Huckebein is part of a combinatorial repression code in the anterior blastoderm

The hierarchy of the segmentation cascade responsible for establishing the Drosophila body plan is composed by gap, pair-rule and segment polarity genes. However, no pair-rule stripes are formed in the anterior regions of the embryo. This lack of stripe formation, as well as other evidence from the literature that is further investigated here, led us to the hypothesis that anterior gap genes might be involved in a combinatorial mechanism responsible for repressing the cis-regulatory modules (CRMs) of hairy (h), even-skipped (eve), runt (run), and fushi-tarazu (ftz) anterior-most stripes. In this study, we investigated huckebein (hkb), which has a gap expression domain at the anterior tip of the embryo. Using genetic methods we were able to detect deviations from the wild-type patterns of the anterior-most pair-rule stripes in different genetic backgrounds, which were consistent with Hkb-mediated repression. Moreover, we developed an image processing tool that, for the most part, confirmed our assumptions. Using an hkb misexpression system, we further detected specific repression on anterior stripes. Furthermore, bioinformatics analysis predicted an increased significance of binding site clusters in the CRMs of h 1, eve 1, run 1 and ftz 1when Hkb was incorporated in the analysis, indicating that Hkb plays a direct role in these CRMs. We further discuss that Hkb and Slp1, which is the other previously identified common repressor of anterior stripes, might participate in a combinatorial repression mechanism controlling stripe CRMs in the anterior parts of the embryo and define the borders of these anterior stripes.     

Enhancer sequences influence the role of the amino-terminal domain of bicoid in transcription

Bicoid (Bcd) is a Drosophila melanogaster morphogenetic gradient that controls embryonic patterning by activating target gene expression in a concentration-dependent manner. In this study we describe experiments to determine how different enhancers respond to Bcd distinctively, focusing on two natural Bcd-responsive enhancer elements, hunchback (hb) and knirps (kni). Our results show that, on the hb enhancer element, the amino-terminal domain of Bcd (residues 1 to 91) plays primarily an inhibitory role, whereas on the kni enhancer element this same Bcd domain plays a positive role at low protein concentrations. We further demonstrate that while the amino-terminal domain is largely dispensable for cooperative binding to the hb enhancer element, it is preferentially required for cooperative binding to the kni enhancer element. Alteration of the arrangement of Bcd binding sites in the kni enhancer element reduces the role of the amino-terminal domain in cooperative DNA binding but increases the effectiveness of the self-inhibitory function. In addition, elimination of symmetric pairs of Bcd binding sites in the kni enhancer element reduces both DNA binding and activation by Bcd. We propose that the amino-terminal domain of Bcd is an enhancer-specific switch that contributes to the protein's ability to activate different target genes in distinct manners.     

Bicoid - morphogen function revisited

Bicoid (Bcd) functions as a morphogen during Drosophila development. Accordingly, bcd mRNA is maternally localized to the anterior pole of the embryo, and Bcd forms an anterior/posterior gradient, which functions in a concentration dependent fashion. Thus, nuclei receiving identical amounts of Bcd should express the same target genes. However, we found that ectopic, uniform expression of Bcd causes anterior gene expression in the posterior with mirror image polarity, indicating that one or several additional factors must provide positional information. Recently, we have shown that one of these factors is Capicua (Cic), a ubiquitous maternal repressor that is down-regulated at the embryonic termini by maternal Torso, a key component of the maternal terminal system. Cic acts on Bcd dependent enhancer elements by repression and thereby controls the posterior limit of Bcd target gene expression. Based on these new findings, we propose that spatial control of gene expression in the anterior region of the embryo is not solely the result of Bcd morphogen action. Rather, it relies on a "morphogenic network" that integrates the terminal system and Bcd activities, providing both polarity and spatial information to the prospective head region of the developing embryo.     

A system of repressor gradients spatially organizes the boundaries of Bicoid-dependent target genes

The homeodomain (HD) protein Bicoid (Bcd) is thought to function as a gradient morphogen that positions boundaries of target genes via threshold-dependent activation mechanisms. Here, we analyze 66 Bcd-dependent regulatory elements and show that their boundaries are positioned primarily by repressive gradients that antagonize Bcd-mediated activation. A major repressor is the pair-rule protein Runt (Run), which is expressed in an opposing gradient and is necessary and sufficient for limiting Bcd-dependent activation. Evidence is presented that Run functions with the maternal repressor Capicua and the gap protein Kruppel as the principal components of a repression system that correctly orders boundaries throughout the anterior half of the embryo. These results put conceptual limits on the Bcd morphogen hypothesis and demonstrate how the Bcd gradient functions within the gene network that patterns the embryo.     

The Snail repressor positions Notch signaling in the Drosophila embryo

The maternal Dorsal nuclear gradient initiates the differentiation of the mesoderm, neurogenic ectoderm and dorsal ectoderm in the precellular Drosophila embryo. Each tissue is subsequently subdivided into multiple cell types during gastrulation. We have investigated the formation of the mesectoderm within the ventral-most region of the neurogenic ectoderm. Previous studies suggest that the Dorsal gradient works in concert with Notch signaling to specify the mesectoderm through the activation of the regulatory gene sim within single lines of cells that straddle the presumptive mesoderm. This model was confirmed by misexpressing a constitutively activated form of the Notch receptor, Notch(IC), in transgenic embryos using the eve stripe2 enhancer. The Notch(IC) stripe induces ectopic expression of sim in the neurogenic ectoderm where there are low levels of the Dorsal gradient. sim is not activated in the ventral mesoderm, due to inhibition by the localized zinc-finger Snail repressor, which is selectively expressed in the ventral mesoderm. Additional studies suggest that the Snail repressor can also stimulate Notch signaling. A stripe2-snail transgene appears to induce Notch signaling in 'na?ve' embryos that contain low uniform levels of Dorsal. We suggest that these dual activities of Snail, repression of Notch target genes and stimulation of Notch signaling, help define precise lines of sim expression within the neurogenic ectoderm.