Chromatin state changes during neural development revealed by in vivo cell-type specific profiling.

A key question in developmental biology is how cellular differentiation is controlled during development. While transitions between trithorax-group (TrxG) and polycomb-group (PcG) chromatin states are vital for the differentiation of ES cells to multipotent stem cells, little is known regarding the role of chromatin states during development of the brain. Here we show that large-scale chromatin remodelling occurs during Drosophila neural development. We demonstrate that the majority of genes activated during neuronal differentiation are silent in neural stem cells (NSCs) and occupy black chromatin and a TrxG-repressive state. In neurons, almost all key NSC genes are switched off via HP1-mediated repression. PcG-mediated repression does not play a significant role in regulating these genes, but instead regulates lineage-specific transcription factors that control spatial and temporal patterning in the brain. Combined, our data suggest that forms of chromatin other than canonical PcG/TrxG transitions take over key roles during neural development

damidseq_pipeline: an automated pipeline for processing DamID sequencing datasets.

UNLABELLED: DamID is a powerful technique for identifying regions of the genome bound by a DNA-binding (or DNA-associated) protein. Currently, no method exists for automatically processing next-generation sequencing DamID (DamID-seq) data, and the use of DamID-seq datasets with normalization based on read-counts alone can lead to high background and the loss of bound signal. DamID-seq thus presents novel challenges in terms of normalization and background minimization. We describe here damidseq_pipeline, a software pipeline that performs automatic normalization and background reduction on multiple DamID-seq FASTQ datasets. AVAILABILITY AND IMPLEMENTATION: Open-source and freely available from http://owenjm.github.io/damidseq_pipeline. The damidseq_pipeline is implemented in Perl and is compatible with any Unix-based operating system (e.g. Linux, Mac OSX). CONTACT: [email protected] SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.We thank Charles Bradshaw for helpful comments on the software. This work was supported by the BBSRC [BB/L00786X/1] and Wellcome Trust [092545]. The Gurdon Institute is supported by core funding from the Wellcome Trust [092096] and CRUK [C6946/A14492].This is the final published version. It first appeared at http://dx.doi.org/10.1093/bioinformatics/btv38

Mastermind Acts Downstream of Notch to Specify Neuronal Cell Fates in theDrosophilaCentral Nervous System

AbstractIn theDrosophilacentral nervous system, cellular diversity is generated through the asymmetric partitioning of cell fate determinants at cell division. Neural precursors (or neuroblasts) divide in a stem cell lineage to generate a series of ganglion mother cells, each of which divides once to produce a pair of postmitotic neurons or glial cells. An exception to this rule is the MP2 neuroblast, which divides only once to generate two neurons. We screened for genes expressed in the MP2 neuroblast and its progeny as a means of identifying the factors that specify cell fate in the MP2 lineage. We identified a P-element insertion line that expresses the reporter gene, tau-β-galactosidase, in the MP2 precursor and its progeny, the vMP2 and dMP2 neurons. The transposon disrupts the neurogenic gene,mastermind,but does not lead to neural hyperplasia. However, the vMP2 neuron is transformed into its sibling cell, dMP2. By contrast, expression of a dominant activated form of the Notch receptor in the MP2 lineage transforms dMP2 to vMP2. Notch signalling requires Mastermind, suggesting that Mastermind acts downstream of Notch to determine the vMP2 cell fate. We show that Mastermind plays a similar role in the neurons derived from ganglion mother cells 1-1a and 4-2a, where it specifies the pCC and RP2sib fates, respectively. This suggests that Notch signalling through Mastermind plays a wider role in specifying neuronal identity in theDrosophilacentral nervous system

Neural stem cell dynamics: the development of brain tumours.

Determining the premalignant lesions that develop into malignant tumours remains a daunting task. Brain tumours are frequently characterised by a block in differentiation, implying that normal developmental pathways become hijacked during tumourigenesis. However, the heterogeneity of stem cells and their progenitors in the brain suggests there are many potential routes to tumour initiation. Studies in Drosophila melanogaster have enhanced our understanding of the tumourigenic potential of distinct cell types in the brain. Here we review recent studies that have improved our knowledge of neural stem cell behaviour during development and in brain tumour models.This work was funded by: Wellcome Trust Senior Investigator Award (103792 to A.H.B.), The Royal Society Darwin Trust Research Professorship (to A.H.B.) and Wellcome Trust PhD Studentship (102454 to A.E.H.). A.H.B acknowledges core funding to The Gurdon Institute from the Wellcome Trust (092096) and CRUK (C6946/A14492)

A newly discovered neural stem cell population is generated by the optic lobe neuroepithelium during embryogenesis in Drosophila melanogaster.

Neural stem cells must balance symmetric and asymmetric cell divisions to generate a functioning brain of the correct size. In both the developing Drosophila visual system and mammalian cerebral cortex, symmetrically dividing neuroepithelial cells transform gradually into asymmetrically dividing progenitors that generate neurons and glia. As a result, it has been widely accepted that stem cells in these tissues switch from a symmetric, expansive phase of cell divisions to a later neurogenic phase of cell divisions. In the Drosophila optic lobe, this switch is thought to occur during larval development. However, we have found that neuroepithelial cells start to produce neuroblasts during embryonic development, demonstrating a much earlier role for neuroblasts in the developing visual system. These neuroblasts undergo neurogenic divisions, enter quiescence and are retained post-embryonically, together with neuroepithelial cells. Later in development, neuroepithelial cells undergo further cell divisions before transforming into larval neuroblasts. Our results demonstrate that the optic lobe neuroepithelium gives rise to neurons and glia over 60 h earlier than was thought previously.Royal Society Darwin Trust Research Professorship and 357 Wellcome Trust Senior Investigator Award 103792 to A.H.B. and Wellcome Trust PhD 358 Studentships 102454, to A.E.H., and 097423 to L.O. A.H.B acknowledges core funding to 359 The Gurdon Institute from the Wellcome Trust (092096) and CRUK (C6946/A14492

Neural stem cell transcriptional networks highlight genes essential for nervous system development

Neural stem cells must strike a balance between self-renewal and multipotency, and differentiation. Identification of the transcriptional networks regulating stem cell division is an essential step in understanding how this balance is achieved. We have shown that the homeodomain transcription factor, Prospero, acts to repress self-renewal and promote differentiation. Among its targets are three neural stem cell transcription factors, Asense, Deadpan and Snail, of which Asense and Deadpan are repressed by Prospero. Here, we identify the targets of these three factors throughout the genome. We find a large overlap in their target genes, and indeed with the targets of Prospero, with 245 genomic loci bound by all factors. Many of the genes have been implicated in vertebrate stem cell self-renewal, suggesting that this core set of genes is crucial in the switch between self-renewal and differentiation. We also show that multiply bound loci are enriched for genes previously linked to nervous system phenotypes, thereby providing a shortcut to identifying genes important for nervous system development

The vasculature as a neural stem cell niche.

Neural stem cells (NSCs) are multipotent, self-renewing progenitors that generate progeny that differentiate into neurons and glia. NSCs in the adult mammalian brain are generally quiescent. Environmental stimuli such as learning or exercise can activate quiescent NSCs, inducing them to proliferate and produce new neurons and glia. How are these behaviours coordinated? The neurovasculature, the circulatory system of the brain, is a key component of the NSC microenvironment, or 'niche'. Instructive signals from the neurovasculature direct NSC quiescence, proliferation, self-renewal and differentiation. During ageing, a breakdown in the niche accompanies NSC dysfunction and cognitive decline. There is much interest in reversing these changes and enhancing NSC activity by targeting the neurovasculature therapeutically. Here we discuss principles of neurovasculature-NSC crosstalk, and the implications for the design of NSC-based therapies. We also consider the emerging contributions to this field of the model organism Drosophila melanogaster.A.H.·B. is funded by Wellcome Trust Senior Investigator Award103792. L.O. is funded by Wellcome Trust PhD Studentship097423. A.H.B acknowledges core funding to the Gurdon Institute from the Wellcome Trust (092096) and CRUK (C6946/A14492)

The Serine Protease Homolog, Scarface, Is Sensitive to Nutrient Availability and Modulates the Development of the <i>Drosophila</i> Blood-Brain Barrier.

The adaptable transcriptional response to changes in food availability not only ensures animal survival but also lets embryonic development progress. Interestingly, the CNS is preferentially protected from periods of malnutrition, a phenomenon known as "brain sparing." However, the mechanisms that mediate this response remain poorly understood. To get a better understanding of this, we used Drosophila melanogaster as a model, analyzing the transcriptional response of neural stem cells (neuroblasts) and glia of the blood-brain barrier (BBB) from larvae of both sexes during nutrient restriction using targeted DamID. We found differentially expressed genes in both neuroblasts and glia of the BBB, although the effect of nutrient deficiency was primarily observed in the BBB. We characterized the function of a nutritional sensitive gene expressed in the BBB, the serine protease homolog, scarface (scaf). Scaf is expressed in subperineurial glia in the BBB in response to nutrition. Tissue-specific knockdown of scaf increases subperineurial glia endoreplication and proliferation of perineurial glia in the blood-brain barrier. Furthermore, neuroblast proliferation is diminished on scaf knockdown in subperineurial glia. Interestingly, reexpression of Scaf in subperineurial glia is able to enhance neuroblast proliferation and brain growth of animals in starvation. Finally, we show that loss of scaf in the blood-brain barrier increases sensitivity to drugs in adulthood, suggesting a physiological impairment. We propose that Scaf integrates the nutrient status to modulate the balance between neurogenesis and growth of the BBB, preserving the proper equilibrium between the size of the barrier and the brain.SIGNIFICANCE STATEMENT The Drosophila BBB separates the CNS from the open circulatory system. The BBB glia are not only acting as a physical segregation of tissues but participate in the regulation of the metabolism and neurogenesis during development. Here we analyze the transcriptional response of the BBB glia to nutrient deprivation during larval development, a condition in which protective mechanisms are switched on in the brain. Our findings show that the gene scarface reduces growth in the BBB while promoting the proliferation of neural stem, assuring the balanced growth of the larval brain. Thus, Scarface would link animal nutrition with brain development, coordinating neurogenesis with the growth of the BBB