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[
International Worm Meeting,
2009]
Why some phenotypic traits are highly plastic or otherwise variable and others are highly invariant (=canalized) in the face of environmental variation remains a puzzle. Data from whole-genome microarray analysis of cDNA collected from large worm cultures suggests that spontaneous mutations reduce the environmental variability of transcript number. In an attempt to better understand the role of transcriptional modulation as a means of maintaining canalized phenotypes, we are investigating the variance between isogenic C. elegans lines as evidenced by the variability in gene expression in much smaller samples. The motivation for using very small samples is to minimize variance propagated by small differences in timing of development. For each of 6 biological replicates, RNA was extracted and pooled from 5 L3 worms, converted to cDNA, amplified, labeled, and hybridized to an Affymetrix GeneChip. Measures of variability between biologically replicated transcriptomes allow us to estimate transcriptional variance when genetic variation is minimized. These data are the foundation for future research comparing ancestral control lines to mutation accumulation lines for which mutational parameters have been well characterized.
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[
International Worm Meeting,
2013]
To what degree natural selection has shaped the rate of spontaneous mutations among different taxa remains an important unresolved question in evolutionary biology. While mutation rates are known to vary among and within taxa, the relative importance of natural selection versus non-adaptive processes has yet to be determined. Classical theory predicts that the strength of natural selection to reduce the deleterious mutation rate should be stronger in asexual and selfing taxa than in outcrossing sexual taxa, leading to an adaptive decrease in mutation rate in the former. However, other theory predicts (1) that "mutator" alleles can hitchhike to high frequency in asexual/selfing taxa, thereby leading to a (non-adaptive) increase in the mutation rate, and (2) the efficiency of selection to reduce mutation rate may be substantially greater in outcrossing than asexual or selfing taxa due to the larger effective population sizes of the former. Whether general trends in exist in nature is currently unknown. Nematodes in the genus Caenorhabditis provide an ideal system to test questions of how mutation rates vary among closely related species with different reproductive strategies. Within the genus the ancestral reproductive state within is outcrossing (gonochorism), however self-fertilization (hermaphroditism) has evolved independently several times. To examine the role of matting system on the evolution of mutation rates we constructed a set of long-term mutation accumulation (MA) lines of the outcrossing species Caenorhabditis remanei. MA lines were maintained by transferring a single male-female pair of worm per generation. After 122 generations of MA, five randomly selected MA lines along with the ancestral control were re-sequenced using Illumina sequencing technology.
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[
International Worm Meeting,
2017]
The rate and molecular spectrum of mutations varies within and among genomes, species and higher taxa. The understanding of the mechanistic causes of variation is far from complete; the understanding of evolutionary causes is even more rudimentary. It has been posited that individuals in poor physiological condition will experience higher rates of mutation than those in good condition. Since deleterious mutations lead to poor condition, the possibility exists that individuals carrying a high mutation load will experience an elevated mutation rate. We tested that hypothesis with a set of "second-order mutation accumulation" (O2MA) lines of the nematode C. elegans. MA lines that had accumulated mutations under minimal selection for ~250 generations ("first-order MA lines", O1MA) were sorted into high-fitness and low-fitness groups, replicated into new sets of O2MA lines, and allowed to accumulate mutations for another ~150 generations of minimal selection. Whole-genome sequencing of 48 O2MA lines and their O1MA ancestors revealed significant variation in base-substitution rate and in total mutation rate both among the O1MA lines measured at generation 250 and in the O2MA lines measured at G400. However, O2MA rate does not depend on ancestral fitness, nor does the specific O1MA ancestry explain a significant fraction of the variance among O2MA lines. Thus, the signal of variation in mutation rate decays on the order of a hundred generations. Multiple logistic regression of mutability on a set of predictor variables revealed that local three-base nucleotide context is the most important predictor of mutability, but that GC content of the 1 Kb surrounding a site and - importantly - local recombination rate are also significant predictors. Mutability explains a large fraction of the variance in standing nucleotide diversity. The deletion rate of low-fitness O2MA lines is less than that of high-fitness O2 lines. However, low-fitness O1MA lines carry more indels of putatively large effect than do high-fitness O1MA lines, whereas low-fitness O2MA lines carry fewer indels of putatively large effect than the high-fitness O2MA lines. Consistent with those findings, the mean mutational effect on experimentally-measured fitness is twice as great in the low-fitness O2MA lines as in the high-fitness lines. Taken together, these results strongly imply that, on average, epistasis is synergistic among new deleterious mutations.
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[
International Worm Meeting,
2019]
Mutation Accumulation (MA) experiments have been a workhorse of evolutionary genetics for over fifty years, and much of what is known about the mutational process at both the phenotypic and molecular level has been learned from MA experiments. The advent of economical whole-genome short-read sequencing greatly increased our ability to characterize the rate and spectrum of base-substitution and short indel mutations. Copy-number variants (CNVs) and structural variants (SVs, e.g., inversions, translocations) have been more difficult to characterize. The paucity of information on the mutational properties of CNVs and SVs is significant because there is compelling evidence that those types of mutations underlie much variation in complex traits. We previously demonstrated by means of a "second-order MA" experiment (i.e., sets of MA lines founded from individual MA lines) with the nematode Caenorhabditis elegans that the rate of base-substitution and (especially) small indel mutations increased over the course of ~250 generations of minimal selection. Preliminary analyses of CNVs based on short-read Illumina sequence data suggested that a similar trend holds for CNVs, but the quantitative estimates of CNV mutation varied by over an order of magnitude depending on the input parameters of the analytical algorithm. To attempt to get a better handle on the CNV and SV mutation rates, we sequenced a small number of MA lines (N=5) with Pacific Biosciences long-read sequencing and used the resulting estimates to attempt to inform the analysis of our more copious short-read data (N>100) MA lines. Preliminary results suggest a conservative long CNV rate of approximately 5% that of small mutations (base subs + short indels). The data are biased toward deletions, but comparisons of the sequence of the ancestor of our MA lines with the C. elegans reference genome indicates that our analytical method has the capacity to detect insertions and inversions as well as deletions. We additionally report comparative MA data from an additional set of 25 C. elegans MA lines derived from a different starting genotype (PB306).
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Baer, Charles F., Lewis, Suzanna, Upadhyay, Ambuj, Ostrow, Dejerianne, Sylvestre, Laurence, Regan, Kerry, Tabman, Brandon, Matsuba, Chikako, Salomon, Matthew
[
International Worm Meeting,
2011]
A long-standing hypothesis in the field of molecular evolution is that the rate of molecular evolution is positively correlated with metabolic rate. Implicit in this idea (sometimes made explicit) is that the mutation rate is positively correlated with metabolic rate. Unfortunately, several factors are confounded with metabolic rate, including generation time, population size and temperature. Further, to the extent that metabolic rate is correlated with temperature (strongly in ectotherms, less strongly in endotherms), direct mutagenic effects of temperature are often further confounded with environmental "stress". Here we report results from an experiment designed to de-confound the effects of temperature, generation time, evolutionary history, and environmental stress to investigate the relationship between temperature and mutation rate. We allowed spontaneous mutations to accumulate under relaxed selection in C. briggsae (PB800 strain) and C. elegans (N2 strain) at low temperature (18 deg C) for 100 generations and high temperature (26 deg C) for 170 generations. The high temperature environment is markedly stressful for C. elegans by the objective criteria of reduced fecundity and survivorship whereas it is not for C. briggsae. We assayed a demographic measure of fitness relative to the unmutated common ancestor ("control") in both sets of mutation accumulation (MA) lines at both temperatures. The results are both interesting and complicated. In all four cases (both sets of MA lines in both species), the cumulative decline in fitness was greater when assayed at high temperature, although the effect was small in the C. briggsae high temperature MA lines. This result suggests that there is a significant class of mutations with (high) temperature-dependent effects. In C. briggsae, high-temperature MA lines decline in fitness faster than low-temperature MA lines, and that result does not depend (much) on the assay temperature. Thus, it appears that temperature itself is mutagenic in C. briggsae. Conversely, in C. elegans, the cumulative effects of mutations are greater when assayed at high temperature than at low temperature, but there is little difference between high and low-temperature MA lines. This result suggests that in C. elegans, on average, there is no relationship between temperature and mutation rate. Direct characterization of the molecular mutational spectrum is underway and will help resolve the relative contributions of mutation rate and allelic effects to the cumulative mutational decay in fitness.