Project description:Oak galls form when gall wasps lay their eggs into part of the tree; in some galls, this attachment point to the host consists of only a few cells. The gall itself comprises entirely of host tissue; however, the initiation, development, and physical appearance are controlled by the inducer. This raises the intriguing question of the molecular mechanisms underlying gall formation, by which one or a small number of cells are reprogrammed and commit to a novel developmental path. Gall wasps undergo two generations each year, and the galls formed by these two generations exhibit markedly different appearances. We sequenced the transcriptomes of both the sexual and asexual generations of Neuropterus quercusbaccarum and Neuroterus numismalis. The transcriptomes of the generations that occur at the same time of year are more similar to each other than they are to the opposite generation of their respective species.
Project description:A striking property of the ancient and obligate mutualism between figs and their pollinating wasps is that fig wasps consistently oviposit in the inner flowers of the fig syconium (gall flowers, which develop into galls that house developing larvae), but typically do not use the outer ring of flowers (seed flowers, which develop into seeds). To better understand differences between gall and seed flowers that might influence oviposition choices, and the unknown mechanisms underlying gall formation, we used a metatranscriptomic approach to analyze eukaryotic gene expression within fig flowers at the time of oviposition choice and early gall development. Consistent with the unbeatable seed hypothesis, which posits that only a portion of fig flowers are physiologically capable of responding to gall induction or supporting larval development, we found significant differences in gene expression assigned to defense and metabolism between gall- and seed flowers in receptive syconia. Transcripts assigned to flavonoids and defense were especially prevalent in receptive gall flowers, and carbohydrate metabolism was significantly up-regulated relative to seed flowers. In turn, high expression of the venom gene icarapin during wasp embryogenesis within galled flowers distinguishes it as a candidate gene for gall initiation. In response to galling, the fig significantly up-regulates the expression of chalcone synthase, which previously has been connected to gall formation in other plants. This study simultaneously evaluates the gene expression profile of both mutualistic partners in a plant-insect mutualism and provides evidence for a stability mechanism in the ancient fig-fig wasp association.
Project description:A striking property of the ancient and obligate mutualism between figs and their pollinating wasps is that fig wasps consistently oviposit in the inner flowers of the fig syconium (gall flowers, which develop into galls that house developing larvae), but typically do not use the outer ring of flowers (seed flowers, which develop into seeds). To better understand differences between gall and seed flowers that might influence oviposition choices, and the unknown mechanisms underlying gall formation, we used a metatranscriptomic approach to analyze eukaryotic gene expression within fig flowers at the time of oviposition choice and early gall development. Consistent with the unbeatable seed hypothesis, which posits that only a portion of fig flowers are physiologically capable of responding to gall induction or supporting larval development, we found significant differences in gene expression assigned to defense and metabolism between gall- and seed flowers in receptive syconia. Transcripts assigned to flavonoids and defense were especially prevalent in receptive gall flowers, and carbohydrate metabolism was significantly up-regulated relative to seed flowers. In turn, high expression of the venom gene icarapin during wasp embryogenesis within galled flowers distinguishes it as a candidate gene for gall initiation. In response to galling, the fig significantly up-regulates the expression of chalcone synthase, which previously has been connected to gall formation in other plants. This study simultaneously evaluates the gene expression profile of both mutualistic partners in a plant-insect mutualism and provides evidence for a stability mechanism in the ancient fig-fig wasp association. We examined two different Ficus flower types at two different time points. Each sample contained a pool of hundreds of individual flowers from multiple sycomia.
Project description:Seasonal photoperiodic changes have strong impact on development in Nasonia vitripennis. Here, Using high-throughput Reduced Representation Bisulfite Sequencing (RRBS) and single-molecule-based sequencing, we generated DNA methylation maps of female wasps maintained in long vs short day. We have identified differential methylated loci that encode the photoperiodic change. analysis of DNA methylation in female wasps maintained in long vs short day, using RRBS followed by Illumina sequencing
Project description:Understanding how plant tissue and organs may be transformed into novel structures by other organisms provides a unique opportunity to study the molecular processes that dictate facets of plant anatomy and morphology. Certain groups of wasps have evolved the ability to transform plant tissues into ornate structures called galls, which provide shelter and nutrition for their larvae. However, the exact mechanism for how gall wasps remodel the plant’s physical structure and metabolism is still largely unknown. At their core, galls alter the morphology and repurpose the function of plant tissue. One common trait that unites all galls is the distinct cellular reprogramming and tumor-like growth that is necessary to produce a gall. There are over 1,400 gall wasps from the family Cynipidae, resulting in a wide diversity of gall structures, shapes, and colors that have been described. Thus, discovering the core molecular determinants that dictate the radical transformation of plant cells will help reveal principles of how plant morphology and function can be rewired by external factors. Little molecular work has been done to elucidate the factors (i.e., genes, proteins, or small molecules) that may be involved in this dramatic repurposing and dedifferentiation of plant tissue. We plan to utilize modern -omics approaches to investigate and identify the molecular factors that underlie the initiation and development of plant galls.
The work (proposal:https://doi.org/10.46936/10.25585/60000461) conducted by the U.S. Department of Energy Joint Genome Institute (https://ror.org/04xm1d337), a DOE Office of Science User Facility, is supported by the Office of Science of the U.S. Department of Energy operated under Contract No. DE-AC02-05CH11231.
Project description:We sequenced DNA from a bulk of Col x Ler F2 hybrid plants (WT and recq4) using Nanopore long-read sequencing and identified crossover sites with COmapper. For nanopore sequencing of gDNA from 1,000 pooled seedlings, 10-day-old seedlings were ground in liquid nitrogen using a mortar and pestle. The ground tissue was resuspended in four volumes of CTAB buffer (1% [w/v] CTAB, 50 mM Tris-HCl pH 8.0, 0.7 M NaCl, 10 mM EDTA) and incubated at 65°C for 30 min. Following chloroform extraction, isopropanol precipitation and removal of RNAs as above, the gDNA pellet was resuspended in 150 μl TE (10 mM Tris-HCl pH 8.0, 0.1 mM EDTA) buffer and gDNA was quantified using a Qubit dsDNA Broad Range assay kit (Thermo Fisher, Q32853). Nine micrograms of gDNA from pollen or seedlings was used to construct a nanopore long-read sequencing library using a Ligation Sequencing Kit V14 (Nanopore, SQK-LSK114). The libraries were sequenced using a PromethION platform (BGI, Hong Kong).
Project description:Goal: Natural populations of Drosophila melanogaster vary widely in their capacity to resist infection by the parasitic wasp Leptopilina boulardi. To date, little is known about the genetic, cellular and molecular basis underpinning the variation in susceptibility to parasitic wasps. In D.melanogaster populations artificially selected for parasitoid resistance, an increase in the number of circulating hemocytes (blood cells) is observed. One possibility is that this is accompanied by changes in hemocyte activation state, resulting in a successful defence response when exposed to parasitic wasps. We tested this possibility by generating populations that are highly resistant and highly susceptible to the parasitic wasps. We generated these populations after 26 generations of experimental evolution, starting from a single interbred wild caught population. Here, we study how selection for resistance to the parasitic wasp L. boulardi changes the number and type of circulating hemocytes in D. melanogaster larvae using scRNA-seq. Methods: Wild caught D. melanogaster females were collected from Cambridge, UK, to establish an outbreed population. From this, three replicated populations were selected for resistance to L. boulardi strain NSRef for 26 generations (Selection). Another three populations were maintained in the laboratory for 26 generations without being exposed to parasitoid-associated selection pressures (No Selection). At generation 26, second instar D. melanogaster larvae (48-63 hours after fertilization) from each population were infected with L. boulardi for three hours (Infection) or maintained without infection (No infection). 48 hours after, circulating hemocytes from third instar D. melanogaster larvae (96-111 hours after fertilization) from each population were collected in PBS and cleaned in OptiPrep solution (1.09g/ml). 10X Single Cell GEX v2 libraries were prepared and sequenced. CellRanger v2 was used to generate sample cell count matrices. Seurat v3 was used to integrate, normalise and cluster the cell types. Results: We identified novel plasmatocyte cell types, immature and mature lamellocytes and crystal cells. We identify a single lineage of cellular differentiation starting from a self-cycling plasmatocyte population, enriched for genes involved in extracellular matrix organisation, and ending in a mature lamellocyte population that has enriched expression of genes involved in signalling, hemostasis and gluconeogenesis. Immature lamellocytes were overrepresented in populations selected for resistance to parasitoids. Infection with parasitoid resulted in an increase in both immature and mature lamellocytes in circulation. Conclusions: Selection for resistance to the parasitic wasps resulted in the constitutive activation of the D. melanogaster immune system, where plasmatocytes differentiate into immature lamellocytes even where they are not exposed to parasitic wasps. We speculate these constitutive changes assist in mounting a successful defence response when exposed to parasitic wasps.