title: Evolution of New cis-Regulatory Motifs Required for Cell-Specific Gene Expression in Caenorhabditis.
abstract: Patterning of C. elegans vulval cell fates relies on inductive signaling. In this induction event, a single cell, the gonadal anchor cell, secretes LIN-3/EGF and induces three out of six competent precursor cells to acquire a vulval fate. We previously showed that this developmental system is robust to a four-fold variation in lin-3/EGF genetic dose. Here using single-molecule FISH, we find that the mean level of expression of lin-3 in the anchor cell is remarkably conserved. No change in lin-3 expression level could be detected among C. elegans wild isolates and only a low level of change-less than 30%-in the Caenorhabditis genus and in Oscheius tipulae. In C. elegans, lin-3 expression in the anchor cell is known to require three transcription factor binding sites, specifically two E-boxes and a nuclear-hormone-receptor (NHR) binding site. Mutation of any of these three elements in C. elegans results in a dramatic decrease in lin-3 expression. Yet only a single E-box is found in the Drosophilae supergroup of Caenorhabditis species, including C. angaria, while the NHR-binding site likely only evolved at the base of the Elegans group. We find that a transgene from C. angaria bearing a single E-box is sufficient for normal expression in C. elegans. Even a short 58 bp cis-regulatory fragment from C. angaria with this single E-box is able to replace the three transcription factor binding sites at the endogenous C. elegans lin-3 locus, resulting in the wild-type expression level. Thus, regulatory evolution occurring in cis within a 58 bp lin-3 fragment, results in a strict requirement for the NHR binding site and a second E-box in C. elegans. This single-cell, single-molecule, quantitative and functional evo-devo study demonstrates that conserved expression levels can hide extensive change in cis-regulatory site requirements and highlights the evolution of new cis-regulatory elements required for cell-specific gene expression.
authors: Institute of Biology of the Ecole Normale Supérieure (IBENS), CNRS, Inserm, Paris, France.|Department of Life Sciences, Imperial College, London, United Kingdom.
authors: Michalis
authors: 0000-0003-1974-7668
authors: M
authors: Barkoulas
authors: Institute of Biology of the Ecole Normale Supérieure (IBENS), CNRS, Inserm, Paris, France.
authors: Amhed M
authors: AM
authors: Vargas Velazquez
authors: Institute of Biology of the Ecole Normale Supérieure (IBENS), CNRS, Inserm, Paris, France.
authors: Alexandre E
authors: 0000-0001-7216-4205
authors: AE
authors: Peluffo
authors: Institute of Biology of the Ecole Normale Supérieure (IBENS), CNRS, Inserm, Paris, France.
authors: Marie-Anne
authors: MA
authors: Félix
mesh_terms: D000818:Animals; D019521:Body Patterning / Q000235:genetics; D017173:Caenorhabditis elegans / Q000235:genetics* / Q000254:growth & development; D029742:Caenorhabditis elegans Proteins / Q000235:genetics*; D002454:Cell Differentiation / Q000235:genetics; D004330:Drosophila / Q000235:genetics / Q000254:growth & development; D024721:E-Box Elements / Q000235:genetics*; D004815:Epidermal Growth Factor / Q000235:genetics*; D005260:Female; D018507:Gene Expression Regulation, Developmental; D011485:Protein Binding / Q000235:genetics; D018160:Receptors, Cytoplasmic and Nuclear / Q000235:genetics / Q000378:metabolism; D012045:Regulatory Sequences, Nucleic Acid / Q000235:genetics; D059010:Single-Cell Analysis; D014844:Vulva / Q000166:cytology / Q000254:growth & development*
chemical_list: D029742:Caenorhabditis elegans Proteins; D018160:Receptors, Cytoplasmic and Nuclear; C075912:Lin-3 protein, C elegans; D004815:Epidermal Growth Factor