry of all 276 strains and identified six distinct ancestral populations. Surprisingly, we saw evidence of admixture between some of the Hawaiian strains and one predominantly swept European population. Taken together, these findings confirm that divergent strains can be collected reliably from the Hawaiian Islands but raise new questions about the direction, magnitude, and timing of gene flow among swept and non-swept populations that has contributed to the global pattern of diversity in the species.

Zhang, Gaotian et al. (2021) International Worm Meeting "The genetic architectures of gene expression variation in wild C. elegans"

Hahnel, Steffen, Andersen, Erik, Roberto, Nicole, Zhang, Gaotian, Lee, Daehan

[International Worm Meeting, 2021]

Natural variation in gene expression is a major source of phenotypic diversity among individuals. Genetic variants underlying gene expression differences are known as expression quantitative trait loci (eQTL). In Caenorhabditis elegans, both local eQTL (located close to the genes they influence) and distant eQTL (located farther away from the genes they influence) have been identified using linkage analysis from a panel of recombinant inbred advanced intercross lines (RIAILs) between the laboratory reference strain, N2, and a wild strain, CB4856. However, the eQTL detected using RIAILs were limited to genetic variants between N2 and CB4856. Here, we investigated the natural variation in gene expression of 205 genetically distinct C. elegans wild strains by performing RNA-sequencing on synchronized young adult hermaphrodites. We obtained reliable expression of 25,896 protein-coding transcripts (16,106 genes). We used genome-wide association (GWA) analysis to identify 3,342 local eQTL for 3,342 transcripts (2,777 genes), and 2,835 distant eQTL for 2,206 transcripts (2,082 genes). We found that most of the narrow-sense heritability for transcript expression variation is explained by detected eQTL. Of the 2,835 distant eQTL, 1,670 eQTL significantly clustered in 54 hotspots across the C. elegans genome. We will further explore causal genes underlying these distant eQTL hotspots and their functions. Additionally, we applied mediation analysis to the eQTL data and other organism-level quantitative traits to elucidate the genetic effects on phenotypic variation mediated by gene expression. For instance, instead of relying on time-consuming investigations of rare genetic variants missed in GWA studies and prior knowledge of the trait as published in our study of C. elegans responses to benzimidazoles (Hahnel et al. 2018), the significant mediating effect of the expression of ben-1, the causal gene, was quickly and successfully identified by mediation analysis. Our results suggest that mediation analysis using expression data facilitates identification of causal genes in GWA studies in C. elegans.

TM Morse et al. (1999) International C. elegans Meeting "Time-series analysis of pirouettes in C. elegans chemotaxis."

TM Morse, JT Pierce, SR Lockery, ML Moravec, TC Ferree, SE Owens, AB Bates

[International C. elegans Meeting, 1999]

Chemotaxis in C. elegans involves a series of abrupt turns (pirouettes) triggered by movement down a gradient of chemical attractant * . Analysis of the time series of concentration change experienced by a worm, together with its pirouette record, suggests a three-stage chemotaxis mechanism in which chemical concentration (C(t)) is differentiated (dC(t)/dt), smoothed (q(t)), and converted into pirouette probability by a nonlinear function (P(q[t])). To test the plausibility of this mechanism, we constructed a computer model in which the smoothing filter and the nonlinearity were estimated from the time series of dC(t)/dt and the pirouette records of real worms. Pirouettes were modeled by sampling randomly, via a Poisson process with probability P(q[t]), from the distribution of direction changes associated with pirouettes in real worms. The average chemotaxis index (time-average of normalized C(t)) of model worms (0.47 +/- 0.05 SD, n = 2000) closely matched the average chemotaxis index of real worms (0.45 +/- 0.04 SD, n = 45), indicating that the three-stage mechanism is quantitatively sufficient to account for C. elegans chemotaxis. To determine the form of the smoothing filter and nonlinearity directly, we have devised a new behavioral assay that subjects unrestrained worms to negative-going impulses in dC/dt as they swim across a sharp border between high and low concentrations of attractant. Preliminary results show that immediately after a border crossing, large impulses make P(t)~= 1, while small impulses make 0 t) >< C. elegans . * Pierce, J.T., and Lockery, S.R., J. Neurosci. (Submitted)

Sreekanth Chalasani et al. (2007) International Worm Meeting "Functional analysis of a neural circuit controlling turning behavior in Caenorhabditis elegans."

Cornelia Bargmann, Jesse Gray, Makoto Tsunozaki, Sreekanth Chalasani, Nikolas Chronis

[International Worm Meeting, 2007]

C. elegans increase its frequency of reversals and turns (jointly termed pirouettes, Pierce-Shimomura et al 1999) after removal of a food stimulus. The AWC and ASK sensory neurons and the AIB interneurons stimulate pirouettes immediately after removal from food, while the AIY and AIA interneurons inhibit pirouettes (Wakabayashi et al 2004, Gray et al 2005). We have found that the sensory neuron AWC releases two neurotransmitters (glutamate and a neuropeptide, NLP-1) when the worm is removed from food. The released glutamate acts to activate AIB and inhibit AIY, promoting reversals. Strains with different reversal frequencies can be generated by manipulating the level of glutamate receptors on interneurons AIB and AIY. Decreasing receptor expression leads to fewer reversals, and increasing receptor expression results in more reversals than in wild-type. The AWC released neuropeptide NLP-1 serves to reduce reversals, suggesting that reversal frequencies are regulated by at least two opposing signaling systems. Consistent with behavioral responses, AWC and AIB respond (by increasing calcium concentration) to removal of stimulus. We plan to extend the imaging studies to other neurons in the circuit. These results provide a plausible molecular explanation that links neurotransmitters, their receptors, and neuronal circuitry to generate behavior. References: Gray, J.M., Hill, J.J., and Bargmann, C.I. (2005). A circuit for navigation in Caenorhabditis elegans. Proc. Natl. Acad. Sci. 102, 3184-3191. Pierce-Shimomura, J.T., Morse, T.M., and Lockery, S.R. (1999). The fundamental role of pirouettes in Caenorhabditis elegans chemotaxis. J. Neurosci 19, 9557-9569. Wakabayashi, T., Kitagawa, I., and Shingai, R. (2004). Neurons regulating the duration of forward locomotion in Caenorhabditis elegans. Neurosci. Res. 50, 103-111.

Sreekanth H Chalasani et al. (2005) International Worm Meeting "Molecular analysis of a neural circuit for navigation in Caenorhabditis elegans."

Sreekanth H Chalasani, Nikolas Chronis, Jesse M Gray, Cornelia I Bargmann

[International Worm Meeting, 2005]

Navigation in C.elegans is achieved by sustained forward movement that is interrupted with reversals and turns (jointly termed pirouettes, Pierce-Shimomura et al 1999). We are interested in the neural circuit that controls the frequency of reversals and turns during exploratory behavior. After worms are taken off bacterial food, they exhibit an initial local search with a high frequency of pirouettes. The AWC and ASK sensory neurons and the AIB interneurons stimulate pirouettes immediately after removal from food, while the AIY interneurons inhibit pirouettes. (Tsalik and Hobert 2003, Wakabayashi et al 2004, Gray et al 2005). How is activity transmitted through this neuronal circuit? The neurotransmitters glutamate and dopamine regulate turning frequency (Hills et al 2004). We found that the vesicular glutamate transporter EAT-4 is essential for the generation of pirouettes after removal from food. Using cell-specific rescue of eat-4 mutants, we show that both AWC and ASK sensory neurons can release glutamate to stimulate pirouettes. The released glutamate appears to be sensed by a glutamate-gated chloride channel (GLC-3) that inhibits the AIY interneurons, and the glutamate-gated cation channel GLR-1, which stimulates the AIB interneurons. These results provide a plausible molecular explanation that links neurotransmitters, their receptors, and neuronal circuitry to generate behavior. We are currently attempting to image neuronal activity in these neurons using genetically encoded calcium sensors. References: Gray, J.M., Hill, J.J., and Bargmann, C.I. (2005). A circuit for navigation in Caenorhabditis elegans. Proc. Natl. Acad. Sci. 102, 3184-3191. Hills, T., Brockie, P.J., and Maricq, A.V. (2004). Dopamine and glutamate control area-restricted search behavior in Caenorhabditis elegans. J. Neurosci 24, 1217-1225. Pierce-Shimomura, T., Morse, T.M., and Lockery, S.R. (1999). The fundamental role of pirouettes in Caenorhabditis elegans chemotaxis. J. Neurosci 19, 9557-9569. Wakabayashi, T., Kitagawa, I., and Shingai, R. (2004). Neurons regulating the duration of forward locomotion in Caenorhabditis elegans. Neurosci. Res. 50, 103-111.

Stephen R Wicks et al. (2001) International C. elegans Meeting "A snip-SNP map of the C. elegans genome: Applications to positional cloning."

Robert H Waterston, Ronald H A Plasterk, Stephen R Wicks, Warren R Gish, Raymond T Yeh

[International C. elegans Meeting, 2001]

Single nucleotide polymorphisms (SNPs) are valuable genetic markers of human disease. They also comprise the highest potential density marker set available for mapping experimentally derived mutations in model organisms such as C. elegans . To facilitate the positional cloning of mutations we have identified polymorphisms in CB4856, an isolate from a Hawaiian isle that shows a uniformly high density of polymorphisms compared to the reference Bristol N2 strain. Based on 5.4 Mbp of aligned sequences, we predict 6222 polymorphisms. Furthermore, 3457 of these markers modify restriction enzyme recognition sites (snip-SNPs) and are therefore easily detected as RFLPs. Of these, we have experimentally confirmed 493 by restriction digest to produce a snip-SNP map of the worm genome (ref 1). A mapping strategy using snip-SNPs and bulked segregant analysis (BSA, ref 2) is outlined. CB4856 is crossed into a mutant strain, and exclusion of CB4856 alleles of a subset of snip-SNPs in mutant progeny is assessed with BSA. The proximity of a linked marker to the mutation is estimated by the relative proportion of each form of the biallelic marker in populations of wildtype and mutant genomes. This step bounds the mutation between flanking snip-SNPs. These flanking markers can be used to detect recombination in individual animals, and only recombinant strains need be phenotyped. By mapping the recombination points in individual animals, it is possible to rapidly zoom in on the site of a mutation. The advantages and limitations of this approach will be discussed. References: 1) Wicks, S.R., Yeh, R.T., Gish, W.R., Waterston, R.H. & Plasterk, R.H.A. (in press) Rapid gene mapping in C. elegans using a high density polymorphism map. Nature Genetics . 2) Michelmore, R.W., Paran, I. & Kesseli, R.V. Identification of markers linked to disease-resistance genes by bulked segregant analysis: a rapid method to detect markers in specific genomic regions by using segregating populations. Proc. Natl. Acad. Sci. USA 88, 9828-9832. (1991).

Yuichi Iino (2007) International Worm Meeting "Two different mechanisms contribute to salt chemotaxis in C. elegans."

Yuichi Iino

[International Worm Meeting, 2007]

"Pirouette mechanism" has been proposed as the main strategy for chemotaxis to water-soluble chemoattractants in C. elegans. Pirouette refers to a bout of sharp turns by which worms quickly change the direction of locomotion. Pirouette occurs in a stochastic manner, but its probability is modulated by the temporal change of chemoattractant concentration. When the concentration of chemoattaractant sensed by a worm declines, it shows a higher probability of pirouette, eventually biasing the worm movement towards the source of chemoattractant (1). Although this mechanism alone can generate chemotaxis, computer simulations based on the observed values show that model worms underperform real worms, suggesting possible existence of another mechanism (1). Therefore, video tracking analysis of chemotaxis was conducted anew using NaCl as a chemoattractant, which lead to identification of a navigation mechanism different from the pirouette mechanism. During runs, worms do not go straight but tend to turn slowly to either direction, apparently in a random fashion. However, the tracking analysis shows that turning is not random but biased by the spatial gradient of chemoattractant concentration. Specifically, the average turning rate is proportional to the chemoattaractant gradient in the direction perpendicular to the direction of locomotion. This mechanism is called the weather vane mechanism; by this mechanism, worms gradually change the direction of locomotion towards the gradient peak. By computer simulations, neither pirouette mechanism alone nor weather vane mechanism alone generates as efficient chemotaxis as real worms, while combination of the two generates a strong chemotaxis, suggesting contribution of both mechanisms to real chemotaxis. These results show that worms detect both temporal and spatial changes of chemical stimuli, and by utilizing both information maximize the chance to reach the source of chemicals. Mutational or physical removal of ASE sensory neurons totally eliminates both mechanisms, suggesting that the same salt-sensing neurons mediate both mechanisms. Further analysis of underlying neural circuits is under way. (1) Pierce-Shimomura, J.T., Morse, T.M. & Lockery, S.R. The fundamental role of pirouettes in Caenorhabditis elegans chemotaxis. J Neurosci 19, 9557-69 (1999).

Olahova M et al. (2010) Aging, Metabolism, Stress, Pathogenesis, and Small RNAs, Madison, WI "Modulation of DAF-16 Activity by a Peroxiredoxin Antioxidant Enzyme with Tissue-Specific Effects on Stress Resistance and Ageing"

Crook H M, Veal E A, Olahova M

[Aging, Metabolism, Stress, Pathogenesis, and Small RNAs, Madison, WI, 2010]

Over the past few years several studies have suggested that links between oxidative stress and ageing may be more complex than initially proposed by oxidative damage theories. For instance, although many long-lived mutant C. elegans have increased stress resistance there is limited evidence that increased antioxidant defences are responsible for their increased lifespan. 2-Cys Peroxiredoxins (Prx) are abundant, conserved thioredoxin peroxidases, which have been shown to have important roles in responses to hydrogen peroxide and in normal longevity. For instance, C. elegans lacking PRDX-2, the ortholog of the Prdx1 tumor suppressor, are highly sensitive to hydrogen peroxide and short-lived. Consistent with an antioxidant role, intestinal PRDX-2 protects against oxidative stress. However, unexpectedly, non-intestinal PRDX-2 acts to reduce the resistance of C. elegans to some forms of oxidative stress, such as arsenite[1]. Here we will present data suggesting that the increased arsenite resistance associated with loss of PRDX-2 from non-intestinal tissues is dependent on the FOXO transcription factor DAF-16. Indeed, we find that DAF-16 is partially activated in PRDX-2-deficient animals. Our preliminary data indicate that reduced insulin/IGF-1-like signaling can increase still further the arsenite resistance of PRDX-2-deficient animals, but only partially rescue the progeric phenotypes associated with PRDX-2-deficiency. Similarly, although intestinal expression of PRDX-2 increases the oxidative stress resistance of prdx-2 mutants beyond that of wild-type animals, it does not increase their lifespan [1]. Together these data suggest that the accelerated ageing associated with loss of PRDX-2 from non-intestinal tissues is not a consequence of reduced resistance to oxidative stress. Moreover, these data demonstrate the importance of taking into consideration tissue-specific activities when assessing how stress-defences may influence the stress resistance and ageing of the whole animal. [1] Olahova, M., Taylor, S.R., Khazaipoul, S., Wang, J., Morgan, B.A., Matsumoto, K., Blackwell, T.K., and Veal, E.A. (2008). A redox-sensitive peroxiredoxin that is important for longevity has tissue- and stress-specific roles in stress resistance. PNAS, 105, 19839-19844.

William C. Chen et al. (2006) European Worm Meeting "Using C. elegans Chromosome Substitution Strains to Map Innate Immunity Loci"

William C. Chen, Sandra S. Slutz, Man-Wah Tan

[European Worm Meeting, 2006]

William C. Chen1, Sandra S. Slutz1, and Man-Wah Tan. The genome of the Hawaiian natural isolate (CB4856) of Caenorhabditis elegans contains thousands of SNPs that differ from Bristol N2. Wicks et al. proposed a method for mapping mutations generated in the N2 background by genotyping snip-SNPs in crosses between mutants and the Hawaiian strain [1]. However, some polymorphisms lead to functional differences between the two strains that affect a variety of phenotypes such as social feeding, germline RNAi, and immunity [2-4]. We found that the Hawaiian strain has increased susceptibility to Pseudomonas aeruginosa. In order to identify the loci responsible for the Hawaiian sensitivity, recombinant inbred lines between Hawaiian and N2 were created. Analysis of these lines shows that changes at multiple loci cause the observed Hawaiian pathogen sensitivity phenotype. Separately, we have EMS mutagenized N2 worms to obtain mutants with enhanced susceptibility to pathogen (Esp); however, we have been unable to map these mutants using Hawaiian due to Hawaiians sensitivity to pathogen. In order to map our Esp mutants and to identify the Hawaiian immunity loci, we have created N2/Hawaiian chromosome substitution strains (CSSs). Each of these six strains (five autosomes and the X chromosome) contains a single homozygous Hawaiian chromosome substituted into an N2 background. CSSs have been used in mice to map quantitative trait loci, and we reasoned that a similar strategy could identify loci that contribute to the Hawaiian phenotype [5]. Importantly, those CSSs that are like N2 for sensitivity to pathogen can be used to map Esp mutations that occur on the same chromosome.. We will report on using the CSSs to identify Hawaiian specific innate immunity loci and the mapping of Esp mutants from our mutagenesis screen. We believe that these CSSs will be a valuable resource for other researchers who have been unable to use the Hawaiian strain for mapping their own mutants due to the differences in phenotypes between Hawaiian and N2. 1. Wicks, S.R., et al., Nature Genetics, 2001. 28(2): p. 160-164.. 2. de Bono, M. and C.I. Bargmann, Cell, 1998. 94(5): p. 679-689; 3. Schulenburg, H. and S. Muller. Parasitology, 2004. 128(4): p. 433-443; 4. Tijsterman, M., et al., Current Biology, 2002. 12(17): p. 1535-1540; 5. Singer, J.B., et al., Science, 2004. 304(5669): p. 445-448.

H A Zariwala et al. (2001) International C. elegans Meeting "The trampoline assay: A new method for measuring the step response of the thermotaxis mechanism in C. elegans."

H A Zariwala, S R Lockery, S Faumont

[International C. elegans Meeting, 2001]

Worms in a thermal gradient migrate to the temperature at which they were cultivated (thermotaxis). Qualitative analysis of the effects of mutations and neuronal ablations on thermosensory behavior[1,2] suggests that thermotaxis may involve a mixture of two strategies: isothermal tracking, in which the head is aligned orthogonal to the thermal gradient, and pirouetting, in which course correction is achieved by a cluster of sharp turns, as in chemotaxis[3]. As a direct test of whether pirouettes play a role in thermotaxis, we devised a new procedure--the "trampoline assay"--in which worms were placed on a thin (~100 m m) agarose film suspended over a chamber filled with buffer at the cultivation temperature (21 o C). After a 5 min. adaptation period, we quickly replaced the first chamber with a second containing buffer at a lower temperature (18 o C), and observed the worm for an additional 5 min. Temperature measurements on the surface of the film indicated that the time constant of the temperature shift was ~4 sec. When we shifted wild type worms (n = 14) to the lower temperature (down-shifts), we observed a transient increase in the probability of initiating sharp turns (time to peak = ~30 sec; decay time constant = ~50 sec). This result is consistent with a role for pirouettes in thermotaxis because, in a spatial temperature gradient, a pirouette induced by a temperature drop tends to return the worm to its cultivation temperature. The increase in sharp turn probability is almost certainly not a mechanical artifact because we saw no increase in turning when the second chamber was at the cultivation temperature (n = 8). Conversely, when we down-shifted the cryophilic mutant ttx-3 (ks5) (n = 15), we observed a sustained decrease in sharp-turn probability. In a spatial temperature gradient, a decrease in turning induced by a temperature drop tends to move the worm toward lower temperatures, consistent with the cryophilic phenotype of ttx-3 . We conclude that pirouettes are likely to be important for thermotaxis, at least for excursions below the cultivation temperature. Experiments are in progress to determine if the same is true for excursions above cultivation temperature. 1. Hedgecock, E.M. & Russel, R.L. PNAS 72, 4061-4065 (1975). 2. Mori, I. & Ohshima, Y. Nature 376, 344-348 (1995). 3. Pierce-Shimomura, J.T. & Lockery, S.R. J. Neurosci. 19, 9557-9569(1999).