Concept: Drosophila melanogaster
DEET (N,N-Diethyl-m-toluamide) is one of the most widely used mosquito repellents. Although DEET has been shown to be extremely effective, recent studies have revealed that certain individual insects are unaffected by its presence. A genetic basis for this has been shown in Aedes aegypti mosquitoes and the fruit fly Drosophila melanogaster, but, for the triatomine bug, Rhodnius prolixus, a decrease in response to DEET occurred shortly after previous exposure, indicating that non-genetic factors may also be involved in DEET “insensitivity”. In this study, we examined host-seeking behaviour and electrophysiological responses of A. aegypti after pre-exposure to DEET. We found that three hours after pre-exposure the mosquitoes showed behavioural insensitivity, and electroantennography revealed this correlated with the olfactory receptor neurons responding less to DEET. The change in behaviour as a result of pre-exposure to DEET has implications for the use of repellents and the ability of mosquitoes to overcome them.
Cervical cancer is one of the leading causes of cancer death in women worldwide. The causative agents of cervical cancers, high-risk human papillomaviruses (HPVs), cause cancer through the action of two oncoproteins, E6 and E7. The E6 oncoprotein cooperates with an E3 ubiquitin ligase (UBE3A) to target the p53 tumour suppressor and important polarity and junctional PDZ proteins for proteasomal degradation, activities that are believed to contribute towards malignancy. However, the causative link between degradation of PDZ proteins and E6-mediated malignancy is largely unknown. We have developed an in vivo model of HPV E6-mediated cellular transformation using the genetic model organism, Drosophila melanogaster. Co-expression of E6 and human UBE3A in wing and eye epithelia results in severe morphological abnormalities. Furthermore, E6, via its PDZ-binding motif and in cooperation with UBE3A, targets a suite of PDZ proteins that are conserved in human and Drosophila, including Magi, Dlg and Scribble. Similar to human epithelia, Drosophila Magi is a major degradation target. Magi overexpression rescues the cellular abnormalities caused by E6+UBE3A coexpression and this activity of Magi is PDZ domain-dependent. Drosophila p53 was not targeted by E6+UBE3A, and E6+UBE3A activity alone is not sufficient to induce tumorigenesis, which only occurs when E6+UBE3A are expressed in conjunction with activated/oncogenic forms of Ras or Notch. Finally, through a genetic screen we have identified the insulin receptor signaling pathway as being required for E6+UBE3A induced hyperplasia. Our results suggest a highly conserved mechanism of HPV E6 mediated cellular transformation, and establish a powerful genetic model to identify and understand the cellular mechanisms that underlie HPV E6-induced malignancy.
Members of the Frizzled family of sevenpass transmembrane receptors signal via the canonical Wnt pathway and also via noncanonical pathways of which the best characterized is the planar polarity pathway. Activation of both canonical and planar polarity signaling requires interaction between Frizzled receptors and cytoplasmic proteins of the Dishevelled family; however, there has been some dispute regarding whether the Frizzled-Dishevelled interactions are the same in both cases. Studies looking at mutated forms of Dishevelled suggested that stable recruitment of Dishevelled to membranes by Frizzled was required only for planar polarity activity, implying that qualitatively different Frizzled-Dishevelled interactions underlie canonical signaling. Conversely, studies looking at the sequence requirements of Frizzled receptors in the fruit fly Drosophila melanogaster for canonical and planar polarity signaling have concluded that there is most likely a common mechanism of action. To understand better Frizzled receptor function, we have carried out a large-scale mutagenesis in Drosophila to isolate novel mutations in frizzled that affect planar polarity activity and have identified a group of missense mutations in cytosolic-facing regions of the Frizzled receptor that block Dishevelled recruitment. Interestingly, although some of these affect both planar polarity and canonical activity, as previously reported for similar lesions, we find a subset that affect only planar polarity activity. These results support the view that qualitatively different Frizzled-Dishevelled interactions underlie planar polarity and canonical Wnt signaling.
The dosage compensation complex (DCC) binds to single X chromosomes in Drosophila males and increases the transcription level of X-linked genes by approximately twofold. Male-specific lethal 2 (MSL2) together with MSL1 mediates the initial recruitment of the DCC to high-affinity sites in the X chromosome. MSL2 contains a DNA-binding cysteine-rich CXC domain that is important for X targeting. In this study, we determined the solution structure of MSL2 CXC domain by NMR spectroscopy. We identified three zinc ions in the CXC domain and determined the metal-to-cysteine connectivities from (1)H-(113)Cd correlation experiments. The structure reveals an unusual zinc-cysteine cluster composed of three zinc ions coordinated by six terminal and three bridging cysteines. The CXC domain exhibits unexpected structural homology to pre-SET motifs of histone lysine methyltransferases, expanding the distribution and structural diversity of the CXC domain superfamily. Our findings provide novel structural insight into the evolution and function of CXC domains.
Observation of how cells divide, grow, migrate and form different parts of a developing organism is crucial for understanding developmental programs. Here, we describe a multicolor imaging tool named Raeppli (after the colorful confetti used at the carnival in Basel). Raeppli allows whole-tissue labeling such that the descendants of the majority of cells in a single organ are labeled and can be followed simultaneously relative to one another. We tested the use of Raeppli in the Drosophila melanogaster wing imaginal disc. Induction of Raeppli during larval stages irreversibly labels >90% of the cells with one of four spectrally separable, bright fluorescent proteins with low bias of selection. To understand the global growth characteristics of imaginal discs better, we induced Raeppli at various stages of development, imaged multiple fixed discs at the end of their larval development and estimated the size of their pouch primordium at those developmental stages. We also imaged the same wing disc through the larval cuticle at different stages of its development; the clones marked by Raeppli provide landmarks that can be correlated between multiple time points. Finally, we used Raeppli for continuous live imaging of prepupal eversion of the wing disc.
In all animals, sleep pressure is under continuous tight regulation. It is universally accepted that this regulation arises from a two-process model, integrating both a circadian and a homeostatic controller. Here we explore the role of environmental social signals as a third, parallel controller of sleep homeostasis and sleep pressure. We show that, in Drosophila melanogaster males, sleep pressure after sleep deprivation can be counteracted by raising their sexual arousal, either by engaging the flies with prolonged courtship activity or merely by exposing them to female pheromones.
To escape danger or catch prey, running vertebrates rely on dynamic gaits with minimal ground contact. By contrast, most insects use a tripod gait that maintains at least three legs on the ground at any given time. One prevailing hypothesis for this difference in fast locomotor strategies is that tripod locomotion allows insects to rapidly navigate three-dimensional terrain. To test this, we computationally discovered fast locomotor gaits for a model based on Drosophila melanogaster. Indeed, the tripod gait emerges to the exclusion of many other possible gaits when optimizing fast upward climbing with leg adhesion. By contrast, novel two-legged bipod gaits are fastest on flat terrain without adhesion in the model and in a hexapod robot. Intriguingly, when adhesive leg structures in real Drosophila are covered, animals exhibit atypical bipod-like leg coordination. We propose that the requirement to climb vertical terrain may drive the prevalence of the tripod gait over faster alternative gaits with minimal ground contact.
A functioning gene drive system could fundamentally change our strategies for the control of vector-borne diseases by facilitating rapid dissemination of transgenes that prevent pathogen transmission or reduce vector capacity. CRISPR/Cas9 gene drive promises such a mechanism, which works by converting cells that are heterozygous for the drive construct into homozygotes, thereby enabling super-Mendelian inheritance. Although CRISPR gene drive activity has already been demonstrated, a key obstacle for current systems is their propensity to generate resistance alleles, which cannot be converted to drive alleles. In this study, we developed two CRISPR gene drive constructs based on the nanos and vasa promoters that allowed us to illuminate the different mechanisms by which resistance alleles are formed in the model organism Drosophila melanogaster. We observed resistance allele formation at high rates both prior to fertilization in the germline and post-fertilization in the embryo due to maternally deposited Cas9. Assessment of drive activity in genetically diverse backgrounds further revealed substantial differences in conversion efficiency and resistance rates. Our results demonstrate that the evolution of resistance will likely impose a severe limitation to the effectiveness of current CRISPR gene drive approaches, especially when applied to diverse natural populations.
The “organic food” market is the fastest growing food sector, yet it is unclear whether organically raised food is nutritionally superior to conventionally grown food and whether consuming organic food bestows health benefits. In order to evaluate potential health benefits of organic foods, we used the well-characterized fruit fly Drosophila melanogaster as a model system. Fruit flies were raised on a diets consisting of extracts of either conventionally or organically raised produce (bananas, potatoes, raisins, soy beans). Flies were then subjected to a variety of tests designed to assess overall fly health. Flies raised on diets made from organically grown produce had greater fertility and longevity. On certain food sources, greater activity and greater stress resistance was additionally observed, suggesting that organic food bestows positive effects on fly health. Our data show that Drosophila can be used as a convenient model system to experimentally test potential health effects of dietary components. Using this system, we provide evidence that organically raised food may provide animals with tangible benefits to overall health.
Environmental adaptation is one of the most fundamental features of organisms. Modern genome science has identified some genes associated with adaptive traits of organisms, and has provided insights into environmental adaptation and evolution. However, how genes contribute to adaptive traits and how traits are selected under an environment in the course of evolution remain mostly unclear. To approach these issues, we utilize “Dark-fly”, a Drosophila melanogaster line maintained in a constant dark condition for more than 60 years. Our previous analysis identified 220,000 single nucleotide polymorphisms (SNPs) in the Dark-fly genome, but did not clarify which SNPs of Dark-fly are truly adaptive for living in the dark. We found here that Dark-fly dominated over the wild-type fly in a mixed population under dark conditions, and based on this domination we designed an experiment for genome re-selection to identify adaptive genes of Dark-fly. For this experiment, large mixed populations of Dark-fly and the wild-type fly were maintained in light conditions or in dark conditions, and the frequencies of Dark-fly SNPs were compared between these populations across the whole genome. We thereby detected condition-dependent selections towards approximately 6% of the genome. In addition, we observed the time-course trajectory of SNP frequency in the mixed populations through generations 0, 22, and 49, which resulted in notable categorization of the selected SNPs into three types with different combinations of positive and negative selections. Our data provided a list of about 100 strong candidate genes associated with the adaptive traits of Dark-fly.