Indeed, these receptors consist in a ligand-binding ectodomain linked via a trans-membrane domain to an intracellular domain that acts as a transcriptional regulator upon its ligand-dependent Cycloheximide release from the membrane. A ligand-dependent conformational change in the ectodomain of Notch is thought to result in ectodomain shedding and intra-membrane processing of Notch. Following the release of the Notch Intra-Cellular Domain, the activated nuclear form of Notch, NICD forms a ternary complex with CSL, a sequence-specific DNA-binding protein known as Suppressor of Hairlessin flies, and a co-activator, known as Mastermindin flies, to regulate the expression of Notch target genes. In the absence of NICD, CSL factors can bind the cis regulatory region and repress the expression of a subset of Notch target genes in both fliesand mammals. Indeed, the human CSL factor CBF1 was initially identified as a transcriptional repressorand several different CSL co-repressors have been identified in mammalian cells. NICD increases the occupancy of CSL binding sites, relieves the transcriptional repression mediated by CSL factors and promotes transcriptional activation. In Drosophila, repression by Su is critical to prevent Notch target genes from being inappropriately activated in some developmental contexts. Su acts in part by recruiting the adaptor protein Hairlessand its co-repressors CtBP and Groucho. While the activity of H appeared to be dispensable in most developmental contexts, including embryogenesis, repression by Su-H complexes is required for cell fate decisions during adult peripheral LDK378 neurogenesis. During pupal development, the activity of H is first required in imaginal tissues for the stable determination of Sensory Organ Precursor cells. SOP specification relies on Notchmediated lateral inhibition such that Notch target genes are repressed in SOPsand activated in surrounding cells. The de-repression of Notch target genes in H mutant SOPs was shown to prevent their stable determination. 
Following their specification, each SOP undergoes a stereotyped series of asymmetric cell divisions to generate the four different cells forming a sensory bristle. The activity of H is also required for proper cell fate determination in the bristle lineage. A reduced level of H in heterozygous or hypomorphic mutant flies led to the transformation of shaft into a second socket, hence resulting into double-socket bristles. Repression by Su-H complexes may act in parallel to other regulatory mechanisms to inhibit the expression of Notch target genes in SOPs. For instance, the transcriptional repressor Longitudinal lackingwas shown to repress the expression of Notch target genes, and to genetically interact with H during adult peripheral neurogenesis. Additionally, the nuclear BEN-solo family protein Insensitivewas recently shown to directly interact with Su and to inhibit in a Hindependent manner the expression of Notch target genes, both in embryos and in a cell-based assay. Our study identified Insb as a novel SOP/neuron-specific nuclear factor that antagonizes Notch to regulate cell fate. First, we have shown that over-expression of Insb inhibited the activity of Notch during sensory organ formation and blocked the expression of a Notch reporter construct in wing discs. This indicated that Insb has the ability to inhibit the expression of Notch target genes. Since the Notch reporter construct used here responded directly to Notch via paired Su binding sites, Insb likely acts via these binding sites, i.e. by modulating the activity of Su-bound complexes. Second, while the activity of insb appeared to be largely dispensable during development.