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El pdf del artículo es el manuscrito de autor (PMID:21664584).-- et al., Developing tissues are patterned by coordinated activities of signaling systems, which can be integrated by a regulatory region of a gene that binds multiple transcription factors or by a transcription factor that is modified by multiple enzymes. Based on a combination of genetic and imaging experiments in the early Drosophila embryo, we describe a signal integration mechanism that cannot be reduced to a single gene regulatory element or a single transcription factor. This mechanism relies on an enzymatic network formed by mitogen-activated protein kinase (MAPK) and its substrates. Specifically, anteriorly localized MAPK substrates, such as Bicoid, antagonize MAPK-dependent downregulation of Capicua, a repressor that is involved in gene regulation along the dorsoventral axis of the embryo. MAPK substrate competition provides a basis for ternary interaction of the anterior, dorsoventral, and terminal patterning systems. A mathematical model of this interaction can explain gene expression patterns with both anteroposterior and dorsoventral polarities. © 2011 Elsevier Inc., SYS acknowledges partial support by NSF via grant DMS-0718604, as well as P50 GM071508 and RO1 GM078079 grants from the NIH. GJ acknowledges support by ICREA by grants from MICINN (BFU2008-01875) and AGAUR (2009SGR-1075). HL was supported by NSF DBI-0649833, NIH NS058465, Sloan Foundation, and DuPont Foundation.

To identify genes expressed in interesting spatial and temporal patterns during development, we mobilized transposons carrying the yeast transcriptional activator, GAL4, to create new insertions throughout the Drosophila genome. At these new sites, neighboring enhancers drive expression of GAL4 in a pattern similar to that of nearby genes (Brand and Perrimon, 1993). Since GAL4 binds to a UAS target sequence, flies containing a GAL4 transposon can activate transcription of a UAS-linked gene, thereby resulting in controlled expression of that secondary gene. Thus, the GAL4 P element is a useful tool both for identifying genes of interest and for ectopically expressing genes in novel tissues or at specific developmental stages. We mobilized a GAL4 insertion on the second chromosome (P{GawB} CY2; Queenan et al., 1997) to generate a collection of GAL4-expressing enhancer trap lines. F1 “jumpstarter” males (w/Y; GAL4/CyO; Sb 2-3/ ) were mated in vials to attached-X (C(1)DX/Y) females. From 200 such crosses, red-eyed, curly-winged, long-bristled sons (bearing a new GAL4 insertion) were selected and mated individually to w/w females to establish 50 lines. Chromosome-mapping data, enhancer trap pattern, and other phenotypic characters demonstrated that four insertions were duplicates, yielding a total of 46 independent lines (Table 1). All X and third chromosome lines are viable, although three lines exhibit reduced viability or fertility (lines 15, 45, and 50). We could not determine the viability of the CyOlinked lines since the balancer chromosome is homozygous lethal. The genotypes of the resulting stocks are: X-chromosome lines, w P{GawB}, second-chromosome lines, y w; Pin/CyO, P{GawB}, and third chromosome lines, y w; III P{GawB}. Line 15 also carries the FM6, y w Bar balancer while lines 45 and 50 contain TM3, Sb Ser. Finally, line number three exhibits a striking eye phenotype consisting of a variable loss of eye pigment in a gradient along the anterior/posterior axis. Surprisingly, this line lacks detectable GAL4 expression in larval eye discs. We characterized sequences flanking a subset of the insertions in our collection by the inverse PCR method (http:www.fruitfly.org/methods/). Table 1 lists neighboring predicted or known genes. To identify and visualize the morphology of cells expressing GAL4, we crossed females from each line to males carrying UAS-taulacZ, which links the -galactosidase enzyme to the tubulin-binding protein Tau (Hildago et al., 1995). Some lines (1, 7, 10 (males only), 14, 22, 25, 26, 32, 35, 36, and 41) were lethal in pupal stages when crossed to this reporter; therefore, to assay ovary and testis expression, we crossed these strains to UAS-lacZ.NZ, which contains a nuclear-localization signal (Y. Hiromi and S. West, unpubl. results). Tables 1 and 2 describe the expression patterns for each line and Figure 1 shows selected patterns that illustrate key points. More detailed descriptions of all the patterns are available upon request. All lines exhibit interesting staining in at least one developmental stage. As commonly noted, we observe patchy or variable GAL4 expression in many lines. For example, GAL4 drives embryonic PNS expression in 18 lines (Table 1). While all embryos within a given strain exhibit a general PNS pattern that is consistent, within an individual embryo, slight variations often occur between segmental repeats (e.g., brackets in Fig. 1a). Only a few lines express GAL4 in a single tissue at a specific developmental stage. For example, line 20 drives expression in the longitudinal visceral mesoderm from stage 12 onwards (Fig. 1c). When crossed to UASGFP, line 20 can be used to visualize this highly migratory tissue in vivo. As Tables 1 and 2 indicate, many lines express GAL4 in identical tissues. Nevertheless, important differences may exist in the distribution of expression within each tissue. For example, Fig. 1f–i shows representative wing discs from four lines that express GAL4 in all imaginal discs, the brain, and the central nervous system (IGD, brain, and CNS). The distinct patterns shown in the wing discs presumably reveal specific regulatory elements that govern expression of nearby