Biology subjects

Andersen, E. C.

Publications and source records attributed to Andersen, E. C..

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bioRxiv2021

easyXpress: An R package to analyze and visualize high-throughput C. elegans microscopy data generated using CellProfiler

High-throughput imaging techniques have become widespread in many fields of biology. These powerful platforms generate large quantities of data that can be difficult to process and visualize efficiently using existing tools. We developed easyXpress to process and review C. elegans high-throughput microscopy data in the R environment. The package provides a logical workflow for the reading, analysis, and visualization of data generated using CellProfilers WormToolbox. We equipped easyXpress with powerful functions to customize the filtering of noise in data, specifically by identifying and removing objects that deviate from expected animal measurements. This flexibility in data filtering allows users to optimize their analysis pipeline to match their needs. In addition, easyXpress includes tools for generating detailed visualizations, allowing the user to interactively compare summary statistics across wells and plates with ease. Researchers studying C. elegans benefit from this streamlined and extensible package as it is complementary to CellProfiler and leverages the R environment to rapidly process and analyze large high-throughput imaging datasets.

bioinformatics↗

bioRxiv2021

Physical constraints on growth dynamics guide C. elegans developmental trajectories and animal shape

Growth control establishes organism size, requiring mechanisms to sense and adjust growth during development. Studies of single cells revealed that size homeostasis uses distinct control methods. In multicellular organisms, mechanisms that regulate single cell growth must integrate control across organs and tissues during development to generate adult size and shape. We leveraged the roundworm Caenorhabditis elegans as a scalable and tractable model to collect precise growth measurements of thousands of individuals, measure feeding behavior, and quantify changes in animal size and shape during a densely sampled developmental time course. As animals transitioned from one developmental stage to the next, we observed changes in body aspect ratio while body volume remained constant. Then, we modeled a physical mechanism by which constraints on cuticle stretch could cause changes in C. elegans body shape. The model-predicted shape changes are consistent with those observed in the data. Theoretically, cuticle stretch could be sensed by the animal to initiate larval-stage transitions, providing a means for physical constraints to influence developmental timing and growth rate in C. elegans. HighlightsO_LIBody size measurements of thousands of animals in a dense developmental time course C_LIO_LIGrowth rate exhibits nonlinear dynamics in both length and width C_LIO_LIChanges in body shape but not volume occur during periods of increased quiescence C_LIO_LIDynamics of animal shape consistent with a length-based threshold in cuticle stretch C_LIO_LIModeling of cuticle stretch dynamics suggests a novel mode for growth control C_LI

developmental biology↗

bioRxiv2021

Long-Term Metabolomics Reference Material

The use of quality control samples in metabolomics ensures data quality, reproducibility and comparability between studies, analytical platforms and laboratories. Long-term, stable and sustainable reference materials (RMs) are a critical component of the QA/QC system, however, the limited selection of currently available matrix matched RMs reduce their applicability for widespread use. To produce a RM in any context, for any matrix that is robust to changes over the course of time we developed IBAT (Iterative Batch Averaging meThod). To illustrate this method, we generated 11 independently grown E. coli batches and made a RM over the course of 10 IBAT iterations. We measured the variance of these materials by NMR and showed that IBAT produces a stable and sustainable RM over time. This E. coli RM was then used as food source to produce a C. elegans RM for a metabolomics experiment. The metabolite extraction of this material alongside 41 independently grown individual C. elegans samples of the same genotype, allowed to estimate the proportion of sample variation in pre-analytical steps. From the NMR data, we found that 40% of the metabolite variance is due to the metabolite extraction process and analysis and 60% is due to sample-to-sample variance. The availability of RMs in untargeted metabolomics is one of the predominant needs of the metabolomics community that reach beyond quality control practices. IBAT addresses this need by facilitating the production of biologically relevant RMs and increasing their widespread use.

biochemistry↗

bioRxiv2021

Natural variation in fertility is correlated with species-wide levels of divergence in Caenorhabditis elegans

Life history traits underlie the fitness of organisms and are under strong natural selection in the face of environmental challenges. A new mutation that positively impacts a life history trait will likely increase in frequency and become fixed in a population (e.g. selective sweep). The identification of the beneficial alleles that underlie selective sweeps provides insights into the mechanisms that occurred during the evolution of species. In the global population of Caenorhabditis elegans, we previously identified selective sweeps that have drastically reduced chromosomal-scale genetic diversity in the species. Here, we measured the fertility (viable offspring) of a collection of wild C. elegans strains, including many recently isolated divergent strains from the Hawaiian islands and found that strains with larger swept genomic regions on multiple chromosomes have significantly higher fertility than strains that do not have evidence of the recent selective sweeps. We used genome-wide association (GWA) mapping to identify three quantitative trait loci (QTL) underlying the fertility variation. Additionally, we mapped previous fertility data of wild C. elegans strains and C. elegans recombinant inbred advanced intercross lines (RIAILs) that were grown in various conditions and detected eight QTL across the genome using GWA and linkage mappings. These QTL show the genetic complexity of life history traits such as fertility across this species. Moreover, the haplotype structure in each GWA QTL region revealed correlations with recent selective sweeps in the C. elegans population. North American and European strains had significantly higher fertility than most strains from Hawaii, a hypothesized origin of the C. elegans species, suggesting that beneficial alleles that cause increased fertility could underlie the selective sweeps during the worldwide expansion of C. elegans.

genetics↗

bioRxiv2021

Mutability of mononucleotide repeats, not oxidative stress, explains the discrepancy between laboratory-accumulated mutations and the natural allele-frequency spectrum in C. elegans

Important clues about natural selection can be gleaned from discrepancies between the properties of segregating genetic variants and of mutations accumulated experimentally under minimal selection, provided the mutational process is the same in the lab as in nature. The ratio of transitions to transversions (Ts/Tv) is consistently lower in C. elegans mutation accumulation (MA) experiments than in nature, which has been argued to be in part due to increased oxidative stress in the lab environment. Using whole-genome sequence data from a set of C. elegans MA lines carrying a mutation (mev-1) that increases the cellular titer of reactive oxygen species (ROS), leading to increased endogenous oxidative stress, we find that the base-substitution spectrum is similar between mev-1 lines, its wild-type progenitor (N2), and another set of MA lines derived from a different wild strain (PB306). By contrast, the rate of short insertions is greater in the mev-1 lines, consistent with studies in other organisms in which environmental stress led to an increase in the rate of insertion-deletion mutations. Further, the mutational properties of mononucleotide repeats in all strains are qualitatively different from those of non-mononucleotide sequence, both for indels and base-substitutions, and whereas the non-mononucleotide spectra are fairly similar between MA lines and wild isolates, the mononucleotide spectra are very different. The discrepancy in mutational spectra between lab MA experiments and natural variation is likely due to a consistent (but unknown) effect of the lab environment that manifests itself via different modes of mutability and/or repair at mononucleotide loci.

evolutionary biology↗

bioRxiv2021

Two novel loci underlie natural differences in Caenorhabditis elegans macrocyclic lactone responses

Parasitic nematodes cause a massive worldwide burden on human health along with a loss of livestock and agriculture productivity. Anthelmintics have been widely successful in treating parasitic nematodes. However, resistance is increasing, and little is known about the molecular and genetic causes of resistance. The free-living roundworm Caenorhabditis elegans provides a tractable model to identify genes that underlie resistance. Unlike parasitic nematodes, C. elegans is easy to maintain in the laboratory, has a complete and well annotated genome, and has many genetic tools. Using a combination of wild isolates and a panel of recombinant inbred lines constructed from crosses of two genetically and phenotypically divergent strains, we identified three genomic regions on chromosome V that underlie natural differences in response to the macrocyclic lactone (ML) abamectin. One locus was identified previously and encodes an alpha subunit of a glutamate-gated chloride channel (glc-1). Here, we validate and narrow two novel loci using near-isogenic lines. Additionally, we generate a list of prioritized candidate genes identified in C. elegans and in the parasite Haemonchus contortus by comparison of ML resistance loci. These genes could represent previously unidentified resistance genes shared across nematode species and should be evaluated in the future. Our work highlights the advantages of using C. elegans as a model to better understand ML resistance in parasitic nematodes. Author SummaryParasitic nematodes infect plants, animals, and humans, causing major health and economic burdens worldwide. Parasitic nematode infections are generally treated efficiently with a class of drugs named anthelmintics. However, resistance to many of these anthelmintic drugs, including macrocyclic lactones (MLs), is rampant and increasing. Therefore, it is essential that we understand how these drugs act against parasitic nematodes and, conversely, how nematodes gain resistance in order to better treat these infections in the future. Here, we used the non-parasitic nematode Caenorhabditis elegans as a model organism to study ML resistance. We leveraged natural genetic variation between strains of C. elegans with differential responses to abamectin to identify three genomic regions on chromosome V, each containing one or more genes that contribute to ML resistance. Two of these loci have not been previously discovered and likely represent novel resistance mechanisms. We also compared the genes in these two novel loci to the genes found within genomic regions linked to ML resistance in the parasite Haemonchus contortus and found several cases of overlap between the two species. Overall, this study highlights the advantages of using C. elegans to understand anthelmintic resistance in parasitic nematodes.

genetics↗

bioRxiv2020

The distribution of mutational effects on fitness in Caenorhabditis elegans inferred from standing genetic variation

The distribution of fitness effects for new mutations is one of the most theoretically important but difficult to estimate properties in population genetics. A crucial challenge to inferring the distribution of fitness effects (DFE) from natural genetic variation is the sensitivity of the site frequency spectrum to factors like population size change, population substructure, and non-random mating. Although inference methods aim to control for population size changes, the influence of non-random mating remains incompletely understood, despite being a common feature of many species. We report the distribution of fitness effects estimated from 326 genomes of Caenorhabditis elegans, a nematode roundworm with a high rate of self-fertilization. We evaluate the robustness of DFE inferences using simulated data that mimics the genomic structure and reproductive life history of C. elegans. Our observations demonstrate how the combined influence of self-fertilization, genome structure, and natural selection can conspire to compromise estimates of the DFE from extant polymorphisms. These factors together tend to bias inferences towards weakly deleterious mutations, making it challenging to have full confidence in the inferred DFE of new mutations as deduced from standing genetic variation in species like C. elegans. Improved methods for inferring the distribution of fitness effects are needed to appropriately handle strong linked selection and selfing. These results highlight the importance of understanding the combined effects of processes that can bias our interpretations of evolution in natural populations.

evolutionary biology↗

bioRxiv2020

Selfing is the safest sex for Caenorhabditis tropicalis

Mating systems have profound effects on genetic diversity and compatibility. The convergent evolution of self-fertilization in three Caenorhabditis species provides a powerful lens to examine causes and consequences of mating system transitions. Among the selfers, C. tropicalis is the least genetically diverse and most afflicted by outbreeding depression. We generated a chromosomal-scale genome for C. tropicalis and surveyed global diversity. Population structure is very strong, and islands of extreme divergence punctuate a genomic background that is highly homogeneous around the globe. Outbreeding depression in the laboratory is caused largely by multiple gene drive elements, genetically consistent with maternal toxin/zygotic antidote systems. Driver loci harbor novel and duplicated genes, and their activity is modified by mito-nuclear background. Segregating drivers dramatically reduce fitness, and simulations show that selfing limits their spread. Frequent selfing in C. tropicalis may therefore be a strategy to avoid drive-mediated outbreeding depression.

evolutionary biology↗

bioRxiv2020

Balancing selection maintains ancient genetic diversity in C. elegans

Across diverse taxa, selfing species have evolved independently from outcrossing species thousands of times. The transition from outcrossing to selfing significantly decreases the effective population size, effective recombination rate, and heterozygosity within a species. These changes lead to a reduction in genetic diversity, and therefore adaptive potential, by intensifying the effects of random genetic drift and linked selection. Within the nematode genus Caenorhabditis, selfing has evolved at least three times and all three species, including in the model organism Caenorhabditis elegans, show substantially reduced genetic diversity relative to outcrossing species. Selfing and outcrossing Caenorhabditis species are often found in the same niches, but we still do not know how selfing species with limited genetic diversity can adapt to these environments. Here, we examine the whole-genome sequences from 609 wild C. elegans strains isolated worldwide and show that genetic variation is concentrated in punctuated hyper-divergent regions that cover 20% of the C. elegans reference genome. These regions are enriched in environmental response genes that mediate sensory perception, pathogen response, and xenobiotic stress response. Population genomic evidence suggests that genetic diversity in these regions has been maintained by long-term balancing selection. Using long-read genome assemblies for 15 wild strains, we show that hyper-divergent haplotypes contain unique sets of genes and show levels of divergence comparable to levels found between Caenorhabditis species that diverged millions of years ago. These results provide an example for how species can avoid the evolutionary "dead end" associated with selfing.

evolutionary biology↗

bioRxiv2019

Deep sampling of Hawaiian Caenorhabditis elegans reveals high genetic diversity and admixture with global populations

Recent efforts to understand the natural niche of the keystone model organism Caenorhabditis elegans have suggested that this species is cosmopolitan and associated with rotting vegetation and fruits. However, most of the strains isolated from nature have low genetic diversity likely because recent chromosome-scale selective sweeps contain alleles that increase fitness in human-associated habitats. Strains from the Hawaii Islands are highly divergent from non-Hawaiian strains. This result suggests that Hawaiian strains might contain ancestral genetic diversity that was purged from most non-Hawaiian strains by the selective sweeps. To characterize the genetic diversity and niche of Hawaiian C. elegans, we sampled across the Hawaiian Islands and isolated 100 new C. elegans strains. We found that C. elegans strains are not associated with any one substrate but are found in cooler climates at high elevations. These Hawaiian strains are highly diverged compared to the rest of the global population. Admixture analysis identified 11 global populations, four of which are from Hawaii. Surprisingly, one of the Hawaiian populations shares recent ancestry with non-Hawaiian populations, including portions of globally swept haplotypes. This discovery provides the first evidence of gene flow between Hawaiian and non-Hawaiian populations. Most importantly, the high levels of diversity observed in Hawaiian strains might represent the complex patterns of ancestral genetic diversity in the C. elegans species before human influence.

ecology↗

bioRxiv2019

Population selection and sequencing of C. elegans wild isolates identifies a region on chromosome III affecting starvation resistance

To understand the genetic basis of complex traits, it is important to be able to efficiently phenotype many genetically distinct individuals. In the nematode Caenorhabditis elegans, individuals have been isolated from diverse populations around the globe and whole-genome sequenced. As a result, hundreds of wild strains with known genome sequences can be used for genome-wide association studies (GWAS). However, phenotypic analysis of these strains can be laborious, particularly for quantitative traits requiring multiple measurements per strain. Starvation resistance is likely a fitness-proximal trait for nematodes, and it is related to metabolic disease risk in humans. However, natural variation in C. elegans starvation resistance has not been characterized, and precise measurement of the trait is time-intensive. Here, we developed a population selection and sequencing-based approach to phenotype starvation resistance in a pool of 96 wild strains. We used restriction site-associated DNA sequencing (RAD-seq) to infer the frequency of each strain among survivors in a mixed culture over time during starvation. We used manual starvation survival assays to validate the trait data, confirming that strains that increased in frequency over time are starvation-resistant relative to strains that decreased in frequency. These results document natural variation in starvation resistance. Further, we found that variation in starvation resistance is significantly associated with variation at a region on chromosome III. Using a near-isogenic line (NIL), we showed the importance of this genomic interval for starvation resistance. This study demonstrates the feasibility of using population selection and sequencing in an animal model for phenotypic analyses of quantitative traits, reveals natural variation of starvation resistance in C. elegans, and identifies a genomic region that contributes to such variation.

genetics↗

bioRxiv2019

Selection and gene flow shape niche-associated copy-number variation of pheromone receptor genes

From quorum sensing in bacteria to pheromone signaling in social insects, chemical communication mediates interactions among individuals in a local population. In Caenorhabditis elegans, ascaroside pheromones can dictate local population density, in which high levels of pheromones inhibit the reproductive maturation of individuals. Little is known about how natural genetic diversity affects the pheromone responses of individuals from diverse habitats. Here, we show that a niche-associated copy-number variation (CNV) of pheromone receptor genes contributes to natural differences in pheromone responses. We found putative loss-of-function deletions that reduce copy number of duplicated pheromone receptor genes (srg-36 and srg-37), which were shown previously to be selected in population-dense laboratory cultures. A common natural deletion in the less functional copy (srg-37) arose from a single ancestral population that spread throughout the world and underlies reduced pheromone sensitivity across the global C. elegans population. This deletion is enriched in wild strains that were isolated from a rotting fruit niche, where proliferating populations are often found. Taken together, these results demonstrate that selection and gene flow together shape the copy number of pheromone receptor genes in natural C. elegans populations to facilitate local adaptation to diverse niches.

evolutionary biology↗

bioRxiv2019

A nematode-specific gene underlies bleomycin-response variation in Caenorhabditis elegans

Bleomycin is a powerful chemotherapeutic drug used to treat a variety of cancers. However, individual patients vary in their responses to bleomycin. The identification of genetic differences that underlie this response variation could improve treatment outcomes by tailoring bleomycin dosages to each patient. We used the model organism Caenorhabditis elegans to identify genetic determinants of bleomycin-response differences by performing linkage mapping on recombinants derived from a cross between the laboratory strain (N2) and a wild strain (CB4856). This approach identified a small genomic region on chromosome V that underlies bleomycin-response variation. Using near-isogenic lines and strains with CRISPR-Cas9 mediated deletions and allele replacements, we discovered that a novel nematode-specific gene (scb-1) is required for bleomycin resistance. Although the mechanism by which this gene causes variation in bleomycin responses is unknown, we suggest that a rare variant present in the CB4856 strain might cause differences in the potential stress-response function of scb-1 between the N2 and CB4856 strains, thereby leading to differences in bleomycin resistance.

genetics↗

bioRxiv2018

Tightly-linked antagonistic-effect loci underlie polygenic demographic variation in C. elegans

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. This view is hard to reconcile with the experience of researchers who define gene functions using mutagenesis, comparing mutants one at a time to the wild type. Here, we use the approach of experimental genetics to show that indeed, most genomic regions carry variants with detectable effects on complex traits. We used high-throughput phenotyping to characterize demography as a multivariate trait in growing populations of Caenorhabditis elegans sensitized by nickel stress. We show that demography under these conditions is genetically complex in a panel of recombinant inbred lines. We then focused on a 1.4-Mb region of the X chromosome. When we compared two near isogenic lines (NILs) that differ only at this region, they were phenotypically indistinguishable. When we used additional NILs to subdivide the region into fifteen intervals, each encompassing ~0.001 of the genome, we found that eleven of intervals have significant effects. These effects are often similar in magnitude to those of genome-wide significant QTLs mapped in the recombinant inbred lines but are antagonized by the effects of variants in adjacent intervals. 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.

genetics↗

bioRxiv2018

Natural variation in arsenic toxicity is explained by differences in branched chain amino acid catabolism

We find that variation in the dbt-1 gene underlies natural differences in Caenorhabditis elegans responses to the toxin arsenic. This gene encodes the E2 subunit of the branched-chain -keto acid dehydrogenase (BCKDH) complex, a core component of branched-chain amino acid (BCAA) metabolism. We causally linked a non-synonymous variant in the conserved lipoyl domain of DBT-1 to differential arsenic responses. Using targeted metabolomics and chemical supplementation, we demonstrate that differences in responses to arsenic are caused by variation in iso-branched chain fatty acids. Additionally, we show that levels of branched chain fatty acids in human cells are perturbed by arsenic treatment. This finding has broad implications for arsenic toxicity and for arsenic-focused chemotherapeutics across human populations. Our study implicates the BCKDH complex and BCAA metabolism in arsenic responses, demonstrating the power of C. elegans natural genetic diversity to identify novel mechanisms by which environmental toxins affect organismal physiology.

genetics↗

bioRxiv2018

Extreme allelic heterogeneity at a Caenorhabditis elegans beta-tubulin locus explains natural resistance to benzimidazoles

Benzimidazoles (BZ) are essential components of the limited chemotherapeutic arsenal available to control the global burden of parasitic nematodes. The emerging threat of BZ resistance among nearly all nematode species necessitates the development of novel strategies to identify genetic and molecular mechanisms underlying this resistance. All detection of parasitic helminth resistance to BZ is focused on the genotyping of three variant sites in the orthologs of the {beta}-tubulin gene found to confer resistance in the free-living nematode Caenorhabditis elegans. Because of the limitations of laboratory and field experiments in parasitic nematodes, it is difficult to look beyond these three sites, and additional BZ resistance is observed in the field. Here, we took an unbiased genome-wide mapping approach in the free-living nematode species C. elegans to identify the genetic underpinnings of natural resistance to the commonly used BZ, albendazole (ABZ). We found a wide range of natural variation in ABZ resistance in natural C. elegans populations. In agreement with known mechanisms of BZ resistance in parasites, we find that a majority of the variation in ABZ resistance among wild C. elegans strains is caused by variation in the {beta}-tubulin gene ben-1. This result shows empirically that resistance to ABZ naturally exists and segregates within the C. elegans population, suggesting that selection in natural niches could enrich for resistant alleles. We identified 25 distinct ben-1 alleles that are segregating at low frequencies within the C. elegans population, including many novel molecular variants. Population genetic analyses indicate that ben-1 variation arose multiple times during the evolutionary history of C. elegans and provide evidence that these alleles likely occurred recently because of local selective pressures. Additionally, we find purifying selection at all five {beta}-tubulin genes, despite predicted loss-of-function resistants variants in ben-1, indicating that BZ resistance in natural niches is a stronger selective pressure than loss of one {beta}-tubulin gene. Furthermore, we use genome-editing to show that the most common parasitic nematode {beta}-tubulin allele that confers BZ resistance, F200Y, confers resistance in C. elegans. Importantly, we identified a novel genomic region that is correlated with ABZ resistance in the C. elegans population but independent of ben-1 and the other {beta}-tubulin loci, suggesting that there are multiple mechanisms underlying BZ resistance. Taken together, our results establish a population-level resource of nematode natural diversity as an important model for the study of mechanisms that give rise to BZ resistance.\n\nAuthor summaryNematode parasites have a tremendous impact on human health with almost two billion people infected worldwide. The control of nematode infections relies mainly on the efficacy of a limited repertoire of anthelmintic compounds, including the benzimidazoles (BZ). Already a significant problem in veterinary medicine, increasing evidence exists for the development of BZ resistance in nematodes that infect humans. Laboratory screens and field surveys identified {beta}-tubulin genes as major determinants of BZ resistance in nematodes but detailed population-wide genetic analyses of resistance mechanisms are only just beginning. Therefore, we took advantage of the free-living model organism Caenorhabditis elegans to study the genetic basis of resistance to the commonly used BZ, albendazole (ABZ) in a natural nematode population. Performing genome-wide association mappings, we were able to identify extreme heterogeneity in the {beta}-tubulin gene ben-1 as a major determinant of ABZ resistance. Moreover, our study provided new insights into the effects of missense and loss-of-function alleles at this locus, and how anthelmintic resistance could have developed within a natural nematode population.

genetics↗

bioRxiv2017

Natural variation in a single amino acid underlies cellular responses to topoisomerase II poisons

Many medications, including chemotherapeutics, are differentially effective from one patient to the next. Understanding the causes of these population-wide differences is a critical step towards the development of personalized treatments and improvements to existing medications. Here, we investigate natural differences in sensitivity to anti-neoplastic drugs that target topoisomerase II, using the model organism Caenorhabditis elegans. We show that wild isolates of C. elegans vary in their sensitivity to these drugs, and we use an unbiased statistical and molecular genetics approach to demonstrate that this variation is explained by a methionine-to-glutamine substitution in topoisomerase II (TOP-2). The presence of a non-polar methionine at this residue increases hydrophobic interactions between TOP-2 and the poison etoposide, as compared to a polar glutamine. We hypothesize that this stabilizing interaction results in increased genomic instability in strains that contain a methionine residue. The residue affected by this substitution is conserved from yeast to humans and is one of the few differences between the two human topoisomerase II isoforms (methionine in hTOPII and glutamine in hTOPII{beta}). We go on to show that this substitution influences binding and cytotoxicity of etoposide and two additional topoisomerase II poisons in human cell lines. These results explain why hTOPII and hTOPII{beta} are differentially affected by various poisons and demonstrate the utility of C. elegans in understanding the genetics of drug responses.

genetics↗