EdU, 5-ethynyl-2’-deoxyuridine
(Thymidine analog incorporated into replicating DNA)
type II, GAL4 driver expressed so-called type II NSC lineages
To identify novel oncogenes, we are using the absence of significant proliferation in the adult Drosophila brain as a “blank slate”. Ectopic proliferation (detected for example by incorporation of replication labels such as EdU or BrdU) in the adult can come about by several mechanisms, including tumourigenesis. We confirm tumourigenesis by transplantation into WT host flies of brain tissue containing these ectopically proliferating cells labelled with a reporter such as Green Fluorescent Protein (GFP): tumour cells invade the host abdomen and can metastasize. Having identified a few neural oncogenes, we are now working on their mechanism of action.
There are two sides of the coin in tumour-microenvironment interactions. We are looking at both how a neural tumour model impacts surrounding non-tumour cells and, conversely, how the microenvironment impacts neural tumour growth. To best address this, we have developed a novel genetic tool that allows induction of labelled tumours of reproducible size, due to control over space (specific neural lineages) and time (high temporal resolution provided by heat-shock), in a single fly stock without a cross. Importantly, these tumours are generated in a GAL4-independent manner, reducing variability introduced by temperature-dependence of GAL4 activity and freeing-up the GAL4/UAS system for tumour microenvironment manipulations by crossing to available stock collections and assessing effect on tumour size.
Quiescence regulation
flflN42, null allele of the gene encoding the regulatory subunit of protein phosphatase 4 falafel, which induces NSC cell-cycle delay/arrest but not the characteristic morphology of quiescent NSCs
Some brain tumours remain dormant (stable in size and location) for long periods whereas others are fatal within months. The cause for this disparity is unknown. We hypothesise that progenitor cell quiescence (reversible cell-cycle exit accompanied by low metabolism) underlies tumour dormancy. Quiescence protects cells from antiproliferative chemotherapy and quiescent medulloblastoma and leukaemia cells can cause disease relapse. Therefore, detection of quiescent cells in tumour resections may be of prognostic value, and understanding quiescence regulation should suggest therapeutic alternatives. No molecular markers exist for quiescence, its identification relying on functional assays (failure to proliferate and retention of EdU or BrdU). In a forward-genetic screen, we have identified a Drosophila mutant (2V327) in which neural stem cells (NSCs) anachronically re-enter quiescence at late larval stages (NSC quiescence is identified by characteristic morphology, with cells extending a fiber of unknown function, accompanying cell-cycle arrest). We have mapped this mutation and are using Drosophila in vivo and a mouse NSC in vitro models to pursue novel hypotheses concerning quiescence regulation.
