Chemical genetics and drug screening in Drosophila cancer models

Drug candidates often fail in preclinical and clinical testing because of reasons of efficacy and/or safety.It would be time- and cost-efficient to have screening models that reduce the rate of such false positive candidates that appear promising at first but fail later.In this regard,it would be particularly useful to have a rapid and inexpensive whole animal model that can pre-select hits from high-throughput screens but before testing in costly rodent assays.Drosophila melanogaster has emerged as a potential whole animal model for drug screening.Of particular interest have been drugs that must act in the context of multi-cellularity such as those for neurological disorders and cancer.A recent review provides a comprehensive summary of drug screening in Drosophila,but with an emphasis on neurodegenerative disorders.Here,we review Drosophila screens in the literature aimed at cancer therapeutics.     

Chemical genetics and drug screening in Drosophila cancer models

Drug candidates often fail in preclinical and clinical testing because of reasons of efficacy and/or safety.It would be time- and cost-efficient to have screening models that reduce the rate of such false positive candidates that appear promising at first but fail later.In this regard,it would be particularly useful to have a rapid and inexpensive whole animal model that can pre-select hits from high-throughput screens but before testing in costly rodent assays.Drosophila melanogaster has emerged as a potential whole animal model for drug screening.Of particular interest have been drugs that must act in the context of multi-cellularity such as those for neurological disorders and cancer.A recent review provides a comprehensive summary of drug screening in Drosophila,but with an emphasis on neurodegenerative disorders.Here,we review Drosophila screens in the literature aimed at cancer therapeutics.     

Chemical Selection of Alcohol Dehydrogenase Negative Mutants in Drosophila

We describe a selection procedure which utilizes the vapor from an unsaturated alcohol, 1-pentene-3-ol, for the detection and isolation of mutant flies with little or no alcohol dehydrogenase activity. ADH-negative flies are unaffected by exposure to the unsaturated alcohol, but ADH positives (wild-types) die after short exposure. The technique can be used to select rare ADH-negative individuals from large populations of wild-type flies.     

Direct sensing of systemic and nutritional signals by haematopoietic progenitors in Drosophila

The Drosophila lymph gland is a haematopoietic organ in which progenitor cells, which are most akin to the common myeloid progenitor in mammals, proliferate and differentiate into three types of mature cell--plasmatocytes, crystal cells and lamellocytes--the functions of which are reminiscent of mammalian myeloid cells. During the first and early second instars of larval development, the lymph gland contains only progenitors, whereas in the third instar, a medial region of the primary lobe of the lymph gland called the medullary zone contains these progenitors, and maturing blood cells are found juxtaposed in a peripheral region designated the cortical zone. A third group of cells referred to as the posterior signalling centre functions as a haematopoietic niche. Similarly to mammalian myeloid cells, Drosophila blood cells respond to multiple stresses including hypoxia, infection and oxidative stress. However, how systemic signals are sensed by myeloid progenitors to regulate cell-fate determination has not been well described. Here, we show that the haematopoietic progenitors of Drosophila are direct targets of systemic (insulin) and nutritional (essential amino acid) signals, and that these systemic signals maintain the progenitors by promoting Wingless (WNT in mammals) signalling. We expect that this study will promote investigation of such possible direct signal sensing mechanisms by mammalian myeloid progenitors.     

Chemical cues influence pupation behavior of Drosophila simulans and Drosophila buzzatii in nature and in the laboratory

In the wild, larvae of several species of Drosophila develop in heterogeneous and rapidly changing environments sharing resources as food and space. In this scenario, sensory systems contribute to detect, localize and recognize congeners and heterospecifics, and provide information about the availability of food and chemical features of environments where animals live. We investigated the behavior of D. simulans and D. buzzatii larvae to chemicals emitted by conspecific and heterospecific larvae. Our goal was to understand the role of these substances in the selection of pupation sites in the two species that cohabit within decaying prickly pear fruits (Opuntia ficus-indica). In these breeding sites, larvae of D. simulans and D. buzzatii detect larvae of the other species changing their pupation site preferences. Larvae of the two species pupated in the part of the fruit containing no or few heterospecifics, and spent a longer time in/on spots marked by conspecifics rather than heterospecifics. In contrast, larvae of the two species reared in isolation from conspecifics pupated randomly over the substrate and spent a similar amount of time on spots marked by conspecifics and by heterospecifics. Our results indicate that early chemically-based experience with conspecific larvae is critical for the selection of the pupation sites in D. simulans and D. buzzatii, and that pupation site preferences of Drosophila larvae depend on species-specific chemical cues. These preferences can be modulate by the presence of larvae of the same or another species.     

Gonadal Mosaicism Induced by Chemical Treatment of Sperm in Drosophila melanogaster

Accurate interpretation of forward genetic screens of chromosomes exposed in mature spermatozoa to a mutagenic chemical requires understanding—incomplete to date—of how exposed chromosomes and their replicas proceed through early development stages from the fertilized ovum to establishment of the germline of the treated male’s offspring. We describe a model for early embryonic development and establishment of the germline of Drosophila melanogaster and a model-validating experiment. Our model proposes that, barring repair, DNA strands modified by treatment with alkylating agents are stable and mutagenic. Each replication of an alkylated strand can result in misreplication and a mutant-bearing daughter nucleus. Daughter nuclei thenceforth replicate faithfully and their descendants comprise the embryonic syncytium. Of the 256 nuclei present after the eighth division, several migrate into the polar plasm at the posterior end of the embryo to found the germline. Based upon distribution of descendants of the alkylated strands, the misreplication rate, and the number of nuclei selected as germline progenitors, the frequency of gonadal mosaicism is predictable. Experimentally, we tracked chromosomes 2 and 3 from EMS-treated sperm through a number of generations, to characterize autosomal recessive lethal mutations and infer gonadal genetic content of the sons of treated males. Over 50% of 106 sons bore germlines that were singly, doubly, or triply mosaic for chromosome 2 or chromosome 3. These findings were consistent with our model, assuming a rate of misreplication between 0.65 and 0.80 at each replication of an alkylated strand. Crossing treated males to mismatch-repair-deficient females had no apparent effect on mutation rate.     

Chemical Patterns, Compartments and a Binary Epigenetic Code in Drosophila

I propose a model which accounts for the geometries and sequencein which compartmental boundary lines arise on the differentimaginal discs, and on the blastoderm of Drosophila melanogaster;and propose that successive lines are recorded by differentbinary switches, to create a binary epigenetic code word specifyingeach disc, and disc compartment. I suppose a biochemical systemundergoing reaction and diffusion acts throughout development.As an imaginal disc grows, a succession of differently shapedchemical concentration patterns form at a discrete set of discsizes. I suppose a specific concentration of one chemical isa threshold. Concentrations above or below threshold switchcells to one or another of two commitments. Then the line acrossthe imaginal disc with the threshold concentration is a predictedcompartmental boundary. The sequence and geometries of suchlines predict the compartmental boundaries seen on the wingdisc, the other discs, and on the blastoderm stage egg. Thecompartmental lines on the wing disc suggest that a terminalcompartment is specified by a combination of binary names recordinga sequence of binary commitments: anterior, not posterior; dorsal,not ventral; wing, not thorax; proximal, not distal. Each combinationcomprises a binary epigenetic code word. Recently I constructedan independent model for transdetermination in Drosophila whichproposed a similar binary epigenetic code for the differentdiscs. The clone restriction lines predicted on the blastodermby my transdetermination model, the chemical pattern model,and analogy with the wing disc, are nearly identical. Severalare already confirmed. The resultant binary code scheme correctlypredicts many relative transdetermination frequencies and accountssimply for the action of most homeotic mutants as genes whichalter a single switch state in one or more discs.     

Chemical modification of cell death in the Bar eye of Drosophila

Summary Cell death during the development of the mutant Bar of Drosophila melanogaster has been further investigated. The temporal and spatial distribution of degenerating cells was determined by light microscopy of serial sections of Bar eye discs. It appears that the presumptive eye cells die during the initial stage of differentiation; i.e., during their organization into groups of cells corresponding to the future ommatidia.The effects of acetamide and cytosine treatment on the development of the Bar eye disc were also examined. Both of these compounds, when fed to larvae, greatly increase facet number in the adult fly. Acetamide increases facet number by inhibiting Bar specific cell death, while cytosine delays degeneration to a later stage in development when ommatidial organization is almost complete.     

Modeling metabolic homeostasis and nutrient sensing in Drosophila: implications for aging and metabolic diseases

Over the past decade, numerous reports have underscored the similarities between the metabolism of Drosophila and vertebrates, with the identification of evolutionarily conserved enzymes and analogous organs that regulate carbohydrate and lipid metabolism. It is now well established that the major metabolic, energy-sensing and endocrine signaling networks of vertebrate systems are also conserved in flies. Accordingly, studies in Drosophila are beginning to unravel how perturbed energy balance impinges on lifespan and on the ensuing diseases when energy homeostasis goes awry. Here, we highlight several emerging concepts that are at the nexus between obesity, nutrient sensing, metabolic homeostasis and aging. Specifically, we summarize the endocrine mechanisms that regulate carbohydrate and lipid metabolism, and provide an overview of the neuropeptides that regulate feeding behavior. We further describe the various efforts at modeling the effects of high-fat or -sugar diets in Drosophila and the signaling mechanisms involved in integrating organ function. Finally, we draw attention to some of the cardinal discoveries made with these disease models and how these could spur new research questions in vertebrate systems.KEY WORDS: Metabolic homeostasis, Nutrient sensing, Drosophila     

Dual detection of fungal infections in Drosophila via recognition of glucans and sensing of virulence factors

The Drosophila immune system discriminates between various types of infections and activates appropriate signal transduction pathways to combat the invading microorganisms. The Toll pathway is required for the host response against fungal and most Gram-positive bacterial infections. The sensing of Gram-positive bacteria is mediated by the pattern recognition receptors PGRP-SA and GNBP1 that cooperate to detect the presence of infections in the host. Here, we report that GNBP3 is a pattern recognition receptor that is required for the detection of fungal cell wall components. Strikingly, we find that there is a second, parallel pathway acting jointly with GNBP3. The Drosophila Persephone protease activates the Toll pathway when proteolytically matured by the secreted fungal virulence factor PR1. Thus, the detection of fungal infections in Drosophila relies both on the recognition of invariant microbial patterns and on monitoring the effects of virulence factors on the host.     

Lactobacillus plantarum promotes Drosophila systemic growth by modulating hormonal signals through TOR-dependent nutrient sensing

There is growing evidence that intestinal bacteria are important beneficial partners of their metazoan hosts. Recent observations suggest a strong link between commensal bacteria, host energy metabolism, and metabolic diseases such as diabetes and obesity. As a consequence, the gut microbiota is now considered a "host" factor that influences energy uptake. However, the impact of intestinal bacteria on other systemic physiological parameters still remains unclear. Here, we demonstrate that Drosophila microbiota promotes larval growth upon nutrient scarcity. We reveal that Lactobacillus plantarum, a commensal bacterium of the Drosophila intestine, is sufficient on its own to recapitulate the?natural microbiota growth-promoting effect. L.?plantarum exerts its benefit by acting genetically upstream of the TOR-dependent host nutrient sensing system controlling hormonal growth signaling. Our results indicate that the intestinal microbiota should also be envisaged as a factor that influences the systemic growth of its host.     

The Drosophila systemic immune response: sensing and signalling during bacterial and fungal infections

A hallmark of the potent, multifaceted antimicrobial defence of Drosophila melanogaster is the challenge-induced synthesis of several families of antimicrobial peptides by cells in the fat body. The basic mechanisms of recognition of various types of microbial infections by the adult fly are now understood, often in great detail. We have further gained valuable insight into the infection-induced gene reprogramming by nuclear factor-kappaB (NF-kappaB) family members under the dependence of complex intracellular signalling cascades. The striking parallels between the adult fly response and mammalian innate immune defences described below point to a common ancestry and validate the relevance of the fly defence as a paradigm for innate immunity.     

Chemical sensing by a novel electrical oscillator: detection and quantitation of polysaccharides in concanavalin A solutions

Electrical oscillations across two platinum electrodes connected to an external circuit and immersed in HEPES buffer solutions of concanavalin A (Con A) were measured at various concentrations of mannan, starch and dextran. The frequency of oscillations was found to change almost linearly with the logarithm of the concentration of mannan between 10(-4) and 1 w/v%; whereas the frequency remained nearly constant with dextran. With starch it slightly increased as the concentration increased. This method was suggested to be useful for quantitative and selective measurement of antigen-antibody reaction and also for detection of various cells with the specific binding site such as cancer cells.