Drosophila Gene Expression During Metamorphosis in Wild Type and Germline Minus Pupae
ABSTRACT: Drosophila melanogaster undergoes a complete metamorphosis, during which time the larval male and female forms transition into sexually dimorphic, reproductive adult forms. To understand this complex morphogenetic process at a molecular-genetic level, we performed whole genome microarray analyses. Genes were identified that were expressed during metamorphosis in both somatic and germline tissues of males and females. Additionally, genes were identified that display sex-specific differences in abundance in both of these tissues at discrete times during metamorphosis. Keywords: time course; wild type; genetic modification; Gene expression was examined at five time points during metamorphosis: 0, 24, 48, 71, and 96 hr After Puparium Formation (APF). Gene expression was examined separately in males and females for both wild type pupae and tudor (tud) progeny. tud progeny have genetically ablated germline tissues. All samples were labeled with Cy5 and compared against a common reference sample labeled with Cy3. The reference sample contained male and female wild type pupae from all stages of metamorphosis. All experiments were conducted in triplicate.
Project description:This SuperSeries is composed of the following subset Series: GSE11311: Drosophila Sex Hierarchy Regulated Gene Expression in 48 hour APF Pupae GSE11313: Drosophila Gene Expression During Metamorphosis in Wild Type and Germline Minus Pupae Keywords: SuperSeries Refer to individual Series
Project description:Drosophila melanogaster undergoes a complete metamorphosis, during which time the larval male and female forms transition into sexually dimorphic, reproductive adult forms. To understand this complex morphogenetic process at a molecular-genetic level, we performed whole genome microarray analyses. In somatic tissues at 48 hour After Puparium Formation (APF), 173 sex-biased transcripts that likely function downstream of the doublesex (dsx) branch of the sex determination hierarchy were identified. The mode of regulation of the sex-specific isoforms of DSX (DSX-F and DSX-M) was examined. It was determined that for most downstream targets, DSX-F and DSX-M regulate gene expression in the same manner, but that one isoform acts as a more potent regulator. Keywords: wild type; genetic modification All microarrays were dual channel with direct comparisons of male versus female or wild type versus mutant. All samples consist of whole body pupae collected at 48 hour After Puparium Formation (APF). For each experiment, four biological replicates were analyzed in a dye-swap design.
Project description:In Drosophila, male-specific FRU (FRUM) is required to establish the potential for courtship behaviors, but the downstream effectors of FRUM during development are largely unknown. A microarray-based approach identified genes that are differentially expressed as a consequence of FRUM in pupae, in both whole body and CNS tissues. Genes were also identified that are sex-differentially expressed in CNS tissues. The FRUM-regulated sets were significantly overrepresented with genes also regulated by the ecdysone regulatory pathway. Two EcR isoforms (EcRA and EcRB1) are expressed in FRUM-expressing neurons during distinct periods of metamorphosis. Males with abrogated EcRA function in FRUM-expressing neurons aggressively court other males. Transcriptional profiles of mutants with abrogated EcRA function in the fru circuit demonstrate that EcR and FRUM regulate common gene sets, including the early gene broad. These results demonstrate a novel role for EcR in specifying male courtship behavior through its actions specifically in the FRUM neural circuitry. All microarrays were dual channel with direct comparisons of male versus female, wild type versus mutant, or experimental versus control. For each experiment, four to eight independent biological samples were analyzed using a dye-swap design. Samples consisted of whole body pupae or dissected CNS collected either at 0 hour After Pupal Formation (APF), 48 hour APF, or 0-24 hour adults.
Project description:Drosophila melanogaster undergoes a complete metamorphosis, during which time the larval male and female forms transition into sexually dimorphic, reproductive adult forms. To understand this complex morphogenetic process at a molecular-genetic level, we performed whole genome microarray analyses. Genes were identified that were expressed during metamorphosis in both somatic and germline tissues of males and females. Additionally, genes were identified that display sex-specific differences in abundance in both of these tissues at discrete times during metamorphosis. Keywords: time course; wild type; genetic modification; Overall design: Gene expression was examined at five time points during metamorphosis: 0, 24, 48, 71, and 96 hr After Puparium Formation (APF). Gene expression was examined separately in males and females for both wild type pupae and tudor (tud) progeny. tud progeny have genetically ablated germline tissues. All samples were labeled with Cy5 and compared against a common reference sample labeled with Cy3. The reference sample contained male and female wild type pupae from all stages of metamorphosis. All experiments were conducted in triplicate.
Project description:Background: Synaptic transmission is required for functional maturation of the CNS and to induce gene transcription involved in neuronal plasticity. Our aim is to identify genes that are transcriptionally regulated by synaptic transmission during development. cDNA microarrays will be used to compare gene expression in normal embryos and embryos with no synaptic transmission. Using the Gal4 UAS system, the tetanus toxin light fragment (TET) will be expressed throughout the nervous system. The effects of TET are well characterised: the synaptic protein n-Synaptobrevin is cleaved, blocking evokes vesicle fusion in TET expressing neurons and as a result synaptic transmission is blocked. Embryos expressing an inactive, mutant version of TET (iTET) show no detectable phenotype. Thus a comparison of gene expression in iTET expressing embryos and TET expressing embryos focuses our screen cleanly and specifically on genes that are regulated by synaptic transmission. At the end of larval life, during metamorphosis, there is a second phase of nervous system development with the same requirement for neural circuit differentiation and maturation as in the embryo. We will compare gene expression in TET expressing and iTET expressing pupae to identify genes which are regulated by synaptic transmission in this second phase of nervous system development. We will focus our analysis on genes which are similarly regulated in both systems. Plan: elav Gal4 (expressed in all the neurons) for the embryos and Heat Shock Gal4 (which allows a temporally controlled expression) for the pupae are used to drive UAS TET and UAS iTET throughout the nervous system. Crosses are set up with flies homozygous for Gal4 or UAS transgenes, so that all the progeny have the same genotype. Total RNA is extracted from 500 carefully staged embryos that have been collected at the time of hatching. Pupae are heat shocked at 24 hours after puparium formation, and heads are then collected 5 days after puparium formation. TET expressing pupae are paralysed but morphologically normal whereas iTET expressing pupae emerge as normal adults. Total RNA will be extracted from 250 heads of each genotype. Probes made from TET and iTET expressing embryos extracts will be hybridised together to the chip and so will be probes made from TET and iTET expressing pupae extracts. Genes regulated in both systems will be compared together.
Project description:Many known miRNAs in fish come from zebrafish and fugu whose genome sequence data are available. The Japanese flounder undergoes typical metamorphosis which is characterized by major morphological, functional, and behavioral changes during growth due to this metamorphosis from larva to juvenile. Metamorphosis is a biological process by which an animal physically develops after birth or hatching, involving a conspicuous and relatively abrupt change in the animal's body structure through cell growth and differentiation. Here, the high-throughput sequencing was adopted to identify the miRNAs during metamorphosis in the Japanese flounder. We found abundant microRNAs during metamorphosis in the Japanese flounder. Small RNAs were sequenced from metamorphosis stages of Japanese flounder
Project description:Insect cuticle plays essential roles in multiple physiological functions. During molting and metamorphosis, tremendous changes occur in silkworm cuticles. Silkworm is a model of Lepidoptera insects; however, little is known about the stage expression profiles of genes in cuticles of silkworm. In the present study, we selected 16 developmental stages, ranging from day 1 of the first instar larvae to day 8 of pupae, to perform microarray-based expression profiles. The data told us that various functions and physiological pathways were activated in the cuticle. Moreover, the expression profiles of cuticular protein genes, as the important components of cuticle, were investigated. The current study provides important insights for the functional study of insect cuticle and the regulation of insect cuticular protein genes. Transcription profiling experiments, 16 developmental stages (samples) were analyzed. Dual-channel experiments, with test samples labeled by Cy5 and common reference samples labeled by Cy3. Common reference sample was used for data normalization. One biological replicate. No dye-swaps.
Project description:Background: MicroRNAs (miRNAs) repress target genes at the post-transcriptional level, and function in the development and cell-lineage pathways of host species. Tissue-specific expression of miRNAs is highly relevant to their physiological roles in the corresponding tissues. However, to date, few miRNAs have been spatially identified in the silkworm. Results: We establish for the first time the spatial expression patterns of nearly 100 miRNAs in multiple normal tissues (organs) of Bombyx mori females and males using microarray and Northern-blotting analyses. In total, only 10 miRNAs were universally distributed (including bmo-let-7 and bmo-bantam), while the majority were expressed exclusively or preferentially in specific tissue types (e.g. bmo-miR-275 and bmo-miR-1). We additionally examined the developmental patterns of miRNA expression during metamorphosis of the body wall, silk glands, midgut and fat body. In total, 63 miRNAs displayed significant alterations in abundance in at least 1 tissue during the developmental transition from larvae to pupae (e.g. bmo-miR-263b and bmo-miR-124). Expression patterns of five miRNAs were significantly increased during metamorphosis in all four tissues (e.g. bmo-miR-27 and bmo-miR-305). Conclusions: In this study, we conducted preliminary spatial measurements of several miRNAs in the silkworm. Periods of rapid morphological change were associated with alterations in miRNA expression patterns in the body wall, silk glands, midgut and fat body during metamorphosis. Accordingly, we propose that this ubiquitous or tissue-specific expression of miRNAs supports their critical roles in tissue specification. The results obtained should facilitate future functional analyses. To determine the global spatial expression patterns of miRNAs in silkworm, we designed a DNA oligonucleotide-based microarray examining 92 unique miRNAs with 106 antisense probes. To determine the extent of tissue-specific changes during the specific developmental events, we assessed changes in miRNA expression in four individual tissues and organs (body wall, silk glands, midgut and fat body) from the larval to pupal stages.
Project description:Expression profiling of Drosophila mir-8 homozygous mutant pupae at a single developmental stage (72 hours APF). Homozygous mutant and wild type pupae were collected at 72 hours APF (After Puparium Formation). Three independent collections were performed. Total RNA was extracted and amplified. Mutant and respective wildtype samples were hybridized together to custom cDNA arrays (DGC1 and 2 EST collections).
Project description:The transcriptome of Blochmannia floridanus, the endosymbiont of the carpenter ant Camponotus floridanus, is presented during eight developmental stages of its holometabolous host by use of a whole genome DNA-macroarray. The detected transcription patterns indicate the presence of local transcription units as well as global regulatory mechanisms. Yet, the overall regulation scale is very modest, rarely exceeding a factor of three which is in line with the low number of transcriptional regulators present in this reduced genome. A large number of genes show differential expression in different life stages and a distinct expression pattern of genes possibly involved in symbiotic function as compared to housekeeping genes is apparent. However, these transcriptional changes are small as compared to the changes in the number of bacteria during host development which is highest in pupae and in young imagines. Control of replication of the bacteria in certain life stages may therefore be the decisive parameter influencing overall gene expression of Blochmannia in the animal. The few highly expressed genes like those encoding molecular chaperones exhibit a significantly higher G+C-content than moderately expressed genes. Keywords: Host stage-dependent gene expression C. floridanus laboratory colonies were maintained under constant conditions at 25°C with a 12 h light cycle in artificial nests and fed twice a week with cockroaches (Nauphoeta cinerea), Bhatkar agar (Bhatkar and Whitcomb, 1970), and honey water (50% wt/wt) ad libitum. Developmental stages were defined as L1 (very young larvae, still clustered), L2 (older larvae, approx. 2-4 mm), P1 (pupae before metamorphosis), P2 (pupae after metamorphosis, still uncolored), P3 (pupae shortly before eclosion with dark abdomens), W1 (workers shortly after eclosion, not completely melanized, no aggressive behavior), W2 (fully melanized adult workers from the nest, age not distinguishable), and W3 (adult workers from isolated subcolonies, at least 4 months old). For RNA preparations, midguts of the respective life stage (L2 - W3) were dissected and bacteria were isolated by homogenization and centrifugation. Endosymbionts from small larvae (L1) were isolated from whole animals. RNA for each cDNA array experiment was isolated from individual bacterial preparations; from L1 to W2, each developmental stage was represented by at least three independent experiments and samples from at least three different ant colonies. Two ant colonies (C79 and C118) were represented in every stage from L1 to W2 with additional samples from six other colonies. Old W3 workers were collected from two isolated subcolonies derived from the same mother colony, each sampled in duplicate.