SummaryThe generation of neurons from neural stem cells requires large-scale changes in gene expression that are controlled to a large extent by proneural transcription factors, such as Ascl1. While recent studies have characterized the differentiation genes activated by proneural factors, less is known on the mechanisms that suppress progenitor cell identity. Here, we show that Ascl1 induces the transcription factor MyT1 while promoting neuronal differentiation. We combined functional studies of MyT1 during neurogenesis with the characterization of its transcriptional program. MyT1 binding is associated with repression of gene transcription in neural progenitor cells. It promotes neuronal differentiation by counteracting the inhibitory activity of Notch signaling at multiple levels, targeting the Notch1 receptor and many of its downstream targets. These include regulators of the neural progenitor program, such as Hes1, Sox2, Id3, and Olig1. Thus, Ascl1 suppresses Notch signaling cell-autonomously via MyT1, coupling neuronal differentiation with repression of the progenitor fate.

The pivotal mechanisms that govern the correct patterning and regionalization of the distinct areas of the mammalian CNS are driven by key molecules that emanate from the so-called secondary organizers at neural plate and tube stages. FGF8 is the candidate morphogenetic molecule to pattern the mesencephalon and rhombencephalon in the isthmic organizer (IsO). Recognizable relevance has been given to the intracellular pathways by which Fgf8 is regulated and modulated. In chick limb bud development, a dual mitogen-activated protein kinase phosphatase-3 (Mkp3) plays a role as a negative feedback modulator of Fgf8 signaling. We have investigated the role of Mkp3 and its functional relationship with the Fgf8 signaling pathway in the mouse IsO using gene transfer microelectroporation assays and protein-soaked bead experiments. Here, we demonstrate that MKP3 has a negative feedback action on the MAPK/ERK-mediated FGF8 pathway in the mouse neuroepithelium.

The present study evaluated the activity of jejunal Na+-K+-ATPase and its sensitivity to inhibition by dopamine in spontaneous hypertensive rats (SHR) and Wistar-Kyoto (WKY) rats during low (LS), normal (NS) and high (HS) salt intake. Basal jejunal Na+-K+-ATPase activity in SHR on LS intake was higher than in WKY rats. Jejunal Na+-K+-ATPase activity in WKY rats, but not in SHR, on LS intake was significantly reduced (20% decrease) by dopamine (1 microM) and SKF 38393 (10 nM), but not quinerolane (10 nM), this being antagonized the D1 receptor antagonist (SKF 83566). Changing from LS to NS or HS intake in WKY rats increased basal jejunal Na+-K+-ATPase activity and attenuated the inhibitory effect of dopamine. In SHR, changing from LS to NS or HS intake increased basal jejunal Na+-K+-ATPase activity. Basal renal Na+-K+-ATPase activity in SHR on LS intake was similar to that in WKY rats and was insensitive to inhibition by dopamine. Changing from LS to NS or HS intake in WKY rats increased basal renal Na+-K+-ATPase activity without affecting the inhibitory effect of dopamine. In SHR, changing from LS to NS or HS intake failed to alter basal renal Na+-K+-ATPase activity. It is concluded that inhibition of jejunal Na+-K+ ATPase activity by D1 dopamine receptor activation is dependent on salt intake in WKY rats, and SHR animals fail to respond to dopamine, irrespective of their salt intake.

BackgroundHemangioblasts are known as the common precursors for primitive hematopoietic and endothelial lineages. Their existence has been supported mainly by the observation that both cell types develop in close proximity and by in vitro differentiation and genetic studies. However, more compelling evidence will arise from tracking their cell fates using a lineage-specific marker.ResultsWe report the identification of a hemangioblast-specific enhancer (Hb) located in the cis-regulatory region of chick Cerberus gene (cCer) that is able to direct the expression of enhanced green fluorescent protein (eGFP) to the precursors of yolk sac blood and endothelial cells in electroporated chick embryos. Moreover, we present the Hb-eGFP reporter as a powerful live imaging tool for visualizing hemangioblast cell fate and blood island morphogenesis.ConclusionsWe hereby introduce the Hb enhancer as a valuable resource for genetically targeting the hemangioblast population as well as for studying the dynamics of vascular and blood cell development.

During mitosis, chromatin condensation is accompanied by a global arrest of transcription. Recent studies suggest transcriptional reactivation upon mitotic exit occurs in temporally coordinated waves, but the underlying regulatory principles have yet to be elucidated. In particular, the contribution of sequence-specific transcription factors (TFs) remains poorly understood. Here we report that Brn2, an important regulator of neural stem cell identity, associates with condensed chromatin throughout cell division, as assessed by live-cell imaging of proliferating neural stem cells. By contrast, the neuronal fate determinant Ascl1 dissociates from mitotic chromosomes. ChIP-seq analysis reveals that Brn2 mitotic-chromosome binding does not result in sequence-specific interactions prior to mitotic exit, relying mostly on electrostatic forces. Nevertheless, surveying active transcription using single-molecule RNA-FISH against immature transcripts, indicates the differential presence of TF near chromatin when exiting mitosis is associated with early (anaphase) versus late (early G1) reactivation of key targets of Brn2 and Ascl1, respectively. Moreover, by using a mitotic-specific dominant negative approach, we show that competing with Brn2 binding during mitotic exit reduces the transcription of its target gene Nestin. Our study shows an important role for differential binding of TFs to mitotic chromosomes, governed by their electrostatic properties, in defining the temporal order of transcriptional reactivation during mitosis-to-G1 transition.