The EGF receptor and N signalling pathways act antagonistically in Drosophila mesothorax bristle patterning.

An early step in the development of the large mesothoracic bristles (macrochaetae) of Drosophila is the expression of the proneural genes of the achaete-scute complex (AS-C) in small groups of cells (proneural clusters) of the wing imaginal disc. This is followed by a much increased accumulation of AS-C proneural proteins in the cell that will give rise to the sensory organ, the SMC (sensory organ mother cell). This accumulation is driven by cis-regulatory sequences, SMC-specific enhancers, that permit self-stimulation of the achaete, scute and asense proneural genes. Negative interactions among the cells of the cluster, triggered by the proneural proteins and mediated by the Notch receptor (lateral inhibition), block this accumulation in most cluster cells, thereby limiting the number of SMCs. Here we show that the proneural proteins trigger, in addition, positive interactions among cells of the cluster that are mediated by the Epidermal growth factor receptor (EGFR) and the Ras/Raf pathway. These interactions, which we denominate 'lateral co-operation', are essential for macrochaetae SMC emergence. Activation of the EGFR/Ras pathway appears to promote proneural gene self-stimulation mediated by the SMC-specific enhancers. Excess EGFR signalling can overrule lateral inhibition and allow adjacent cells to become SMCs and sensory organs. Thus, the EGFR and Notch pathways act antagonistically in notum macrochaetae determination.

Proneural gene self-stimulation in neural precursors: an essential mechanism for sense organ development that is regulated by Notch signaling.

To learn about the acquisition of neural fate by ectodermal cells, we have analyzed a very early sign of neural commitment in Drosophila, namely the specific accumulation of achaete-scute complex (AS-C) proneural proteins in the cell that becomes a sensory organ mother cell (SMC). We have characterized an AS-C enhancer that directs expression specifically in SMCs. This enhancer promotes Scute protein accumulation in these cells, an event essential for sensory organ development in the absence of other AS-C genes. Interspecific sequence comparisons and site-directed mutagenesis show the presence of several conserved motifs necessary for enhancer action, some of them binding sites for proneural proteins. These and other data indicate that the enhancer mediates scute self-stimulation, although only in the presence of additional activating factors, which most likely interact with conserved motifs reminiscent of NF-kappaB-binding sites. Cells neighboring the SMC do not acquire the neural fate because the Notch signaling pathway effectors, the Enhancer of split bHLH proteins, block this proneural gene self-stimulatory loop, possibly by antagonizing the action on the enhancer of the NF-kappaB-like factors or the proneural proteins. These data suggest a mechanism for SMC committment.

Cis-regulation of achaete and scute: shared enhancer-like elements drive their coexpression in proneural clusters of the imaginal discs.

The pattern of bristles and other sensory organs on the adult cuticle of Drosophila is prefigured in the imaginal discs by the pattern of expression of the proneural achaete (ac) and scute (sc) genes, two members of the ac-sc complex (AS-C). These genes are simultaneously expressed by groups of cells (the proneural clusters) located at constant positions in discs. Their products (transcription factors of the basic-helix-loop-helix family) allow cells to become sensory organ mother cells (SMCs), a fate normally realized by only one or a few cells per cluster. Here we show that the highly complex pattern of proneural clusters is constructed piecemeal, by the action on ac and sc of site-specific, enhancer-like elements distributed along most of the AS-C (approximately 90 kb). Fragments of AS-C DNA containing these enhancers drive reporter lacZ genes in only one or a few proneural clusters. This expression is independent of the ac and sc endogenous genes, indicating that the enhancers respond to local combinations of factors (prepattern). We show further that the cross-activation between ac and sc, discovered by means of transgenes containing either ac or sc promoter fragments linked to lacZ and thought to explain the almost identical patterns of ac and sc expression, does not occur detectably between the endogenous ac and sc genes in most proneural clusters. Our data indicate that coexpression is accomplished by activation of both ac and sc by the same set of position-specific enhancers.