The bHLH Repressor Deadpan Regulates the Self-renewal and Specification of Drosophila Larval Neural Stem Cells Independently of Notch

Abstract

Neural stem cells (NSCs) are able to self-renew while giving rise to neurons and glia that comprise a functional nervous system. However, how NSC self-renewal is maintained is not well understood. Using the Drosophila larval NSCs called neuroblasts (NBs) as a model, we demonstrate that the Hairy and Enhancer-of-Split (Hes) family protein Deadpan (Dpn) plays important roles in NB self-renewal and specification. The loss of Dpn leads to the premature loss of NBs and truncated NB lineages, a process likely mediated by the homeobox protein Prospero (Pros). Conversely, ectopic/over-expression of Dpn promotes ectopic self-renewing divisions and maintains NB self-renewal into adulthood. In type II NBs, which generate transit amplifying intermediate neural progenitors (INPs) like mammalian NSCs, the loss of Dpn results in ectopic expression of type I NB markers Asense (Ase) and Pros before these type II NBs are lost at early larval stages. Our results also show that knockdown of Notch leads to ectopic Ase expression in type II NBs and the premature loss of type II NBs. Significantly, dpn expression is unchanged in these transformed NBs. Furthermore, the loss of Dpn does not inhibit the over-proliferation of type II NBs and immature INPs caused by over-expression of activated Notch. Our data suggest that Dpn plays important roles in maintaining NB self-renewal and specification of type II NBs in larval brains and that Dpn and Notch function independently in regulating type II NB proliferation and specification.

Rationally designed fluorogenic protease reporter visualizes spatiotemporal dynamics of apoptosis in vivo

“…5 D and E, and Fig. S4 C and D), consistent with previous Dpn overexpression studies (22)(23)(24)27). To examine the spatiotemporal dynamics of apoptosis during tumorigenesis of the larval brain, we imaged dpn OE and wild-type larval brains at different time points throughout larval development.…”

“…5A and Fig. S4) (22)(23)(24). Throughout neural development, the stem cells of Drosophia, known as neuroblasts (NBs), proliferate to generate the nervous system of both larval and adult stages via asymmetric division, whereby NBs self-renew and generate daughter cells (Fig.…”

“…5C) (25). Dpn is normally expressed in the NBs and intermediate neural progenitors (INPs) of the developing Drosophila brain and has roles in maintaining NB selfrenewal and specification of the type II NB identity in larval brains (22)(23)(24). We ectopically overexpressed Dpn (dpn OE ) using the insc-GAL4 (GAL4 1407 inserted in inscuteable promoter) driving expression in NBs, INPs, and GMCs (ganglion mother cells), which has been shown to lead to excessive NB proliferation at the expense of more differentiated cells, representing a tumorlike state (22)(23)(24).…”

Earmuff restricts progenitor cell potential by attenuating the competence to respond to self-renewal factors

“…Together, these data strongly suggest that Erm restricts developmental potential by antagonizing Dpn function. Importantly, although not affecting the maintenance of type II neuroblasts (Zacharioudaki et al, 2012;Zhu et al, 2012) (Fig. 5F), clonally removing the function of dpn strongly suppressed the supernumerary neuroblast phenotype in erm-null brains (Fig.…”

“…The neuroblast self-renewal factors include Dpn, Klumpfuss (Klu), Enhancer of split mγ [E(spl)mγ] and Notch (Weng et al, 2010;San-Juán and Baonza, 2011;Xiao et al, 2012;Zacharioudaki et al, 2012;Zhu et al, 2012). Removal of Notch function alone or dpn and E(spl)mγ function simultaneously leads to premature neuroblast differentiation, whereas overexpression of any of the neuroblast self-renewal factors in type II neuroblasts leads to massive formation of supernumerary neuroblasts.…”