A Screen for Modifiers of Notch Signaling Uncovers Amun, a Protein With a Critical Role in Sensory Organ Development

Abstract

Notch signaling is an evolutionarily conserved pathway essential for many cell fate specification events during metazoan development. We conducted a large-scale transposon-based screen in the developing Drosophila eye to identify genes involved in Notch signaling. We screened 10,447 transposon lines from the Exelixis collection for modifiers of cell fate alterations caused by overexpression of the Notch ligand Delta and identified 170 distinct modifier lines that may affect up to 274 genes. These include genes known to function in Notch signaling, as well as a large group of characterized and uncharacterized genes that have not been implicated in Notch pathway function. We further analyze a gene that we have named Amun and show that it encodes a protein that localizes to the nucleus and contains a putative DNA glycosylase domain. Genetic and molecular analyses of Amun show that altered levels of Amun function interfere with cell fate specification during eye and sensory organ development. Overexpression of Amun decreases expression of the proneural transcription factor Achaete, and sensory organ loss caused by Amun overexpression can be rescued by coexpression of Achaete. Taken together, our data suggest that Amun acts as a transcriptional regulator that can affect cell fate specification by controlling Achaete levels.

A broad expression profile of the GMR-GAL4 driver in Drosophila melanogaster

“…We found the GMR>Dl flies not only show the eye phenotype as previously reported (Shalaby et al, 2009; data not shown), but also display a wing phenotype. In most cases, the distal part of vein L5 was lost in the GMR>Dl wings ( Figure 1C and D), as compared to the wild-type control ( Figure 1A and B).…”

Parallel Genetic and Proteomic Screens Identify Msps as a CLASP–Abl Pathway Interactor in Drosophila

“…While a number of screens have already made use of the Df collection (Bonds et al 2007) or the transposon insertion collection (Kankel et al 2007;Chang et al 2008;Shalaby et al 2009) individually, the logistics of obtaining and screening all of the 16,000 lines is not feasible for many labs. Thus, we applied a hierarchical strategy for our screen ( Figure 1A) that would take advantage of both collections and also would assay LOF vs. neomorphic effects.…”

“…The collection of deletions (Dfs; Parks et al 2004) was used for the primary screen, while the collection of transposon insertions (Thibault et al 2004) was utilized for the secondary screens. The use of these strains for genome-wide genetic interaction screening has been previously described (Kankel et al 2007;Chang et al 2008;Shalaby et al 2009). To confirm candidate interactors, all interacting transposons were crossed in at least two separate experiments to GMR-GAL4, UAS-CLASP-GFP and eye phenotypes were examined and imaged.…”

The developmental biology of genetic Notch disorders

“…Genetic and environmental factors thus likely shift the intrinsic duration or strength of Notch signaling, altering mutation tolerance in different individuals. Indeed, screens for modifiers of Notch-dependent phenotypes or signaling per se in Drosophila (Go and Artavanis-Tsakonas, 1998;Shalaby et al, 2009), mouse (Rubio-Aliaga et al, 2007 and in vitro (Mourikis et al, 2010) have revealed many candidate modifiers of Notchdependent disease. For example, loss of Itch (a negative regulator of Notch signaling) in mice interacts with gain of Notch1 in developing thymocytes to produce autoimmune disease, while loss of one allele of Dll3 in Notch1 heterozygous mice results in axial segmentation defects in 30% of double heterozygous mice (Loomes et al, 2007a).…”