Biology subjects

Large, C. R. L.

Publications and source records attributed to Large, C. R. L..

8 recordsLinked to original sources

bioRxiv2025

Variation in social feeding behavior and interactions among Caenorhabditis nematodes

The ability to respond to complex stimuli and environmental cues is essential for organisms to survive and reproduce. Responding to a wide range of stimuli requires a neuronal network that can integrate cues and execute behavioral responses. Evolution of behaviors occurs ubiquitously in most established ecological niches, even among closely related species. To uncover the genetic and neuronal drivers of evolving behaviors, we have taken advantage of the large and relatively ancient divergence in the Caenorhabditis genus to ask how different Caenorhabditis nematodes respond to environmental stimuli and whether behavioral traits are shared or distinct. Here, we assayed foraging behaviors of twelve members of the Caenorhabditis clade, including members of both the elegans and japonica supergroup, and the basal taxon C. monodelphis. For each species, we analyzed social feeding and bordering behaviors, which are well characterized in C. elegans. These behaviors are the functional readout of complex sensory integration of multiple sensory cues including pheromones, touch, O2/CO2 concentration, and attractive and noxious stimuli. We hypothesized that the evolutionary divergence between species would correlate to divergence in these behaviors. We observed a wide variation in social aggregate feeding and bordering behaviors of hermaphrodite and female animals, but the variation did not correlate with evolutionary relatedness of the species. Addition of male animals with female or hermaphrodite animals of the same species increased aggregation behavior of subset of species, but not others. Combination of a second species with C. elegans significantly reduced aggregate feeding behavior of C. elegans, but not the other species. Intraspecies and interspecies interactions therefore modifies behavioral paradigms. Overall, we find that foraging and social feeding behaviors vary widely across Caenorhabditis species, likely due to species-specific responses and integration of environmental and context sensory cues. In general, the clade represents a compelling model to dissect evolution of behavior across diverse environments and a large timescale.

evolutionary biology↗

bioRxiv2024

Overdrive is essential for targeted sperm elimination by Segregation Distorter

Intra-genomic conflict driven by selfish chromosomes is a powerful force that shapes the evolution of genomes and species. In the male germline, many selfish chromosomes bias transmission in their own favor by eliminating spermatids bearing the competing homologous chromosomes. However, the mechanisms of targeted gamete elimination remain mysterious. Here, we show that Overdrive (Ovd), a gene required for both segregation distortion and male sterility in Drosophila pseudoobscura hybrids, is broadly conserved in Dipteran insects but dispensable for viability and fertility. In D. melanogaster, Ovd is required for targeted Responder spermatid elimination after the histone-to-protamine transition in the classical Segregation Distorter system. We propose that Ovd functions as a general spermatid quality checkpoint that is hijacked by independent selfish chromosomes to eliminate competing gametes.

genetics↗

bioRxiv2024

Lineage-resolved analysis of embryonic gene expression evolution in C. elegans and C. briggsae

What constraints govern the evolution of gene expression patterns across development remains a fundamental question of evolutionary biology. The advent of single-cell sequencing opens the possibility of learning these constraints by systematically profiling homologous cells across different organisms. The nematode C. elegans is a well-studied model for embryonic development, and its invariant lineage that is conserved with other Caenorhabditis species makes it an ideal model to directly compare gene expression between homologous progenitor and terminal cell types across evolution. We have measured the spatiotemporal divergence of gene expression across embryogenesis by collecting, annotating, and comparing the transcriptomes of homologous embryonic progenitors and terminal cell types, using a dataset comprising >200,000 C. elegans cells and >190,000 C. briggsae cells. We find a high level of similarity in gene expression programs between the species despite tens of millions of years of evolutionary divergence, consistent with their conserved developmental lineages. Even still, thousands of genes show divergence in their cell-type specific expression patterns, and these are enriched for categories involved in environmental response and behavior. Comparing the degree of expression conservation across cell types reveals that certain cell types such as neurons, have diverged more than others such as the intestine and body wall muscle. Taken together, this work identifies likely constraints on the evolution of developmental gene expression and provides a powerful resource for addressing diverse evolutionary questions.

evolutionary biology↗

bioRxiv2023

Template switching between the leading and lagging strands at replication forks generates inverted copy number variants through hairpin-capped extrachromosomal DNA

Inherited and de novo copy number variants (CNVs) are increasingly found to be correlated with human developmental and cancerous phenotypes. Several models for template switching during replication have been proposed to explain the generation of these gross chromosomal rearrangements. We proposed a model of template switching (ODIRA--origin dependent inverted repeat amplification) in which simultaneous ligation of the leading and lagging strands at diverging replication forks could generate segmental inverted triplications through an extrachromosomal inverted circular intermediate. Here, we created a genetic assay using split-ura3 cassettes to trap the proposed inverted intermediate. However, instead of recovering circular inverted intermediates, we found inverted linear chromosomal fragments ending in native telomeres--suggesting that a template switch had occurred at the centromere-proximal fork of a replication bubble. As telomeric inverted hairpin fragments can also be created through double strand breaks we tested whether replication errors or repair of double stranded DNA breaks were the most likely initiating event. The results from CRISPR/Cas9 cleavage experiments and growth in the replication inhibitor hydroxyurea indicate that it is a replication error, not a double stranded break that creates the inverted junctions. Since inverted amplicons of the SUL1 gene occur during long-term growth in sulfate-limited chemostats, we sequenced evolved populations to look for evidence of linear intermediates formed by an error in replication. All of the data are compatible with a two-step version of the ODIRA model in which sequential template switching at short inverted repeats between the leading and lagging strands at replication forks generates inverted interstitial triplications. AUTHOR SUMMARYChromosomal rearrangements are a potent source of genetic variation in humans and other organisms. One specific type of rearrangement involves the increase in copies of segments of the genome. The variation in gene dosage that these rearrangements can cause has been associated with a wide range of neurological and other human disorders. A specific puzzling form of copy number increase consists of three tandem copies with the central copy in inverted orientation. How this rearrangement occurs is of great interest, yet the mechanisms responsible are only inferred by examining the sequence of final inverted products. Yeast provides a unique model system to explore the underlying molecular defects that give rise to inverted triplications. While the favored hypothesis suggests that double stranded DNA repair is the causative agent, we find that a particular form of template switching between strands at the replication fork, not a double stranded DNA break, is the initiating event. Using the awesome power of yeast genetics, we provide evidence in two different assays for this unique replication error that we call ODIRA (for Origin Dependent Inverted Repeat Amplification) and propose that it can also explain this form of copy number variant seen in human evolution and disease.

genomics↗

bioRxiv2021

yEvo: Experimental evolution in high school classrooms selects for novel mutations and epistatic interactions that impact clotrimazole resistance in S. cerevisiae.

Antifungal resistance in pathogenic fungi is a growing global health concern. Non-pathogenic laboratory strains of Saccharomyces cerevisiae are a useful model for studying mechanisms of antifungal resistance that are relevant to understanding the same processes in pathogenic fungi. We developed a series of lab modules in which high school students used experimental evolution to study antifungal resistance by isolating azole-resistant S. cerevisiae and examining the genetic basis of resistance. All 99 sequenced clones from these experiments possessed mutations previously shown to impact azole resistance, demonstrating the efficacy of our protocols. We additionally found recurrent mutations in an mRNA degradation pathway and an uncharacterized mitochondrial protein (Csf1) that have possible mechanistic connections to azole resistance. The scale of replication in this high school-led initiative allowed us to identify epistatic interactions, as evidenced by pairs of mutations that occur in the same clone more frequently than expected by chance (positive epistasis) or less frequently (negative epistasis). We validated one of these pairs, a negative epistatic interaction between gain-of-function mutations in the multidrug resistance transcription factors Pdr1 and Pdr3. This high school-university collaboration can serve as a model for involving members of the broader public in the scientific process to make meaningful discoveries in biomedical research.

genetics↗

bioRxiv2020

Genomic stability and adaptation of beer brewing yeasts during serial repitching in the brewery

Ale brewing yeast are the result of admixture between diverse strains of Saccharomyces cerevisiae, resulting in a heterozygous tetraploid that has since undergone numerous genomic rearrangements. As a result, comparisons between the genomes of modern related ale brewing strains show both extensive aneuploidy and mitotic recombination that has resulted in a loss of intragenomic diversity. Similar patterns of intraspecific admixture and subsequent selection for one haplotype have been seen in many domesticated crops, potentially reflecting a general pattern of domestication syndrome between these systems. We set out to explore the evolution of the ale brewing yeast, to understand both polyploid evolution and the process of domestication in the ecologically relevant environment of the brewery. Utilizing a common brewery practice known as repitching, in which yeasts are reused over multiple beer fermentations, we generated population time courses from multiple breweries utilizing similar strains of ale yeast. Applying whole-genome sequencing to the time courses, we have found that the same structural variations in the form of aneuploidy and mitotic recombination of particular chromosomes reproducibly rise to detectable frequency during adaptation to brewing conditions across multiple related strains in different breweries. Our results demonstrate that domestication of ale strains is an ongoing process and will likely continue to occur as modern brewing practices develop.

evolutionary biology↗

bioRxiv2019

Phenotypic and genotypic consequences of CRISPR/Cas9 editing of the replication origins in the rDNA of Saccharomyces cerevisiae

The complex structure and repetitive nature of eukaryotic ribosomal DNA (rDNA) is a challenge for genome assembly, and thus, the consequences of sequence variation in rDNA remain unexplored. However, renewed interest in the role that rDNA variation may play in diverse cellular functions, aside from ribosome production, highlights the need for a method that would permit genetic manipulation of the rDNA. Here, we describe a CRISPR/Cas9 based strategy to edit the rDNA locus in the budding yeast Saccharomyces cerevisiae. Using this approach, we modified the endogenous rDNA origin of replication in each repeat by deleting or replacing its consensus sequence. We characterized the transformants that have successfully modified their rDNA locus and propose a mechanism for how CRISPR/Cas9 mediated editing of the rDNA occurs. In addition, we carried out extended growth and life span experiments to investigate the long-term consequences that altering the rDNA origin of replication has on cellular health. We find that long-term growth of the edited clones results in faster growing suppressors that have acquired segmental aneusomy of the rDNA containing region of chr XII or aneuploidy of chromosomes XII, II, or IV. Furthermore, we find that all edited isolates suffer a reduced life span, irrespective of their levels of extrachromosomal rDNA circles. Our work demonstrates that it is possible to quickly, efficiently and homogeneously edit the rDNA locus via CRISPR/Cas9. It serves as a model for modifying other parts of the rDNA and, more generally, for editing other tandemly repeated sequences in higher eukaryotes.

genetics↗

bioRxiv2018

Temperature preference biases parental genome retention during hybrid evolution

Interspecific hybridization can introduce genetic variation that aids in adaptation to new or changing environments. Here we investigate how the environment, and more specifically temperature, interacts with hybrid genomes to alter parental genome representation over time. We evolved Saccharomyces cerevisiae x Saccharomyces uvarum hybrids in nutrient-limited continuous culture at 15{degrees}C for 200 generations. In comparison to previous evolution experiments at 30{degrees}C, we identified a number of temperature specific responses, including the loss of the S. cerevisiae allele in favor of the cryotolerant S. uvarum allele for several portions of the hybrid genome. In particular, we discovered a genotype by environment interaction in the form of a reciprocal loss of heterozygosity event on chromosome XIII. Which species haplotype is lost or maintained is dependent on the parental species temperature preference and the temperature at which the hybrid was evolved. We show that a large contribution to this directionality is due to temperature sensitivity at a single locus, the high affinity phosphate transporter PHO84. This work helps shape our understanding of what forces impact genome evolution after hybridization, and how environmental conditions may favor or disfavor hybrids over time.

evolutionary biology↗