As such, attempts to design therapeutics based on these prevalent functions should take into account the effects on Notch signaling, since the relationship between Notch sig naling and carcinogenesis is context dependent. Conclusions Awd belongs to the Nm23 family of protein that is evolu tionarily sellectchem conserved from Drosophila to mammals. Our in vivo analyses demonstrate that loss of awd gene function blocks Notch signaling by altering the receptor processing after the S2 cleavage and causes Notch accumulation in early endosomes. Furthermore, we obtained evidence indi cating that Awd is required for Rab5 function in early en dosome formation. Nm23 has been an enigmatic gene function. It is a house keeping gene involved in nucleotide synthesis and energy metabolism, and yet exhibiting Inhibitors,Modulators,Libraries specific developmental func tions.

It was the first metastasis suppressor gene identified, yet exhibits oncogenic functions in some cancer cohorts. We have previously shown that ei ther loss of function or over expression Inhibitors,Modulators,Libraries of awd can affect different aspects of epithelial morphogenesis. That is, loss of function awd results in over accumulation of adherens junction components and piling up of the epithelium, while over expression of awd results in reduced adherens junc tions and disintegration of epithelial structure. These findings provided some explanation of the biphasic function of Nm23 in tumorigenesis. In light of the studies presented here, an additional level of complexity should be considered since Notch signaling can exert different cellular functions in different tissues and at different times during patho physiological alterations of the same tissues.

Methods Drosophila Inhibitors,Modulators,Libraries strains and genetics Stocks were raised on standard cornmeal yeast agar medium at 25 C. The stock carrying the protein null awd allele, awdj2A4, has been described. The Inhibitors,Modulators,Libraries awdj2A4 al lele combined with the FRT chromosome FRT82B has been described. Cell clones mutant for awdj2A4 were gen erated through mitotic recombination using the FLP FRT system, either with Inhibitors,Modulators,Libraries the hs flp recombinase transgene or using the directed mosaic technique with the UAS flp transgene under control of the ubiquitous somatic cell driver en2. 4 Gal4e22c. To obtain over expression of specific transgenes in awdj2A4 mutant follicle cells we used either the directed mosaics or the MARCM tech niques.

The transgenic line carrying the constitutively active variant of the YFP Rab5 fusion genes was obtained from the Bloomington Stock Center. The UAS NICD and the GbeSu m8 lacZ lines were a kind gift from S. Bray of University Dorsomorphin of Cambridge. The UAS NEXT line was a kind gift from M. Fortini of Thomas Jefferson University. The genotypes of flies and larvae used for the analyses are described in Additional file 10, Supplementary experimental procedures.