Dr. Bender proposes to continue his studies on the Drosophila ecdysone receptor (EcR) gene which he initiated in the Hogness lab as a postdoctoral fellow. Three isoforms of EcR exist; they are denoted EcR-A, EcR-B1 and EcR-B2 and have identical DNA and hormone binding domains but differing amino termini. These isoforms arise from a single gene as a consequence of two promoters. The distal promoter produces EcR-A while the proximal promoter produces EcR-B1 and and EcR-B2. These isoforms exhibit different patterns of developmental expression, but it is not clear at this time whether these forms are functionally different. Determining the functional significance of these isoforms is one of two primary goals of this proposal. Dr. Bender has made an effort to characterize the Drosophila EcR gene. He isolated a series of EcR mutants and found them to be either of two types. Apparent nulls inactivate all three isoforms and are embryonic lethals. The other alleles inactivate only the B1 isoform and have a non-pupariating lethal phase. Transgenic strains bearing the three isoforms under hsp70 promoter control were constructed and were used to rescue a null allele. All three isoforms could effect partial rescue, although the B1 isoform was approximately 5X more effective. Therefore these experiments did not resolve the question of whether the isoforms have unique roles. This proposal has two main specific aims. In the first, Dr. Bender proposes to isolate more mutant alleles of EcR with the principal goal of independently inactivating each of the isoforms. The principal screen will be a transposon mutagenesis and this screen and the ways in which the mutants will be analyzed both molecularly and functionally is described in great detail. The second main specific aim is to identify additional genes involved in the ecdysone response by screening for enhancers or suppressors of EcR/usp double mutants. Dr. Bender has shown that EcR/usp double transheterozygotes have low viability, so this screen should offer a sensitive way to identify mutant alleles that affect either the availability of the ecdysone hormone or the ability to respond to its presence. Dr. Bender states that approximately 30 chromosome 2 EcR enhancer mutants have been identified in his screens conducted to date, and methods to characterize these enhancers and to expand the screen to chromosome 3 are described.
| Rayburn, Lowell Y M; Rhea, Jeanne; Jocoy, Steven R et al. (2009) The proprotein convertase amontillado (amon) is required during Drosophila pupal development. Dev Biol 333:48-56 |
| Carney, G E; Wade, A A; Sapra, R et al. (1997) DHR3, an ecdysone-inducible early-late gene encoding a Drosophila nuclear receptor, is required for embryogenesis. Proc Natl Acad Sci U S A 94:12024-9 |
