Analysis of the "engrailed"-expressing neuroblast lineages in "Drosophila" brain development

The segment polarity gene engrailed (en) encodes a homeodomain transcription factor which is expressed in metamerically reiterated stripes in the embryonic neuroectoderm, in some primary neuroblasts and their progeny; usually located at the posterior boundary of each embryonic CNS neuromere. In this thesis, we have analyzed the expression of en in the larval and adult brains. In the late larval brain, four secondary neuroblast lineages (three protocerebral lineages and one deutocerebral lineage) express en in specific subsets. However, in the adult brain, only three of the four lineages express en.
In a first study (Chapter 2), we have characterized the neuropile innervation pattern of en-expressing central brain neuroblast lineages in embryonic, larval and adult stages. Firstly, based on en expression data and anatomical criteria, we are able to link primary lineages in the larva to secondary, adult-specific lineages. Secondly, the neurons of the en-expressing lineages form most arborizations, particularly their proximal branches, in the same brain neuropile compartments throughout development. Thirdly, the en-positive lineages of differing neuromeric origin and therefore, from different brain neuromeres innervate a non-overlapping set of neuropile compartments. Thus, the lineages appear to respect boundaries between neuromere-specific compartments in the brain and our findings support a model for neuromere-specific brain neuropile. Moreover, using genetic labeling techniques, this attempt is the first of its kind that links larval and adult brain anatomy at higher resolution.
In our second study (Chapter 3), we have analysed the total number and pattern of en-expressing, adult-specific cells in each of the four, identified larval neuroblast lineages mentioned above. Firstly, there are lineage-specific differences in number as well as expression of en in the four lineages examined. Secondly, this difference is established due to programmed cell death, which has a pronounced effect on the number of cells; approximately half of the immature adult-specific neurons in three of the four lineages are eliminated by cell death during development. Furthermore, programmed cell death selectively affects en-positive versus en-negative cells in a lineage-specific manner and, thus, controls the relative number of en-expressing neurons in each lineage. Our data provide evidence for a hemilineage-specific cell death model; i.e, either half of a lineage is targeted by programmed cell death. Further testing of this model by analysis of single and two cell clones in one of the four lineages also supports the proposed model. Finally, Notch signalling is involved in the regulation of en expression and consequently, is implicated to play a role in generation of the hemilineages. This study is the first of its kind to demonstrate the prominent role of lineage-specific programmed cell death in the generation of neuronal number and lineage diversity in the Drosophila postembryonic, central brain.
In conclusion, the use of en as a molecular marker has helped us characterise brain anatomy in greater detail. In addition, the analysis of the en-expressing neurons in the central brain has revealed a surprisingly predominant and lineage-specific role of programmed cell death in the control of neuronal number.