Jeremy A. Lynch

Localized maternal orthodenticle patterns anterior and posterior in the long germ wasp Nasonia

The Bicoid (Bcd) gradient in Drosophila has long been a model for the action of a morphogen in establishing embryonic polarity. However, it is now clear that bcd is a unique feature of higher Diptera. An evolutionarily ancient gene, orthodenticle (otd), has a bcd-like role in the beetle Tribolium. Unlike the Bcd gradient, which arises by diffusion of protein from an anteriorly localized messenger RNA, the Tribolium Otd gradient forms by translational repression of otd mRNA by a posteriorly localized factor. These differences in gradient formation are correlated with differences in modes of embryonic patterning. Drosophila uses long germ embryogenesis, where the embryo derives from the entire anterior–posterior axis, and all segments are patterned at the blastoderm stage, before gastrulation. In contrast, Tribolium undergoes short germ embryogenesis: the embryo arises from cells in the posterior of the egg, and only anterior segments are patterned at the blastoderm stage, with the remaining segments arising after gastrulation from a growth zone. Here we describe the role of otd in the long germband embryo of the wasp Nasonia vitripennis. We show that Nasonia otd maternal mRNA is localized at both poles of the embryo, and resulting protein gradients pattern both poles. Thus, localized Nasonia otd has two major roles that allow long germ development. It activates anterior targets at the anterior of the egg in a manner reminiscent of the Bcd gradient, and it is required for pre-gastrulation expression of posterior gap genes.

A method for parental RNA interference in the wasp Nasonia vitripennis.

The wasp Nasonia vitripennis is emerging as a useful model organism in which to address a variety of biological questions, due, in part, to its ease of laboratory use, unique aspects of its biology and the sequencing of its genome. In order to take full advantage of the potential of this organism, methods for manipulating gene function are needed. To this end, a protocol for parental RNA interference (pRNAi) in N. vitripennis is described. pRNAi entails injecting pupae with double-stranded RNA, allowing the injected wasps to eclose and examining the progeny for developmental defects. This basic protocol is described in the context of the life cycle of N. vitripennis. This technique has been useful in elucidating the function of most, although not all, genes tested to date, and has potential applications beyond embryonic patterning. pRNAi experiments in Nasonia can be completed in as little as 2 weeks.

Symmetry Breaking During Drosophila Oogenesis

The orthogonal axes of Drosophila are established during oogenesis through a hierarchical series of symmetry-breaking steps, most of which can be traced back to asymmetries inherent in the architecture of the ovary. Oogenesis begins with the formation of a germline cyst of 16 cells connected by ring canals. Two of these 16 cells have four ring canals, whereas the others have fewer. The first symmetry-breaking step is the selection of one of these two cells to become the oocyte. Subsequently, the germline cyst becomes surrounded by somatic follicle cells to generate individual egg chambers. The second symmetry-breaking step is the posterior positioning of the oocyte within the egg chamber, a process mediated by adhesive interactions with a special group of somatic cells. Posterior oocyte positioning is accompanied by a par gene-dependent repolarization of the microtubule network, which establishes the posterior cortex of the oocyte. The next two steps of symmetry breaking occur during midoogenesis after the volume of the oocyte has increased about 10-fold. First, a signal from the oocyte specifies posterior follicle cells, polarizing a symmetric prepattern present within the follicular epithelium. Second, the posterior follicle cells send a signal back to the oocyte, which leads to a second repolarization of the oocyte microtubule network and the asymmetric migration of the oocyte nucleus. This process again requires the par genes. The repolarization of the microtubule network results in the transport of bicoid and oskar mRNAs, the anterior and posterior determinants, respectively, of the embryonic axis, to opposite poles of the oocyte. The asymmetric positioning of the oocyte nucleus defines a cortical region of the oocyte where gurken mRNA is localized, thus breaking the dorsal-ventral symmetry of the egg and embryo.

EGF signaling and the origin of axial polarity among the insects.

The eggs of insects are unusual in that they often have bilateral symmetry when they are laid, indicating that both anterior-posterior (AP) and dorsal-ventral (DV) symmetries are broken during oogenesis. The molecular basis of this process is well understood in Drosophila melanogaster, in which symmetry breaking events for both axes depend on the asymmetric position of the oocyte nucleus and on germline-soma signaling mediated by the Tgf alpha-like epidermal growth factor (EGF) ligand Gurken. Germline-soma signaling interactions centered around the oocyte nucleus have been proposed in other insect species, but the molecular nature of these interactions has not been elucidated. We have examined the behavior of the oocyte nucleus and the function of EGF signaling components in the ovaries of the wasp Nasonia vitripennis, the beetle Tribolium castaneum, and the cricket Gryllus bimaculatus. We have found that EGF signaling has broadly conserved roles in mediating the encapsulation of oocytes by the somatic follicle cell layer, in establishing polarity of the egg chambers, and in setting up the DV axis of the embryo. These results provide insights into the evolutionary origins of the unique strategy employed by insects to establish embryonic axial polarity during oogenesis.

The evolution of dorsal–ventral patterning mechanisms in insects

The gene regulatory network (GRN) underpinning dorsal-ventral (DV) patterning of the Drosophila embryo is among the most thoroughly understood GRNs, making it an ideal system for comparative studies seeking to understand the evolution of development. With the emergence of widely applicable techniques for testing gene function, species with sequenced genomes, and multiple tractable species with diverse developmental modes, a phylogenetically broad and molecularly deep understanding of the evolution of DV axis formation in insects is feasible. Here, we review recent progress made in this field, compare our emerging molecular understanding to classical embryological experiments, and suggest future directions of inquiry.

The phylogenetic origin of oskar coincided with the origin of maternally provisioned germ plasm and pole cells at the base of the holometabola

The establishment of the germline is a critical, yet surprisingly evolutionarily labile, event in the development of sexually reproducing animals. In the fly Drosophila, germ cells acquire their fate early during development through the inheritance of the germ plasm, a specialized maternal cytoplasm localized at the posterior pole of the oocyte. The gene oskar (osk) is both necessary and sufficient for assembling this substance. Both maternal germ plasm and oskar are evolutionary novelties within the insects, as the germline is specified by zygotic induction in basally branching insects, and osk has until now only been detected in dipterans. In order to understand the origin of these evolutionary novelties, we used comparative genomics, parental RNAi, and gene expression analyses in multiple insect species. We have found that the origin of osk and its role in specifying the germline coincided with the innovation of maternal germ plasm and pole cells at the base of the holometabolous insects and that losses of osk are correlated with changes in germline determination strategies within the Holometabola. Our results indicate that the invention of the novel gene osk was a key innovation that allowed the transition from the ancestral late zygotic mode of germline induction to a maternally controlled establishment of the germline found in many holometabolous insect species. We propose that the ancestral role of osk was to connect an upstream network ancestrally involved in mRNA localization and translational control to a downstream regulatory network ancestrally involved in executing the germ cell program.

A major role for zygotic hunchback in patterning the Nasonia embryo

Developmental genetic analysis has shown that embryos of the parasitoid wasp Nasonia vitripennis depend more on zygotic gene products to direct axial patterning than do Drosophila embryos. In Drosophila, anterior axial patterning is largely established by bicoid, a rapidly evolving maternal-effect gene, working with hunchback, which is expressed both maternally and zygotically. Here, we focus on a comparative analysis of Nasonia hunchback function and expression. We find that a lesion in Nasonia hunchback is responsible for the severe zygotic headless mutant phenotype, in which most head structures and the thorax are deleted, as are the three most posterior abdominal segments. This defines a major role for zygotic Nasonia hunchback in anterior patterning, more extensive than the functions described for hunchback in Drosophila or Tribolium. Despite the major zygotic role of Nasonia hunchback, we find that it is strongly expressed maternally, as well as zygotically. Nasonia Hunchback embryonic expression appears to be generally conserved; however, the mRNA expression differs from that of Drosophila hunchback in the early blastoderm. We also find that the maternal hunchback message decays at an earlier developmental stage in Nasonia than in Drosophila, which could reduce the relative influence of maternal products in Nasonia embryos. Finally, we extend the comparisons of Nasonia and Drosophila hunchback mutant phenotypes, and propose that the more severe Nasonia hunchback mutant phenotype may be a consequence of differences in functionally overlapping regulatory circuitry.

Genome of the Asian longhorned beetle (Anoplophora glabripennis), a globally significant invasive species, reveals key functional and evolutionary innovations at the beetle-plant interface

BackgroundRelatively little is known about the genomic basis and evolution of wood-feeding in beetles. We undertook genome sequencing and annotation, gene expression assays, studies of plant cell wall degrading enzymes, and other functional and comparative studies of the Asian longhorned beetle, Anoplophora glabripennis, a globally significant invasive species capable of inflicting severe feeding damage on many important tree species. Complementary studies of genes encoding enzymes involved in digestion of woody plant tissues or detoxification of plant allelochemicals were undertaken with the genomes of 14 additional insects, including the newly sequenced emerald ash borer and bull-headed dung beetle.ResultsThe Asian longhorned beetle genome encodes a uniquely diverse arsenal of enzymes that can degrade the main polysaccharide networks in plant cell walls, detoxify plant allelochemicals, and otherwise facilitate feeding on woody plants. It has the metabolic plasticity needed to feed on diverse plant species, contributing to its highly invasive nature. Large expansions of chemosensory genes involved in the reception of pheromones and plant kairomones are consistent with the complexity of chemical cues it uses to find host plants and mates.ConclusionsAmplification and functional divergence of genes associated with specialized feeding on plants, including genes originally obtained via horizontal gene transfer from fungi and bacteria, contributed to the addition, expansion, and enhancement of the metabolic repertoire of the Asian longhorned beetle, certain other phytophagous beetles, and to a lesser degree, other phytophagous insects. Our results thus begin to establish a genomic basis for the evolutionary success of beetles on plants.

Comparisons of the embryonic development of Drosophila, Nasonia, and Tribolium.

Studying the embryogenesis of diverse insect species is crucial to understanding insect evolution. Here, we review current advances in understanding the development of two emerging model organisms: the wasp Nasonia vitripennis and the beetle Tribolium castaneum in comparison with the well‐studied fruit fly Drosophila melanogaster. Although Nasonia represents the most basally branching order of holometabolous insects, it employs a derived long germband mode of embryogenesis, more like that of Drosophila, whereas Tribolium undergoes an intermediate germband mode of embryogenesis, which is more similar to the ancestral mechanism. Comparing the embryonic development and genetic regulation of early patterning events in these three insects has given invaluable insights into insect evolution. The similar mode of embryogenesis of Drosophila and Nasonia is reflected in their reliance on maternal morphogenetic gradients. However, they employ different genes as maternal factors, reflecting the evolutionary distance separating them. Tribolium, on the other hand, relies heavily on self‐regulatory mechanisms other than maternal cues, reflecting its sequential nature of segmentation and the need for reiterated patterning. WIREs Dev Biol 2012, 1:16–39. doi: 10.1002/wdev.3

Heads and tails: evolution of antero-posterior patterning in insects.

In spite of their varied appearances, insects share a common body plan whose layout is established by patterning genes during embryogenesis. We understand in great molecular detail how the Drosophila embryo patterns its segments. However, Drosophila has a type of embryogenesis that is highly derived and varies extensively as compared to most insects. Therefore, the study of other insects is invaluable for piecing together how the ancestor of all insects established its segmented body plan, and how this process can be plastic during evolution. In this review, we discuss the evolution of Antero-Posterior (A-P) patterning mechanisms in insects. We first describe two distinct modes of insect development - long and short germ development - and how these two modes of patterning are achieved. We then summarize how A-P patterning occurs in the long-germ Drosophila, where most of our knowledge comes from, and in the well-studied short-germ insect, Tribolium. Finally, using examples from other insects, we highlight differences in patterns of expression, which suggest foci of evolutionary change.