Abstract
The developing Drosophila brain is a well-studied model system for neurogenesis and stem cell biology. In the Drosophila central brain, around 200 neural stem cells called neuroblasts undergo repeated rounds of asymmetric cell division. These divisions typically generate a larger self-renewing neuroblast and a smaller ganglion mother cell that undergoes one terminal division to create two differentiating neurons. Although single mitotic divisions of neuroblasts can easily be imaged in real time, the lack of long term imaging procedures has limited the use of neuroblast live imaging for lineage analysis. Here we describe a method that allows live imaging of cultured Drosophila neuroblasts over multiple cell cycles for up to 24 hours. We describe a 4D image analysis protocol that can be used to extract cell cycle times and growth rates from the resulting movies in an automated manner. We use it to perform lineage analysis in type II neuroblasts where clonal analysis has indicated the presence of a transit-amplifying population that potentiates the number of neurons. Indeed, our experiments verify type II lineages and provide quantitative parameters for all cell types in those lineages. As defects in type II neuroblast lineages can result in brain tumor formation, our lineage analysis method will allow more detailed and quantitative analysis of tumorigenesis and asymmetric cell division in the Drosophila brain.
Highlights
The simplicity of the Drosophila central nervous system (CNS) and the variety of genetic tools to modify and monitor cell behavior make it an ideal system to study mechanisms of neurogenesis
NBs and intermediate neural progenitor (INP) establish an internal polarity axis, localizing self-renewing Par complex proteins atypical protein kinase C (aPKC), Par 6 and Baz to the apical domain (Figure 1B-G) and differentiating factors such as Mira to the basal domain to be inherited by the INP or GMC, respectively (Figure 1B-G) [9,10]
As in vivo, cultured type I, type II NBs and INPs asymmetrically distributed apical and basal polarity proteins to result in asymmetric progeny cell fates
Summary
The simplicity of the Drosophila central nervous system (CNS) and the variety of genetic tools to modify and monitor cell behavior make it an ideal system to study mechanisms of neurogenesis. Drosophila neuroblasts (NBs) divide asymmetrically to generate one larger self-renewing and a smaller cell that differentiates into neurons and glia after a limited number of transit amplifying divisions [1]. Larval NBs generate an intrinsic axis of polarity by localizing apical and basal polarity proteins on opposite sides of the cell cortex. The cell fate determinants Numb, Prospero (Pros) and Brain tumor (Brat) localize to the opposite side at the basal cortex and, through binding to mediator proteins such as Miranda (Mira) and Partner-of-Numb (Pon), segregate into the differentiating daughter cell [11,12]. Once inherited by the GMC, Numb, Pros and Brat inhibit self-renewal and promote cell cycle exit and differentiation [13,14,15,16]
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
