Project description:RNA was extracted using Trizol from heads of 5 Heliconius species: H. erato, H. charithonia, H. melpomene, H. doris and H. sara. An illumina TruSeq kit was used to generate RNA-Seq libraries which were sequenced using 100 bp paired-end sequencing. Sequence data was used to compare gene expression between males and females within species. Differentially expressed genes were annotated for functions and compared across species. Data was also used to detect for dosage compensation in all species.
Project description:The staphylococcal accessory regulator A (sarA) impacts the extracellular accumulation of Staphylococcus aureus virulence factors at the level of intracellular production and extracellular protease-mediated degradation. To assess the relative impact of these two functions, we previously used a proteomics approach that measures protein abundance as a function of all proteoforms to demonstrate that mutation of sarA results in increased levels of extracellular proteases and assess the impact of this on the accumulation of S. aureus exoproteins1. While this approach confirmed that protease-mediated degradation has a significant impact on the S. aureus exoproteome, it was potentially limited in that it did not take into account the possibility that large, stable proteolytic products from a given protein could result in false negatives when quantified by total proteoforms. Here, we present an expanded proteomics approach that utilizes a dual quantitative method for measuring abundance at both the total proteoform and full-length exoprotein levels. Specifically, proteins present in conditioned medium from overnight, stationary phase cultures of the USA300 strain LAC, an isogenic sarA mutant, and a sarA mutant unable to produce any of the known extracellular proteases (sarA/protease) were resolved using one-dimensional gel electrophoresis. Using methods that focus on total proteoforms vs. methods that focus specifically on full-length proteins, quantitative proteomic comparisons of sarA vs sarA/protease mutants identified proteins that were degraded in a protease dependent manner owing to mutation of sarA, while comparisons of a sarA/protease mutant vs the LAC parent strain identified proteins in which abundance was altered in a sarA mutant in a protease-independent manner. Furthermore, the proteins uniquely identified by the full-length data analysis approach eliminated false negatives observed in the total proteoform analysis. This approach provided for a more comprehensive and robust analysis of the impact of mutating sarA and protease-mediated degradation on the S. aureus exoproteome.
Project description:We use RNAseq data to perform differential gene expression to identify genes controlling structural colouration in two co-mimetic species of Heliconius butterfly - Heliconius erato and Heliconius melpomene. We use comparisons between iridescent and non-iridescent subspecies of Helcionius erato (H. e. cyrbia and H. e. demophoon, respectively) and Helcionius melpomene (H. m. cythera and H. m. rosina, respectively) at two separate developmental stages, 50% and 70% of development. In addition, in the iridescent subspecies of both H. erato and H. melpomene, we compared the iridescent wing regions (forewing and hindwing combined) to the non-iridescent androconial wing region using differential gene expression.
Project description:We use RNAseq data to perform differential gene expression analysis to identify genes controlling structural colouration in two co-mimetic species of Heliconius butterfly - Heliconius erato and Heliconius melpomene. We use comparisons between iridescent and non-iridescent subspecies of Helcionius erato (H. e. cyrbia and H. e. demophoon, respectively) and Helcionius melpomene (H. m. cythera and H. m. rosina, respectively) at two separate developmental stages, 50% and 70% of development. In addition, in the iridescent subspecies of both H. erato and H. melpomene, we compared the iridescent wing regions (forewing and hindwing combined) to the non-iridescent androconial wing (anterior hindwing) region using differential gene expression.
Project description:Activated myofibroblasts play an essential role in tissue fibrogenesis by producing extracellular matrixes (ECM). ECMs replace normal functioning tissue, reducing tissue plasticity and impairing functions of the organ. Recent studies suggested that pericytes are a major source of myofibroblast precursors. We previously showed that Smad Anchor for Receptor Activation (SARA) prevents cellular phenotypic transdifferentiation toward mesenchymal cells, and depletion of SARA induces transdifferentiation of epithelial cells to fibroblast-like phenotype. Here, we generated a transgenic mice that overexpress SARA specifically pericytes by using PDGFRβ-Cre (SARATg, PDGFRβ-Cre). When subjected to either subcutaneous injection of bleomycin or intraperitoneal administration of aristolochic acid, which induces skin and kidney fibrosis, respectively, SARATg, PDGFRβ-Cre mice developed significantly less fibrosis compared to SARAWT, PDGFRβ-Cre mice. To decipher molecular signature and pericyte trajectory under fibrotic conditions and effects of SARA overexpression, we isolated PDGFR+ cells from skin or kidney of SARATg or WT, PDGFRβ-Cre, Z/EG mice with or without fibrotic stimuli and performed scRNAseq analyses. In both skin and kidney sample sets, we found pericytes and immune cell populations are the major cell components among the PDGFR+ GFP+ cells. We found that pericyte populations are divided into canonical and non-canonical sub-populations and the latter has assumed myofibroblast characteristics. Trajectory mapping revealed a single path from canonical to non-canonical pericyte sub-populations and SARA overexpression truncated the trajectory. These results suggest that SARA prevents pericyte transdifferentiation into myofibroblasts under fibrotic condition, and therefore anti-fibrotic.