Project description:The adaptable transcriptional response to changes in food availability not only ensures animal survival, but also permits progressing with embryonic development. Interestingly, the central nervous system is preferentially protected to periods of malnutrition, a phenomenon known as ‘brain sparing’. However, the mechanisms that mediates this genetic response remains poorly understood. To get a better understanding of this, we used Drosophila melanogaster as a genetic model, analysing the transcriptional response of larval neural stem cells (neuroblasts) and glial cells of the blood-brain barrier during nutrient restriction using the targeted DamID technique.
Project description:In this study we use targeted DamID to assay RNA polymerase occupancy across the genome specifically within the seam cells and hypodermis of C. elegans to identify genes expressed in these cell-types.
Project description:To adapt RNA polymerase DamID (RAPID) for FLP-mediated spatial control in Caenorhabditis elegans, we inserted a Dam::rpb-6 fusion gene downstream of a FRT-flanked mCh::his-58 cassette and under control of the hsp-16.41 promoter. We introduced a single copy of this construct into the C. elegans genome and crossed the resulting line with a dpy-7p::FLP driver to enable basal Dam::rpb-6 expression in the hypodermis. Nematodes were cultured at 20 degrees Celcius to ensure low levels of Dam::RPB-6 expression in the hypodermis and total genomic DNA was purified from L4 larvae. DNA from animals expressing GFP::Dam was used to control for unspecific methylation. The genome-wide association profile of Dam::RPB-6 was determined by deep sequencing, which revealed a list of 2331 protein coding genes with FDR < 0.05. Original RAPID reference: Gomez-Saldivar et al (2020) Tissue-Specific Transcription Footprinting Using RNA PoI DamID (RAPID) in Caenorhabditis elegans. Genetics 216, 931–945. doi:10.1534/genetics.120.303774
Project description:Targeted DamID was performed to identify binding targets of P-element somatic inhinitor (Psi), Myc and RNA Polymerase (using the RpII18 subunit common to all 3 polymerases). Scalloped(sd)-GAL4 was used to restrict expression to the wing disc.
Project description:Exploring downstream regulation by the transcription factor FoxO is necessary to understand its function as a tumor suppressor or promoter in different biological contexts. Here, we employed RNA-seq profiling on single clones isolated using laser capture microdissection from Drosophila larval eye imaginal discs to identify FoxO targets that restrict the proliferation of Tsc1-deficient cells under nutrient restriction (NR).
Project description:Maternal undernutrition during pregnancy followed by ad libitum access to nutrients during postnatal life induces postnatal metabolic disruptions in multiple species. As skeletal muscle is a major metabolic organ, RNAseq was performed on the longissimus dorsi muscles of slaughter-weight adult females that had been exposed to nutrient-restriction in utero.
Project description:We addressed the molecular mechanisms and biological processes underlying the differences in cerebrum, liver, and muscle gene expression of 50 days fetuses in response to early maternal nutrient restriction. To this end, we employed tissue-to-tissue and tissue-specific network approaches using the genome-wide expression profile of 14 samples for two experimental conditions [Control – CON (n = 7) and 40% nutrient restriction – RES (n = 7)]. After RNA quality control, read mapping was performed with STAR aligner to the bovine ARS-UCD1.2 reference genome. Count normalization was performed by using the VST function from DESEq2. Gene networks were constructed using the PCIT algorithm, whereas differentially co-expressed genes were identified based on the R-package DGCA. Significant gene-gene co-expression was taken based on r > | 0.9| and then filtered by differentially expressed genes and transcription factors. Networks were visualized in Cytoscape, and functional analyses were carried out on Cluego and ShinyGo tools. We found that the cerebrum, liver, and muscle tissues were affected by nutrient restriction leading to differences in transcriptional regulation between CON and RES groups. Furthermore, we shed light on the nutrient-sensing pathways across tissues such as mTOR, PI3K/Akt, and insulin. This project was supported by Agriculture and Food Research Initiative Competitive Grant no. 2016-67016-24946 from the USDA National Institute of Food and Agriculture as well as the USDA-NIFA-AFRI (2018-67011-31708).
Project description:The aim of this study was to identify fetal adaptations to gene expression in our mouse model of intrauterine growth restriction (IUGR). Starting at E6.5, pregnant CD-1 mice received either control (ad libitum) or 70% of ad libitum food (maternal nutrient restriction [MNR]). At E18.5, fetal right liver lobes were collected from 2 pups/litter in litters with 11-15 pups. Frozen liver was homogenized for RNA isolations. Isolated RNA was sequenced on a HiSeq2500 with an average read depth of 23M reads and a range of 17-31M reads per sample. Reads were aligned to the mm9 transcriptome with Bowtie2. Normalization and differential expression was done with DESeq2, ALDex2 and EdgeR, and genes were considered in the study if they were consistent in 2 or more tools (FDR <0.1).