Keywords: d. neuronal specification; c. neural stem cells
The Drosophila medulla, which is the largest neuropil in the optic lobe, is as an excellent model system in which to study the mechanisms that regulate neurogenesis. Its 40,000 neurons, which comprise over 90 cell types, are generated from a neuroepithelial crescent termed the outer proliferation center (OPC). Beginning at the onset of the third larval instar, and continuing for two days, a proneural wave converts neuroepithelial cells into neuroblasts (NBs), which subsequently divide asymmetrically to generate the neurons and glia of the medulla cortex. It has previously been shown that two axes of positional information act on OPC NBs to generate neural diversity. In the temporal axis, a cascade of five genes--Hth, Ey, Slp1, D and Tll--are sequentially expressed in each of the medulla NBs as they age. In the spatial axis, the OPC crescent from which the NBs are generated is sub-divided into eight compartments (patterned by four genes: Vsx1, Optix, Rx, and Hh). Distinct neuronal types are generated by NBs based on their spatio-temporal address. Here, we describe a third patterning axis that further diversifies neuronal fates in the medulla. We show that the neuroepithelium from which the NBs are generated is itself temporally patterned by the expression of five genes--Imp, Syp, Chinmo, Mamo and E93--over the two-day period of neurogenesis. This long-range temporal patterning of the neuroepithelium confers NBs from the same spatio-temporal address with unique identities based on the developmental time that they are generated. Using clonal- and EdU-birthdating analyses, we show that NBs in the Vsx1-Hth spatio-temporal window surprisingly generate not one, but four distinct neuronal types over the course of neurogenesis. These neurons (Tm23, TmY15, Pm3 and TmY12) are generated in distinct developmental windows and their specification is dependent on the activity of the long-range temporal factors; in syp mutants, early-born TmY15 neurons are expanded in number, whereas late-born Pm3 neurons are lost. We further show that long-range temporal patterning extends beyond the Hth temporal NB window to concurrently pattern NBs in the Ey, Slp1 and D temporal windows as well. Finally, we demonstrate that the birthdate of medulla neurons correlates with their final anterio-posterior position in the adult cortex and propose that long-range temporal patterning functions as a mechanism to couple neurogenesis with circuit assembly.