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Gain-of-function screens in Drosophila are an effective method with which to identify genes that affect the development of particular structures or cell types. It has been found that a fraction of 2-10% of the genes tested, depending on the particularities of the screen, results in a discernible phenotype when overexpressed. However, it is not clear to what extent a gain-of-function phenotype generated by overexpression is informative about the normal function of the gene. Thus, very few reports attempt to correlate the loss- and overexpression phenotype for collections of genes identified in gain-of-function screens. In this work we use RNA interference and in situ hybridization to annotate a collection of 123 P-GS insertions that in combination with different Gal4 drivers affect the size and/or patterning of the wing. We identify the gene causing the overexpression phenotype by expressing, in a background of overexpression, RNA interference for the genes affected by each P-GS insertion. Then, we compare the loss and gain-of-function phenotypes obtained for each gene and relate them to its expression pattern in the wing disc. We find that 52% of genes identified by their overexpression phenotype are required during normal development. However, only in 9% of the cases analyzed was there some complementarity between the gain- and loss-of-function phenotype, suggesting that, in general, the overexpression phenotypes would not be indicative of the normal requirements of the gene.

The Notch signaling pathway plays an important role in development and physiology. In Drosophila, Notch is activated by its Delta or Serrate ligands, depending in part on the sugar modifications present in its extracellular domain. O-fucosyltransferase-1 (OFUT1) performs the first glycosylation step in this process, O-fucosylating various EGF repeats at the Notch extracellular domain. Besides its O-fucosyltransferase activity, OFUT1 also behaves as a chaperone during Notch synthesis and is able to down regulate Notch by enhancing its endocytosis and degradation. We have reevaluated the roles that O-fucosylation and the synthesis of GDP-fucose play in the regulation of Notch protein stability. Using mutants and the UAS/Gal4 system, we modified in developing tissues the amount of GDP-mannose-deshydratase (GMD), the first enzyme in the synthesis of GDP-fucose. Our results show that GMD activity, and likely the levels of GDP-fucose and O-fucosylation, are essential to stabilize the Notch protein. Notch degradation observed under low GMD expression is absolutely dependent on OFUT1 and this is also observed in Notch Abruptex mutants, which have mutations in some potential O-fucosylated EGF domains. We propose that the GDP-fucose/OFUT1 balance determines the ability of OFUT1 to endocytose and degrade Notch in a manner that is independent of the residues affected by Abruptex mutations in Notch EGF domains.

The development of the Drosophila melanogaster wing depends on the correct regulation of cell survival, growth, proliferation, differentiation, and pattern formation. These processes, and the genes controlling then, are common to the development of epithelia in many different organisms. To identify additional genes contributing to wing development we have carried out a genetic screen in mosaic wings carrying clones of homozygous mutant cells. We obtained 12 complementation groups corresponding to genes with a proven role in wing formation such as smoothened, thick veins, mothers against dpp, expanded, and fat and 71 new complementation groups affecting the pattern of veins and the size of wing. We mapped one of these groups to the mediator15 gene (med15), a component of the Mediator complex. We show that Med15 and other members of the Mediator complex are required, among other processes, for the transcription of decapentaplegic target genes.

The formation of the Drosophila wing involves developmental processes such as cell proliferation, pattern formation, and cell differentiation that are common to all multicellular organisms. The genes controlling these cellular behaviors are conserved throughout the animal kingdom, and the genetic analysis of wing development has been instrumental in their identification and functional characterization. The wing is a postembryonic structure, and most loss-of-function mutations are lethal in homozygous flies before metamorphosis. In this manner, loss-of-function genetic screens aiming to identify genes affecting wing formation have not been systematically utilized. As an alternative, a number of genetic searches have utilized the phenotypic consequences of gene gain-of-expression, as a method more efficient to search for genes required during imaginal development. Here we present the results of a gain-of-function screen designed to identify genes involved in the formation of the wing veins. We generated 13,000 P-GS insertions of a P element containing UAS sequences (P-GS) and combined them with a Gal4 driver expressed mainly in the developing pupal veins. We selected 500 P-GSs that, in combination with the Gal4 driver, result in modifications of the veins, changes in the morphology of the wing, or defects in the differentiation of the trichomes. The P-element insertion sites were mapped to the genomic sequence, identifying 373 gene candidates to participate in wing morphogenesis and vein formation.