Project description:Loss of nautilus (MyoD) gene function results in a variable phenotype affecting muscle formation in embryos and larvae, larval movement, pupal eclosion, egg deposition, adult mobility and survival. 8-miR cluster deletion disrupts muscle formation in the embryo while affecting protein production from the nautilus, dMef2 , regulators of the muscle transcriptional network. We propose the complex phenotype in the nautilus null is due to the disruption of the regulatory interactions provided by the 8-miR cluster. The results demonstrate that nautilus is an integral regulator of the miRNA circuitry buffering the transcriptional network directing muscle development. Two-condition experiment, wild type (w1118) vs. mutant (8-miR cluster null). Biological replicates: 3 wild type, 3 mutants, independently isolated. One of the biological replicate was dye swaped to avoid dye bias.
Project description:Loss of nautilus (MyoD) gene function results in a variable phenotype affecting muscle formation in embryos and larvae, larval movement, pupal eclosion, egg deposition, adult mobility and survival. mir-3 over expression disrupts muscle formation in the embryo while affecting protein production from the dMef2 and tinman genes, global regulators of the muscle transcriptional network. We propose the complex phenotype in the nautilus null is due to the disruption of the regulatory interactions provided by the 8-miR cluster. The results demonstrate that nautilus is an integral regulator of the miRNA circuitry buffering the transcriptional network directing muscle development. Two-condition experiment, wild type (w1118) vs. mutant (mir-3 ectopic expressionl). Biological replicates: 3 wild type, 3 mutants, independently isolated. One of the biological replicate was dye swaped to avoid dye bias.
Project description:To determine if the Drosophila MyoD homolog, nautilus, was activating any miRNA loci, similar to vertebrate MyoD, we compared the miRNA expression profiles between wild-type (w1118) and nautilus null embryos during the window of maximum nautilus expression (6-8hr AEL), using LNA arrays specifically designed to quantify miRNA levels in Drosophila (Exiqon). Expression levels for mir-309, mir-3, mir-286, mir-4, mir-5, and mir-6 from the 8-miR cluster, were significantly decreased in nautilus null embryos. It suggests that the intergenic 8-miR cluster, encoding eight miRNAs, is regulated by nautilus. Two-condition experiment, wild type (w1118) vs. mutant (nautilus null). Biological replicates: 3 wild type, 3 mutants, independently isolated.
Project description:Embryos were collected, aged, mock-treated/treated with 40Gy gamma radiation, and allowed to recover for 1.5 hours. Targets from 3 sets of wild type (yw, w1118) and 2 sets of mutant (Dmp53NS) biological replicates were generated and the expression profiles were determined using Affymetrix Drosophila Genechip 1 arrays. Comparisons between the sample groups allow the identification of genes with radiation-responsive and Dmp53-dependent expression patterns. Experiment Overall Design: 3 sets of paired control and irradiated Wild Type and 2 sets of paired control and irradiated Mutant (Dmp53NS) biological replicates were analyzed
Project description:The initially homogeneous epithelium of the early Drosophila embryo differentiates into regional subpopulations with different behaviours and physical properties that are needed for morphogenesis. The factors at top of the genetic hierarchy that control these behaviours are known, but many of their targets are not. To understand how proteins work together to mediate differential cellular activities, we studied in an unbiased manner the proteomes and phosphoproteomes of the three main cell populations along the dorso-ventral axis during gastrulation using mutant embryos that represent the different populations. We detected 6111 protein groups and 6259 phosphosites of which 3399 and 3433 respectively, were differentially regulated. The changes in phosphosite abundance did not correlate with changes in host protein abundance, showing phosphorylation to be a regulatory step during gastrulation. Hierarchical clustering of protein groups and phosphosites identified clusters that contain known fate determinants such as Doc1, Sog, Snail and Twist. The recovery of the appropriate known marker proteins in each of the different mutants we used validated the approach, but also revealed that two mutations that both interfere with the dorsal fate pathway, Toll10B and serpin27aex do this in very different manners. Diffused network analyses within each cluster point to microtubule components as one of the main groups of regulated proteins. Functional studies on the role of microtubules provide the proof of principle that microtubules have different functions in different domains along the DV axis of the embryo. This repository is related to PXD046050 which represents the label-free proteome.
Project description:Post-transcriptional gene regulatory mechanisms are fundamental to the determination of gene expression dynamics and especially crucial for the earliest stages of animal development in which transcription is nearly silent. Here we performed high-resolution total RNA-sequencing and quantitative mass spectrometry analysis simultaneously on Drosophila maternal-to-zygotic transition (MZT) to reveal the dynamic interchange between transcript and protein expression level at the earliest stage of animal development. Intriguingly, we found that both stable transcript expression and an increase in translation efficiency can be crucial for protein upregulation prior to zygotic gene activation (ZGA), whereas such features diminished following ZGA. Further, this study is the first to report the proteome-wide quantitative changes in protein ubiquitination in Drosophila MZT. Our results indicate that timely ubiquitination of the distinct target proteins during MZT are essential for the downregulation of protein expression levels. Profiling of the RNA-associated proteome changes in Drosophila MZT suggested that RNA-binding activities can be regulated without the respective change in net protein expression levels for over 200 proteins, including Pcid2, Sym, and Cpsf73. Taken together, we find that stable expression of transcript can be crucial for the protein expression level upregulation during the earliest stage of MZT, and also report that proteome wide and quantitative remodeling of the distinct post translational regulatory mechanisms can be crucial for the gene expression level changes and progression of the earliest stages of animal development.
Project description:The initially homogeneous epithelium of the early Drosophila embryo differentiates into regional subpopulations with different behaviours and physical properties that are needed for morphogenesis. The factors at top of the genetic hierarchy that control these behaviours are known, but many of their targets are not. To understand how proteins work together to mediate differential cellular activities, we studied in an unbiased manner the proteomes and phosphoproteomes of the three main cell populations along the dorso-ventral axis during gastrulation using mutant embryos that represent the different populations. We detected 6111 protein groups and 6259 phosphosites of which 3399 and 3433 respectively, were differentially regulated. The changes in phosphosite abundance did not correlate with changes in host protein abundance, showing phosphorylation to be a regulatory step during gastrulation. Hierarchical clustering of protein groups and phosphosites identified clusters that contain known fate determinants such as Doc1, Sog, Snail and Twist. The recovery of the appropriate known marker proteins in each of the different mutants we used validated the approach, but also revealed that two mutations that both interfere with the dorsal fate pathway, Toll10B and serpin27aex do this in very different manners. Diffused network analyses within each cluster point to microtubule components as one of the main groups of regulated proteins. Functional studies on the role of microtubules provide the proof of principle that microtubules have different functions in different domains along the DV axis of the embryo.
Project description:cdipt is an essential gene in the synthesis of phosphatidylinositol (PtdIns) in the zebrafish, Danio rerio. The zebrafish mutant cdipt^hi559Tg (ZL782) carries a retroviral insertion which inactivates cdipt. Homozygous mutants exhibit hepatocellular endoplasmic reticulum (ER) stress and non-alcoholic fatty liver disease (NAFLD) pathologies at 5 days post fertilization (dpf). This study reveals a novel link between PtdIns, ER stress, and steatosis. We compared whole animal gene expression profiles of hi559 mutant larvae with phenotypically wild type larvae from a heterozygote incross in triplicate.
Project description:Rad21 is a subunit of cohesin. The main function of cohesin is to hold replicated chromosomes together until cells divide, but it also plays a role in gene expression. To find out which genes might be regulated by cohesin, a study was conducted to look for global changes in gene expression in zebrafish embryos lacking cohesin component Rad21. The zebrafish Rad21 mutant used for expression analysis was rad21nz171, an allele isolated in a forward genetic screen for regulators of runx1. Experiment Overall Design: RNA from rad21nz171 mutant and wild type zebrafish embryos collected at 24 hours post-fertilization (h.p.f.) and 48 h.p.f. was hybridized to Affymetrix microarrays (Gene Chip zebrafish genome arrays cat. no. 900488). Four pools of 50 embryos for each genotype and time point were used as the RNA source, and RNA from each pool was hybridized independently such that the experiment had four biological replicates.
Project description:We have used trypsin-mediated wounding to amplify the transcriptional response to clean puncture wounding. We studied the transcriptional response of clean puncture wounding and trypsin puncture wounding in relation to unwounded wild-type late-stage embryos. Trypsin puncture wounding generally increases the puncture wounding fold induction relative to unwounded controls.