TheDrosophila Broad-ComplexEarly Gene Directly Regulates Late Gene Transcription during the Ecdysone-Induced Puffing Cascade

AbstractThe ensemble of tissue-specific changes that drivesDrosophilametamorphosis is initiated by the steroid hormone ecdysone and proceeds through a transcriptional cascade comprised of primary response transcriptional regulators and secondary response structural genes. TheBroad-Complex(BR-C) primary response early gene is composed of several distinct genetic functions and encodes a family of related transcription factor isoforms. Our objective in this study was to determine whether individual BR-C isoforms directly regulate secondary response target genes. A cluster of 10 salivary gland-specific secondary responseL71late genes are dependent on theBR-C rbp+genetic function. Transgenic animals expressing individual BR-C isoforms were tested for their ability to provide theBR-C rbp+genetic function by monitoring the transcriptional activation of theL71genes. We found that the BR-C Z1 isoforms could complement the transcriptional defects seen inrbpmutants but the Z2, Z3, and Z4 isoforms could not. We conclude that theBR-C rbp+function is provided by the BR-C Z1 isoform in prepupal salivary glands.L71gene rescue was restricted to the prepupal salivary gland, suggesting the involvement of additional factors inL71gene regulation. Interestingly, we found that the overexpression of Z3 or Z4 isoforms inBR-C+salivary glands repressedL71expression, indicating that BR-C proteins might also function as transcriptional repressors. Molecular mapping and characterization of the regulatory sequences that controlL71-6expression revealed several Z1 isoform binding sites. Mutagenesis of these Z1 binding sites resulted in the failure to activate late gene expressionin vivowhen measured by transgenic reporter genes. We conclude that theBR-Cearly gene directly activates late gene transcription by interacting with late genecis-acting regulatory elements and that this interaction is responsible for the temporal linkage of early and late ecdysone-induced gene expression