The Co-activator CREB-binding Protein Participates in Enhancer-dependent Activities of Bicoid

Abstract

Bicoid (Bcd) is a transcriptional activator required for early embryonic patterning in Drosophila. Despite extensive studies, it currently remains unclear how Bcd activates transcription and what proteins participate in its activation process. In this report, we describe experiments to analyze the role of the Drosophila co-activator dCBP in Bcd-mediated activation. In Drosophila S2 cells, the Bcd activity is increased by the co-transfection of plasmids expressing dCBP and reduced by doublestranded RNA-mediated interference against dCBP. We further show that Bcd and dCBP can interact with each other and that Bcd-interacting domains of dCBP can cause dominant negative effects on Bcd activity in S2 cells. Our comparison of two Bcd-responsive enhancers, hunchback (hb) and knirps (kni), reveals a differential role of dCBP in facilitating Bcd activation. A dCBP mutant defective in its histone acetyltransferase activity exhibits a reduced, but not abolished, co-activator function for Bcd. Our chromatin immunoprecipitation experiments show that dCBP can increase not only the occupancy of Bcd itself at the enhancers but also the recruitment of general transcription factors to the promoter. Together, these experiments suggest that dCBP is an enhancer-dependent co-activator of Bcd, facilitating its activation through multiple mechanisms.

Bicoid cooperative DNA binding is critical for embryonic patterning in Drosophila

“…However, more is likely to be involved than DNA site occupancy, because the affinity of some Bicoid-site enhancers does not strictly correlate with the A-P expression (23) and because other transcription regulators play a critical role in determining A-P expression (15). Critical differences in the arrangement of Bicoid binding sites within an enhancer also effect synergistic interactions by Bicoid with coactivators, such as CREB-BP, thus altering transcription output (20). Such mechanisms are independent of Bicoid site-occupancy per se.…”

“…For example, we did not explicity test transcriptional synergy (6,53), which may be important for Bicoid activity (17,20,54). Also, in Drosophila embryos, and perhaps in other organisms, there may be independent mechanisms (e.g., stau-dependent) to ensure that morphogenetic gradients undergo ''error correction'' to enhance both accuracy and precision in their interpretation (13,55).…”

“…Some of these mysteries might be explained on the basis of Bicoid's interactions with other proteins (16)(17)(18)(19)(20). For example, Bicoid interacts with the Sap18-Rpd3 histone deacetylase complex, which likely converts Bicoid from an activator to a repressor in the anterior tip of the embryo (14,18,21).…”

Functional Role of CREB-Binding Protein in the Circadian Clock System of Drosophila melanogaster

“…For the construction of transgenic flies in which CBP expression is downregulated via a GAL4/UAS system, a doublestranded RNA (dsRNA) construct for CBP was designed according to the genomic cDNA hybrid method (22). The DNA fragment from bp 6721 to 7454 of CBP cDNA and genomic DNA including the cDNA with internal and adjacent 3Ј introns were amplified by PCR from pDF378 (16) and fly genomic DNA, respectively. The fragments were ligated together into the pUAST vector, and the transgenic construct was injected with pUCHsp⌬2-3 into w 1118 embryos, from which several germ line transformants were established.…”

Structure-Function Analysis of the Drosophila melanogaster Caudal Transcription Factor Provides Insights into Core Promoter-preferential Activation

“…dCBP has previously been shown to function as a Bicoid co-activator in S2 cells (77,78). giant (gt) is a Bicoid target gene (79 -81) whose core promoter contains a TATA box, an Inr, and a DPE motif (38).…”