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 laboratory as in nature. The base-substitution spectrum differs between C. elegans laboratory mutation accumulation (MA) experiments and the standing site-frequency spectrum, which has been argued to be in part owing to increased oxidative stress in the laboratory environment. Using genome sequence data from C. elegans MA lines carrying a mutation (mev-1) that increases the cellular titer of reactive oxygen species (ROS), leading to increased oxidative stress, we find the base-substitution spectrum is similar between mev-1, its wild-type progenitor (N2), and another set of MA lines derived from a different wild strain (PB306). Conversely, the rate of short insertions is greater in mev-1, consistent with studies in other organisms in which environmental stress increased the rate of insertion-deletion mutations. Further, the mutational properties of mononucleotide repeats in all strains are different from those of nonmononucleotide sequence, both for indels and base-substitutions, and whereas the nonmononucleotide spectra are fairly similar between MA lines and wild isolates, the mononucleotide spectra are very different, with a greater frequency of A:T → T:A transversions and an increased proportion of ±1-bp indels. The discrepancy in mutational spectra between laboratory MA experiments and natural variation is likely owing to a consistent (but unknown) effect of the laboratory environment that manifests itself via different modes of mutability and/or repair at mononucleotide loci.

Reverse Plasticity Underlies Rapid Evolution by Clonal Selection within Populations of Fibroblasts Propagated on a Novel Soft Substrate.

Mechanical properties such as substrate stiffness are a ubiquitous feature of a cell's environment. Many types of animal cells exhibit canonical phenotypic plasticity when grown on substrates of differing stiffness, in vitro and in vivo. Whether such plasticity is a multivariate optimum due to hundreds of millions of years of animal evolution, or instead is a compromise between conflicting selective demands, is unknown. We addressed these questions by means of experimental evolution of populations of mouse fibroblasts propagated for approximately 90 cell generations on soft or stiff substrates. The ancestral cells grow twice as fast on stiff substrate as on soft substrate and exhibit the canonical phenotypic plasticity. Soft-selected lines derived from a genetically diverse ancestral population increased growth rate on soft substrate to the ancestral level on stiff substrate and evolved the same multivariate phenotype. The pattern of plasticity in the soft-selected lines was opposite of the ancestral pattern, suggesting that reverse plasticity underlies the observed rapid evolution. Conversely, growth rate and phenotypes did not change in selected lines derived from clonal cells. Overall, our results suggest that the changes were the result of genetic evolution and not phenotypic plasticity per se. Whole-transcriptome analysis revealed consistent differentiation between ancestral and soft-selected populations, and that both emergent phenotypes and gene expression tended to revert in the soft-selected lines. However, the selected populations appear to have achieved the same phenotypic outcome by means of at least two distinct transcriptional architectures related to mechanotransduction and proliferation.

Balancing selection maintains ancient polymorphisms at conserved enhancers for the olfactory receptor genes of a Chinese marine fish.

The study of balancing selection, as a selective force maintaining adaptive genetic variation in gene pools longer than expected by drift, is currently experiencing renewed interest due to the increased availability of new data, methods of analysis, and case studies. In this investigation, evidence of balancing selection operating on conserved enhancers of the olfactory receptor (OR) genes is presented for the Chinese sleeper (Bostrychus sinensis), a coastal marine fish that is emerging as a model species for evolutionary studies in the Northwest Pacific marginal seas. Coupled with tests for Gene Ontology enrichment and transcription factor binding, population genomic data allow for the identification of an OR cluster in the sleeper with a downstream flanking region containing three enhancers that are conserved with human and other fish species. Phylogenetic and population genetic analyses indicate that the enhancers are under balancing selection as evidenced by their translineage polymorphisms, excess common alleles, and increased within-group diversities. Age comparisons between the translineage polymorphisms and most recent common ancestors of neutral genealogies substantiate that the former are old, and thus, due to ancient balancing selection. The survival and reproduction of vertebrates depend on their sense of smell, and thereby, on their ORs. In addition to locus duplication and allelic variation of structural genes, this study highlights a third mechanism by which receptor diversity can be achieved for detecting and responding to the huge variety of environmental odorants (i.e., by balancing selection acting on OR gene expression through their enhancer variability).

Evolution of the Mutational Process under Relaxed Selection in Caenorhabditis elegans.

The mutational process varies at many levels, from within genomes to among taxa. Many mechanisms have been linked to variation in mutation, but understanding of the evolution of the mutational process is rudimentary. Physiological condition is often implicated as a source of variation in microbial mutation rate and may contribute to mutation rate variation in multicellular organisms.Deleterious mutations are an ubiquitous source of variation in condition. We test the hypothesis that the mutational process depends on the underlying mutation load in two groups of Caenorhabditis elegans mutation accumulation (MA) lines that differ in their starting mutation loads. "First-order MA" (O1MA) lines maintained under minimal selection for ∼250 generations were divided into high-fitness and low-fitness groups and sets of "second-order MA" (O2MA) lines derived from each O1MA line were maintained for ∼150 additional generations. Genomes of 48 O2MA lines and their progenitors were sequenced. There is significant variation among O2MA lines in base-substitution rate (µbs), but no effect of initial fitness; the indel rate is greater in high-fitness O2MA lines. Overall, µbs is positively correlated with recombination and proximity to short tandem repeats and negatively correlated with 10 bp and 1 kb GC content. However, probability of mutation is sufficiently predicted by the three-nucleotide motif alone. Approximately 90% of the variance in standing nucleotide variation is explained by mutability. Total mutation rate increased in the O2MA lines, as predicted by the "drift barrier" model of mutation rate evolution. These data, combined with experimental estimates of fitness, suggest that epistasis is synergistic.

The mutational decay of male-male and hermaphrodite-hermaphrodite competitive fitness in the androdioecious nematode C. elegans.

Androdioecious Caenorhabditis have a high frequency of self-compatible hermaphrodites and a low frequency of males. The effects of mutations on male fitness are of interest for two reasons. First, when males are rare, selection on male-specific mutations is less efficient than in hermaphrodites. Second, males may present a larger mutational target than hermaphrodites because of the different ways in which fitness accrues in the two sexes. We report the first estimates of male-specific mutational effects in an androdioecious organism. The rate of male-specific inviable or sterile mutations is ⩽5 × 10-4/generation, below the rate at which males would be lost solely due to those kinds of mutations. The rate of mutational decay of male competitive fitness is ~ 0.17%/generation; that of hermaphrodite competitive fitness is ~ 0.11%/generation. The point estimate of ~ 1.5X faster rate of mutational decay of male fitness is nearly identical to the same ratio in Drosophila. Estimates of mutational variance (VM) for male mating success and competitive fitness are not significantly different from zero, whereas VM for hermaphrodite competitive fitness is similar to that of non-competitive fitness. Two independent estimates of the average selection coefficient against mutations affecting hermaphrodite competitive fitness agree to within two-fold, 0.33-0.5%.

A review of nanotechnology in enzyme cascade to address challenges in pre-treating biomass.

Nanotechnology has become a revolutionary technique for improving the preliminary treatment of lignocellulosic biomass in the production of biofuels. Traditional methods of pre-treatment have encountered difficulties in effectively degrading the intricate lignocellulosic composition, thereby impeding the conversion of biomass into fermentable sugars. Nanotechnology has enabled the development of enzyme cascade processes that present a potential solution for addressing the limitations. The focus of this review article is to delve into the utilization of nanotechnology in the pretreatment of lignocellulosic biomass through enzyme cascade processes. The review commences with an analysis of the composition and structure of lignocellulosic biomass, followed by a discussion on the drawbacks associated with conventional pre-treatment techniques. The subsequent analysis explores the importance of efficient pre-treatment methods in the context of biofuel production. We thoroughly investigate the utilization of nanotechnology in the pre-treatment of enzyme cascades across three distinct sections. Nanomaterials for enzyme immobilization, enhanced enzyme stability and activity through nanotechnology, and nanocarriers for controlled enzyme delivery. Moreover, the techniques used to analyse nanomaterials and the interactions between enzymes and nanomaterials are introduced. This review emphasizes the significance of comprehending the mechanisms underlying the synergy between nanotechnology and enzymes establishing sustainable and environmentally friendly nanotechnology applications.

Second-generation biorefineries: single platform for the conversion of lignocellulosic wastes to environmentally important biofuels.

The continuously increasing demands for various fossil fuels to achieve the day-to-day needs of the human population are growing and causing adverse effects on the environment and leading to the depletion of their natural resources. To overcome such drastic problems and minimize the production of greenhouse gases, lignocellulose biomass, which is an abundant and bio-renewable source present on earth with excellent properties and composition, has been used for decades to develop biofuels that can easily take over the place of conventional fuels. Lignocellulose biomass comprises polymeric sugars, i.e., cellulose and hemicellulose, and aromatic polymer, lignin, which are responsible for producing various bio-based products. However, utilizing lignocellulosic wastes for such purposes is needed but their recalcitrant structure makes it difficult to achieve their full usage. For this, several pretreatment approaches are developed to loosen the complexity between sugars and lignin. In some way, few of the conventional pretreatment methods are expensive, non-eco-friendly, and produce undesired by-products, causing a lower yield and reusability of enzymes used in the reaction. Utilizing novel pretreatment strategies that are cost-effective, help in increasing the yield of products, and are environment-friendly is required. Thus, incorporating nanoparticles and nanomaterials in the development of pretreatment and other strategies for the production of bio-based products is currently thriving. This review is designed in such a way that the readers can easily get brief knowledge about the production of important biofuels developed within second-generation biorefineries using lignocellulosic biomass. It also summarizes the importance of nanotechnology in different steps of biofuel development.

Direct inference of the distribution of fitness effects of spontaneous mutations from recombinant inbred Caenorhabditis elegans mutation accumulation lines.

The distribution of fitness effects of new mutations plays a central role in evolutionary biology. Estimates of the distribution of fitness effect from experimental mutation accumulation lines are compromised by the complete linkage disequilibrium between mutations in different lines. To reduce the linkage disequilibrium, we constructed 2 sets of recombinant inbred lines from a cross of 2 Caenorhabditis elegans mutation accumulation lines. One set of lines ("RIAILs") was intercrossed for 10 generations prior to 10 generations of selfing; the second set of lines ("RILs") omitted the intercrossing. Residual linkage disequilibrium in the RIAILs is much less than in the RILs, which affects the inferred distribution of fitness effect when the sets of lines are analyzed separately. The best-fit model estimated from all lines (RIAILs + RILs) infers a large fraction of mutations with positive effects (∼40%); models that constrain mutations to have negative effects fit much worse. The conclusion is the same using only the RILs. For the RIAILs, however, models that constrain mutations to have negative effects fit nearly as well as models that allow positive effects. When mutations in high linkage disequilibrium are pooled into haplotypes, the inferred distribution of fitness effect becomes increasingly negative-skewed and leptokurtic. We conclude that the conventional wisdom-most mutations have effects near 0, a handful of mutations have effects that are substantially negative, and mutations with positive effects are very rare-is likely correct, and that unless it can be shown otherwise, estimates of the distribution of fitness effect that infer a substantial fraction of mutations with positive effects are likely confounded by linkage disequilibrium.

Current status of metabolic engineering of microorganisms for bioethanol production by effective utilization of pentose sugars of lignocellulosic biomass.

Lignocellulosic biomass, consisting of homo- and heteropolymeric sugars, acts as a substrate for the generation of valuable biochemicals and biomaterials. The readily available hexoses are easily utilized by microbes due to the presence of transporters and native metabolic pathways. But, utilization of pentose sugar viz., xylose and arabinose are still challenging due to several reasons including (i) the absence of the particular native pathways and transporters, (ii) the presence of inhibitors, and (iii) lower uptake of pentose sugars. These challenges can be overcome by manipulating metabolic pathways/glycosidic enzymes cascade by using genetic engineering tools involving inverse-metabolic engineering, ex-vivo isomerization, Adaptive Laboratory Evolution, Directed Metabolic Engineering, etc. Metabolic engineering of bacteria and fungi for the utilization of pentose sugars for bioethanol production is the focus area of research in the current decade. This review outlines current approaches to biofuel development and strategies involved in the metabolic engineering of different microbes that can uptake pentose for bioethanol production.

Head-to-head comparison of three experimental methods of quantifying competitive fitness in C. elegans.

Organismal fitness is relevant in many contexts in biology. The most meaningful experimental measure of fitness is competitive fitness, when two or more entities (e.g., genotypes) are allowed to compete directly. In theory, competitive fitness is simple to measure: an experimental population is initiated with the different types in known proportions and allowed to evolve under experimental conditions to a predefined endpoint. In practice, there are several obstacles to obtaining robust estimates of competitive fitness in multicellular organisms, the most pervasive of which is simply the time it takes to count many individuals of different types from many replicate populations. Methods by which counting can be automated in high throughput are desirable, but for automated methods to be useful, the bias and technical variance associated with the method must be (a) known, and (b) sufficiently small relative to other sources of bias and variance to make the effort worthwhile. The nematode Caenorhabditis elegans is an important model organism, and the fitness effects of genotype and environmental conditions are often of interest. We report a comparison of three experimental methods of quantifying competitive fitness, in which wild-type strains are competed against GFP-marked competitors under standard laboratory conditions. Population samples were split into three replicates and counted (1) "by eye" from a saved image, (2) from the same image using CellProfiler image analysis software, and (3) with a large particle flow cytometer (a "worm sorter"). From 720 replicate samples, neither the frequency of wild-type worms nor the among-sample variance differed significantly between the three methods. CellProfiler and the worm sorter provide at least a tenfold increase in sample handling speed with little (if any) bias or increase in variance.