EcR isoforms in Drosophila: testing tissue-specific requirements by targeted blockade and rescue.
The three Drosophila EcR isoforms differ only at their N termini; thus, they share the conserved ligand-binding domain transcriptional activation function (AF2) and only differ in the unconserved A/B region, which contains a second, isoform-specific, activation function (AF1). We have developed a dominant-negative mutant EcR (EcR-DN), expressed it in flies with the GAL4/UAS system, and used it to block ecdysone signaling in eight tissues or groups of tissues. Localized EcR-DN arrests ecdysone-dependent development in the target cells and often--because of a molting checkpoint--arrests development globally. Simultaneously expressing individual wild-type EcR isoforms in the same target tissues suppresses the EcR-DN phenotype and identifies the rescuing isoform as sufficient to support the development of the target. Every isoform, and even an N-terminal truncated EcR that lacks any AF1, supports development in the fat body, eye discs, salivary glands, EH-secreting neurosecretory cells and in the dpp expression domain, implying that AF1 is dispensable in these tissues. By contrast, only EcR-A is able to support development in the margins of the wing discs, and only EcR-B2 can do so in the larval epidermis and the border cells of the developing egg chamber. In light of our results, the simplest explanations for the widespread spatial and temporal variations in EcR isoform titers appear untenable.
Cherbas
Lucy
L
Department of Biology, Indiana University, Bloomington, IN 47405, USA. Icherbas@bio.indiana.edu
Hu
Xiao
X
Zhimulev
Igor
I
Belyaeva
Elena
E
Cherbas
Peter
P
D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D002875:Chromosomes / Q000378:metabolism; D029721:Drosophila Proteins / Q000235:genetics / Q000378:metabolism*; D004331:Drosophila melanogaster / Q000235:genetics / Q000254:growth & development* / Q000378:metabolism; D004440:Ecdysone / Q000378:metabolism; D005260:Female; D018507:Gene Expression Regulation, Developmental; D018390:Gene Targeting; D017930:Genes, Reporter; D007814:Larva / Q000033:anatomy & histology / Q000502:physiology; D008297:Male; D008969:Molecular Sequence Data; D009154:Mutation; D010641:Phenotype; D017956:Photoreceptor Cells, Invertebrate / Q000166:cytology / Q000254:growth & development; D020033:Protein Isoforms / Q000235:genetics / Q000378:metabolism; D017434:Protein Structure, Tertiary; D011987:Receptors, Steroid / Q000235:genetics / Q000378:metabolism*; D012469:Salivary Glands / Q000378:metabolism; D016415:Sequence Alignment; D015398:Signal Transduction / Q000502:physiology
D029721:Drosophila Proteins; D020033:Protein Isoforms; D011987:Receptors, Steroid; C055469:ecdysone receptor; D004440:Ecdysone