Identification of C. elegans DAF-12-binding sites, response elements, and target genes.
Intracellular receptor DAF-12 regulates dauer formation and developmental age and affects Caenorhabditis elegans lifespan. Genetic analyses place DAF-12 at the convergence of several signal transduction pathways; however, the downstream effectors and the molecular basis for the receptor's multiple physiological outputs are unknown. Beginning with C. elegans genomic DNA, we devised a procedure for multiple rounds of selection and amplification that yielded fragments bearing DAF-12-binding sites. These genomic fragments mediated DAF-12-dependent transcriptional regulation both in Saccharomyces cerevisiae and in C. elegans; that is, they served as functional DAF-12 response elements. We determined that most of the genomic fragments that displayed DAF-12 response element activity in yeast were linked to genes that were regulated by DAF-12 in C. elegans; indeed, the response element-containing fragments typically resided within clusters of DAF-12-regulated genes. DAF-12 target gene regulation was developmental program and stage specific, potentially predicting a fit of these targets into regulatory networks governing aspects of C. elegans reproductive development and dauer formation.
Shostak
Yuriy
Y
Program in Biochemistry and Molecular Biology, University of California, San Francisco, California 94143-2280, USA.
Van Gilst
Marc R
MR
Antebi
Adam
A
Yamamoto
Keith R
KR
D000818:Animals; D001483:Base Sequence; D001665:Binding Sites / Q000502:physiology; D017173:Caenorhabditis elegans; D029742:Caenorhabditis elegans Proteins / Q000235:genetics* / Q000378:metabolism* / Q000502:physiology; D005454:Fluorescence Polarization; D005786:Gene Expression Regulation; D005809:Genes, Regulator / Q000235:genetics / Q000502:physiology*; D017930:Genes, Reporter / Q000235:genetics; D049452:Green Fluorescent Proteins; D008969:Molecular Sequence Data; D009841:Oligonucleotides; D018160:Receptors, Cytoplasmic and Nuclear / Q000235:genetics* / Q000378:metabolism* / Q000502:physiology; D020218:Response Elements / Q000235:genetics*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012441:Saccharomyces cerevisiae; D017422:Sequence Analysis, DNA; D015398:Signal Transduction / Q000502:physiology
D029742:Caenorhabditis elegans Proteins; C410905:DAF-12 protein, C elegans; D009841:Oligonucleotides; D018160:Receptors, Cytoplasmic and Nuclear; D049452:Green Fluorescent Proteins