Lijie Sun

Genome-wide analysis of phenobarbital-inducible genes in Drosophila melanogaster

An oligoarray analysis was conducted to determine the differential expression of genes due to phenobarbital exposure in Drosophila melanogaster (w1118 strain) third instar larvae. Seventeen genes were observed to be induced with increased expression by a statistical analysis of microarrays approach with a q ≤ 0.05. At q ≤ 0.12, four more genes (Cyp12d1, DmGstd4, and two genes with unknown function) were found to be up‐regulated, and 11 genes with unknown function were found to be down‐regulated. Fifteen of these genes, Cyp4d14, Cyp6a2, Cyp6a8, Cyp12d1, Cyp6d5, Cyp6w1, CG2065, DmGstd6, DmGstd7, Amy‐p/Amy‐d, Ugt86Dd, GC5724, Jheh1, Jheh2 and CG11893, were verified using quantitative real time polymerase chain reaction. Some of these genes have been shown to be over‐transcribed in metabolically DDT‐resistant Drosophila strains.

Transcriptional signatures in response to wheat germ agglutinin and starvation in Drosophila melanogaster larval midgut.

One function of plant lectins such as wheat germ agglutinin is to serve as defences against herbivorous insects. The midgut is one critical site affected by dietary lectins. We observed marked cellular, structural and gene expression changes in the midguts of Drosophila melanogaster third instar larvae that were fed wheat germ agglutinin. Some of these changes were similar to those observed in the midguts of starved D. melanogaster. Dietary wheat germ agglutinin caused shortening, branching, swelling, distortion and in some cases disintegration of the midgut microvilli. Starvation was accompanied primarily by shortening of the microvilli. Microarray analyses revealed that dietary wheat germ agglutinin evoked differential expression of 61 transcripts; seven of these were also differentially expressed in starved D. melanogaster. The differentially transcribed gene clusters in wheat germ agglutinin‐fed larvae were associated with (1) cytoskeleton organization; (2) digestive enzymes; (3) detoxification reactions; and (4) energy metabolism. Four possible transcription factor binding motifs were associated with the differentially expressed genes. One of these exhibited substantial similarity to MyoD, a transcription factor binding motif associated with cellular structures in mammals. These results are consistent with the hypothesis that wheat germ agglutinin caused a starvation‐like effect and structural changes of midgut cells of D. melanogaster third‐instar larvae.

Systems-Scale Analysis Reveals Pathways Involved in Cellular Response to Methamphetamine.

Background Methamphetamine (METH), an abused illicit drug, disrupts many cellular processes, including energy metabolism, spermatogenesis, and maintenance of oxidative status. However, many components of the molecular underpinnings of METH toxicity have yet to be established. Network analyses of integrated proteomic, transcriptomic and metabolomic data are particularly well suited for identifying cellular responses to toxins, such as METH, which might otherwise be obscured by the numerous and dynamic changes that are induced. Methodology/Results We used network analyses of proteomic and transcriptomic data to evaluate pathways in Drosophila melanogaster that are affected by acute METH toxicity. METH exposure caused changes in the expression of genes involved with energy metabolism, suggesting a Warburg-like effect (aerobic glycolysis), which is normally associated with cancerous cells. Therefore, we tested the hypothesis that carbohydrate metabolism plays an important role in METH toxicity. In agreement with our hypothesis, we observed that increased dietary sugars partially alleviated the toxic effects of METH. Our systems analysis also showed that METH impacted genes and proteins known to be associated with muscular homeostasis/contraction, maintenance of oxidative status, oxidative phosphorylation, spermatogenesis, iron and calcium homeostasis. Our results also provide numerous candidate genes for the METH-induced dysfunction of spermatogenesis, which have not been previously characterized at the molecular level. Conclusion Our results support our overall hypothesis that METH causes a toxic syndrome that is characterized by the altered carbohydrate metabolism, dysregulation of calcium and iron homeostasis, increased oxidative stress, and disruption of mitochondrial functions.

Bowman-Birk inhibitor affects pathways associated with energy metabolism in Drosophila melanogaster

Bowman‐Birk inhibitor (BBI) is toxic when fed to certain insects, including the fruit fly, Drosophila melanogaster. Dietary BBI has been demonstrated to slow growth and increase insect mortality by inhibiting the digestive enzymes trypsin and chymotrypsin, resulting in a reduced supply of amino acids. In mammals, BBI influences cellular energy metabolism. Therefore, we tested the hypothesis that dietary BBI affects energy‐associated pathways in the D. melanogaster midgut. Through microarray and metabolomic analyses, we show that dietary BBI affects energy utilization pathways in the midgut cells of D. melanogaster. In addition, ultrastructure studies indicate that microvilli are significantly shortened in BBI‐fed larvae. These data provide further insights into the complex cellular response of insects to dietary protease inhibitors.

Negative Cross-Resistance: Past, Present, and Future Potential

This chapter explores the current status of negative cross-resistance (NCR) in the peer-reviewed literature, examines discovery strategies in more detail, discusses how to deploy the resulting NCR compounds, and addresses the potential limitations and possible future opportunities for such an approach in resistance management. NCR has been observed across a variety of species and chemical classes. However, to date, it has not typically been used in wide-scale insect resistance management (IRM). The lack of forthcoming NCR products may be due to logical business models that necessitate the cost effective development of new products due to the needs of the marketplace. In this regard, NCR products will likely only be developed in response to verified resistance to currently marketed high-value products. An additional reason for the lack of NCR compounds may have been the practical limitations in the methodologies needed to efficiently discover them. Development of NCR compounds can be achieved through a variety of methods, for example, use of large-scale screening processes modified from those currently used for screening for novel pesticides. Such screening approaches could involve field resistant insects, or in some specific cases transgenic D. melanogaster expressing the resistance trait. Additionally, advances in molecular cloning and expression of peptides in display technologies could allow for the rapid development of NCR products as soon as resistance occurs in insect populations in the field. Rational design of traditional chemistries as well as proteins is also well established. Thus, field-resistant insects, high throughput transgenic live insect systems, phage display technologies, and rational design approaches, or any combination of these, could be used to assess a wide array of receptor/toxin combinations to model a best fit for NCR toxins useful in the field.

Negative Cross-Resistance

During the 20th century, pesticide use has become integral for current agricultural practices, as has the challenge associated with pesticide resistance. Traditional resistance management plans have often used a “use and discard” approach, changing the chemical to target a different mode of action in the pest species once resistance becomes a problem in the field. An alternative strategy is to identify compounds that confer negative cross-resistance (NCR), where the NCR compound is more toxic to pesticide resistant insects as compared to their pesticide susceptible counterparts. Examples of NCR exist in the literature, however, a systematic approach to discover and use these compounds has been lacking in industrial agriculture. In the following chapter we explore both the limitations and the potential for use of NCR strategies in relation to resistance management.

Understanding Resistance and Induced Responses of Insects to Xenobiotics and Insecticides in the Age of “Omics” and Systems Biology

We tend to think of the word resistance in terms of evolutionary changes in an insect population that occur in response to repetitive exposures to pesticides or other xenobiotics used to manage insect pests in crops, homes, and gardens, or on livestock or humans (including disease vectors). Resistance can also be defined in broader terms since insects are “resistant” to many naturally occurring abiotic and biotic factors they encounter in their environment. In this chapter, we will outline the concepts associated with pesticide resistance, as well as provide examples of some of the known mechanisms associated with resistance. In addition, we will also discuss the broader context of how we can use emergent “omics” tools, such as genomics, proteomics, and metabolomics, to better understand and discover resistance mechanisms. As RNAi is an emerging potential approach for insect control, we also discuss the potential for resistance in insect populations to RNAi pest control strategies. Finally, we will discuss how we can use this information to develop strategies that minimize the impact of insects on human health, food, and property.

Chapter 11 – Negative Cross-Resistance: History, Present Status, and Emerging Opportunities

During the 20th century, pesticide use has become integral for current agricultural practices, as has the challenge associated with pesticide resistance. Traditional resistance management plans have often used a “use and discard” approach, changing the chemical to target a different mode of action in the pest species once resistance becomes a problem in the field. An alternative strategy is to identify compounds that confer negative cross-resistance (NCR), where the NCR compound is more toxic to pesticide resistant insects as compared to their pesticide susceptible counterparts. Examples of NCR exist in the literature, however, a systematic approach to discover and use these compounds has been lacking in industrial agriculture. In the following chapter we explore both the limitations and the potential for use of NCR strategies in relation to resistance management.

Negative Cross-Resistance. History, Present Status, and Emerging Opportunities.

During the 20th century, pesticide use has become integral for current agricultural practices, as has the challenge associated with pesticide resistance. Traditional resistance management plans have often used a “use and discard” approach, changing the chemical to target a different mode of action in the pest species once resistance becomes a problem in the field. An alternative strategy is to identify compounds that confer negative cross-resistance (NCR), where the NCR compound is more toxic to pesticide resistant insects as compared to their pesticide susceptible counterparts. Examples of NCR exist in the literature, however, a systematic approach to discover and use these compounds has been lacking in industrial agriculture. In the following chapter we explore both the limitations and the potential for use of NCR strategies in relation to resistance management.