A key aim of neurodevelopmental research is to understand how precursor cells decide to stop dividing and commence their terminal differentiation at the correct time and place. Here, we show that minibrain (mnb), the Drosophila ortholog of the Down syndrome candidate gene DYRK1A, is transiently expressed in newborn neuronal precursors known as ganglion cells (GCs). Mnb promotes the cell cycle exit of GCs through a dual mechanism that regulates the expression of the cyclin-dependent kinase inhibitor Dacapo, the homolog of vertebrate p27Kip1 (Cdkn1b). Mnb upregulates the expression of the proneural transcription factor (TF) Asense, which promotes Dacapo expression. Mnb also induces the expression of Prospero, a homeodomain TF that in turn inhibits the expression of Deadpan, a pan-neural TF that represses dacapo. In addition to its effects on Asense and Prospero, Mnb also promotes the expression of the neuronal-specific RNA regulator Elav, strongly suggesting that Mnb facilitates neuronal differentiation. These actions of Mnb ensure the precise timing of neuronal birth, coupling the mechanisms that regulate neurogenesis, cell cycle control and terminal differentiation of neurons.

The Down syndrome and microcephaly related gene Mnb/Dyrk1A encodes an evolutionary conserved protein kinase subfamily that plays important roles in neurodevelopment. minibrain (mnb) mutants of Drosophila melanogaster (Dm) exhibit reduced adult brains due to neuronal deficits generated during larval development. These deficits are the consequence of the apoptotic cell death of numerous neuronal precursors that fail to properly exit the cell cycle and differentiate. We have recently found that in both the Dm larval brain and the embryonic vertebrate central nervous system (CNS), a transient expression of Mnb/Dyrk1A promotes the cell cycle exit of newborn neuronal precursors by upregulating the expression of the cyclin-dependent kinase inhibitor p27kip1 (called Dacapo in Dm). In the larval brain, Mnb performs this action by regulating the expression of three transcription factors, Asense (Ase), Deadpan (Dpn) and Prospero (Pros), which are key regulators of the self-renewal, proliferation, and terminal differentiation of neural progenitor cells. We have here studied in detail the cellular/temporal expression pattern of Ase, Dpn, Pros and Mnb, and have analyzed possible regulatory effects among them at the transitions from neurogenic progenitors to postmitotic neuronal precursors in the Dm larval brain. The emerging picture of this analysis reveals an intricate regulatory network in which Mnb appears to play a pivotal role helping to delineate the dynamics of the expression patterns of Ase, Dpn and Pros, as well as their specific functions in the aforementioned transitions. Our results also show that Ase, Dpn and Pros perform several cross-regulatory actions and contribute to shape the precise cellular/temporal expression pattern of Mnb. We propose that Mnb/Dyrk1A plays a central role in CNS neurogenesis by integrating molecular mechanisms that regulate progenitor self-renewal, cell cycle progression and neuronal differentiation.

In the developing Drosophila optic lobe, neuroepithelial (NE) cells are transformed progressively into neurogenic progenitors called neuroblasts (NBs). The progenitors undergoing this transition are identified by the expression of the Acheate Scute Complex (AS-C) factor Lethal of Scute (L'sc). Here we found that Asense (Ase), another AS-C factor, presents a peak of expression in the cells neighboring those transition L'sc expressing cells. This peak of Ase identifies a new transition step and it is necessary and sufficient to promote the NE to NB transition. Thus, our data provide the first direct evidence for a proneural role of Ase in CNS neurogenesis. Furthermore, we found that the peak of Ase is induced in a non-cell autonomous manner by L'sc through the activation of Notch signaling in the adjacent cells. This suggests that the two classic proneural activities, promoting neurogenesis and Notch signaling, have been split between Ase and L'sc. Thus, our data fit with a model in which the key proneural role of Ase is integrated with Notch and L'sc activities, facilitating the progressive transformation of NE cells into NBs.