Developmentally regulated subnuclear genome reorganization restricts neural progenitor competence in Drosophila.

Stem and/or progenitor cells often generate distinct cell types in a stereotyped birth order and over time lose competence to specify earlier-born fates by unknown mechanisms. In Drosophila, the Hunchback transcription factor acts in neural progenitors (neuroblasts) to specify early-born neurons, in part by indirectly inducing the neuronal transcription of its target genes, including the hunchback gene. We used in vivo immuno-DNA FISH and found that the hunchback gene moves to the neuroblast nuclear periphery, a repressive subnuclear compartment, precisely when competence to specify early-born fate is lost and several hours and cell divisions after termination of its transcription. hunchback movement to the lamina correlated with downregulation of the neuroblast nuclear protein, Distal antenna (Dan). Either prolonging Dan expression or disrupting lamina interfered with hunchback repositioning and extended neuroblast competence. We propose that neuroblasts undergo a developmentally regulated subnuclear genome reorganization to permanently silence Hunchback target genes that results in loss of progenitor competence.

3D Carbon Fiber Skeleton Film Modified with Gradient Cu Nanoparticles as Auxiliary Anode Regulates Dendrite-Free, Bottom-Up Zinc Deposition.

Achieving stable Zn plating/stripping under high current density and large area capacity remains a major challenge for metal Zn anodes. To address this issue, common filter paper is utilized to construct 3D carbon fiber skeleton film modified with gradient Cu nanoparticles (CFF@Cu). The original zincophobic hydrophilic CFF is transformed into gradient zincophilic and reversed gradient hydrophilic composite, due to the gradient distribution of Cu nanoparticles. When CFF@Cu is placed above Zn foil as an auxiliary anode, Zn foil anode exhibits stable, reversible, and dendrite-free Zn plating/stripping for 1200 h at 10 mA cm-2 and 2 mAh cm-2, 2000 h at 2 mA cm-2 and 2 mAh cm-2, 340 h at 10 mA cm-2 and 10 mAh cm-2. Additionally, nucleation barrier of Zn, Zn2+ transport and deposition kinetics are improved. The deposits on the Zn foil anode become homogeneous, dense, and fine. Side reactions and by-products are effectively inhibited. The excellent performance is mainly attributed to the gradient zincophilic field in 3D CFF. A portion of Zn2+ is captured by Cu and deposited within CFF@Cu from bottom to top, which reduces and homogenizes Zn2+ flux on Zn foil, as well as weakens and homogenizes electric field on Zn foil.

Transcriptional profiling from whole embryos to single neuroblast lineages in Drosophila.

Embryonic development results in the production of distinct tissue types, and different cell types within each tissue. A major goal of developmental biology is to uncover the "parts list" of cell types that comprise each organ. Here we perform single cell RNA sequencing (scRNA-seq) of the Drosophila embryo to identify the genes that characterize different cell and tissue types during development. We assay three different timepoints, revealing a coordinated change in gene expression within each tissue. Interestingly, we find that the elav and Mhc genes, whose protein products are widely used as markers for neurons and muscles, respectively, show broad pan-embryonic expression, indicating the importance of post-transcriptional regulation. We next focus on the central nervous system (CNS), where we identify genes whose expression is enriched at each stage of neuronal differentiation: from neural progenitors, called neuroblasts, to their immediate progeny ganglion mother cells (GMCs), followed by new-born neurons, young neurons, and the most mature neurons. Finally, we ask whether the clonal progeny of a single neuroblast (NB7-1) share a similar transcriptional identity. Surprisingly, we find that clonal identity does not lead to transcriptional clustering, showing that neurons within a lineage are diverse, and that neurons with a similar transcriptional profile (e.g. motor neurons, glia) are distributed among multiple neuroblast lineages. Although each lineage consists of diverse progeny, we were able to identify a previously uncharacterized gene, Fer3, as an excellent marker for the NB7-1 lineage. Within the NB7-1 lineage, neurons which share a temporal identity (e.g. Hunchback, Kruppel, Pdm, and Castor temporal transcription factors in the NB7-1 lineage) have shared transcriptional features, allowing for the identification of candidate novel temporal factors or targets of the temporal transcription factors. In conclusion, we have characterized the embryonic transcriptome for all major tissue types and for three stages of development, as well as the first transcriptomic analysis of a single, identified neuroblast lineage, finding a lineage-enriched transcription factor.

Drosophila nucleostemin 3 is required to maintain larval neuroblast proliferation.

Stem cells must maintain proliferation during tissue development, repair and homeostasis, yet avoid tumor formation. In Drosophila, neural stem cells (neuroblasts) maintain proliferation during embryonic and larval development and terminate cell cycle during metamorphosis. An important question for understanding how tissues are generated and maintained is: what regulates stem cell proliferation versus differentiation? We performed a genetic screen which identified nucleostemin 3 (ns3) as a gene required to maintain neuroblast proliferation. ns3 is evolutionarily conserved with yeast and human Lsg1, which encode putative GTPases and are essential for organism growth and viability. We found NS3 is cytoplasmic and it is required to retain the cell cycle repressor Prospero in neuroblast cytoplasm via a Ran-independent pathway. NS3 is also required for proper neuroblast cell polarity and asymmetric cell division. Structure-function analysis further shows that the GTP-binding domain and acidic domain are required for NS3 function in neuroblast proliferation. We conclude NS3 has novel roles in regulating neuroblast cell polarity and proliferation.

High-Performance Na-O2 Batteries Enabled by Oriented NaO2 Nanowires as Discharge Products.

Na-O2 batteries are emerging rechargeable batteries due to their high theoretical energy density and abundant resources, but they suffer from sluggish kinetics due to the formation of large-size discharge products with cubic or irregular particle shapes. Here, we report the unique growth of discharge products of NaO2 nanowires inside Na-O2 batteries that significantly boosts the performance of Na-O2 batteries. For this purpose, a high-spin Co3O4 electrocatalyst was synthesized via the high-temperature oxidation of pure cobalt nanoparticles in an external magnetic field. The discharge products of NaO2 nanowires are 10-20 nm in diameter and ∼10 µm in length, characteristics that provide facile pathways for electron and ion transfer. With these nanowires, Na-O2 batteries have surpassed 400 cycles with a fixed capacity of 1000 mA h g-1, an ultra-low over-potential of ∼60 mV during charging, and near-zero over-potential during discharging. This strategy not only provides a unique way to control the morphology of discharge products to achieve high-performance Na-O2 batteries but also opens up the opportunity to explore growing nanowires in novel conditions.

Lineage-specific effects of Notch/Numb signaling in post-embryonic development of the Drosophila brain.

Numb can antagonize Notch signaling to diversify the fates of sister cells. We report here that paired sister cells acquire different fates in all three Drosophila neuronal lineages that make diverse types of antennal lobe projection neurons (PNs). Only one in each pair of postmitotic neurons survives into the adult stage in both anterodorsal (ad) and ventral (v) PN lineages. Notably, Notch signaling specifies the PN fate in the vPN lineage but promotes programmed cell death in the missing siblings in the adPN lineage. In addition, Notch/Numb-mediated binary sibling fates underlie the production of PNs and local interneurons from common precursors in the lAL lineage. Furthermore, Numb is needed in the lateral but not adPN or vPN lineages to prevent the appearance of ectopic neuroblasts and to ensure proper self-renewal of neural progenitors. These lineage-specific outputs of Notch/Numb signaling show that a universal mechanism of binary fate decision can be utilized to govern diverse neural sibling differentiations.

High frequency characteristics of Fe65Co35 alloy cluster-assembled films prepared by energetic cluster deposition.

Size-monodispersed Fe65Co35 alloy clusters whose average sizes ranged from 7 to 12 nm were produced by a plasma-gas-condensation (PGC)-type cluster deposition apparatus. Fe65Co35 alloy cluster-assembled films were further prepared at room temperature by energetic cluster deposition method. Positively charged clusters in a cluster beam were accelerated electrically and deposited onto a negatively biased substrate together with neutral clusters from the same cluster source, leading to the formation of a high-density Fe65Co35 alloy cluster-assembled films with good soft magnetic properties. High frequency magnetic characteristics of these films were studied at room temperature using a high-frequency permeameter (RMF-3000, Ryowa). The real part (micro') of permeability for the Fe65Co35 alloy cluster-assembled films prepared at bias voltage V(a) = -20 kV has a large value of micro' = 135 at f = 1.5 GHz, and imaginary part (micro") of permeability has a maximum (micro" approximately 190) at about 2.5 GHz.

A developmental framework linking neurogenesis and circuit formation in the Drosophila CNS.

The mechanisms specifying neuronal diversity are well characterized, yet it remains unclear how or if these mechanisms regulate neural circuit assembly. To address this, we mapped the developmental origin of 160 interneurons from seven bilateral neural progenitors (neuroblasts) and identify them in a synapse-scale TEM reconstruction of the Drosophila larval central nervous system. We find that lineages concurrently build the sensory and motor neuropils by generating sensory and motor hemilineages in a Notch-dependent manner. Neurons in a hemilineage share common synaptic targeting within the neuropil, which is further refined based on neuronal temporal identity. Connectome analysis shows that hemilineage-temporal cohorts share common connectivity. Finally, we show that proximity alone cannot explain the observed connectivity structure, suggesting hemilineage/temporal identity confers an added layer of specificity. Thus, we demonstrate that the mechanisms specifying neuronal diversity also govern circuit formation and function, and that these principles are broadly applicable throughout the nervous system.

Genetic mosaic with dual binary transcriptional systems in Drosophila.

MARCM (mosaic analysis with a repressible cell marker) involves specific labeling of GAL80-minus and GAL4-positive homozygous cells in otherwise heterozygous tissues. Here we demonstrate how the concurrent use of two independent binary transcriptional systems may facilitate complex MARCM studies in the Drosophila nervous system. By fusing LexA with the VP16 acidic activation domain (VP16) or the GAL4 activation domain (GAD), we obtained both GAL80-insensitive and GAL80-suppressible transcriptional factors. LexA::VP16 can mediate MARCM-independent binary transgene induction in mosaic organisms. The incorporation of LexA::GAD into MARCM, which we call dual-expression-control MARCM, permits the induction of distinct transgenes in different patterns among GAL80-minus cells in mosaic tissues. Lineage analysis with dual-expression-control MARCM suggested the presence of neuroglioblasts in the developing optic lobes but did not indicate the production of glia by postembryonic mushroom body neuronal precursors. In addition, dual-expression-control MARCM with a ubiquitous LexA::GAD driver revealed many unidentified cells in the GAL4-GH146-positive projection neuron lineages.

A novel temporal identity window generates alternating Eve+/Nkx6+ motor neuron subtypes in a single progenitor lineage.

BACKGROUND: Spatial patterning specifies neural progenitor identity, with further diversity generated by temporal patterning within individual progenitor lineages. In vertebrates, these mechanisms generate "cardinal classes" of neurons that share a transcription factor identity and common morphology. In Drosophila, two cardinal classes are Even-skipped (Eve)+ motor neurons projecting to dorsal longitudinal muscles, and Nkx6+ motor neurons projecting to ventral oblique muscles. Cross-repressive interactions prevent stable double-positive motor neurons. The Drosophila neuroblast 7-1 (NB7-1) lineage uses a temporal transcription factor cascade to generate five distinct Eve+ motor neurons; the origin and development of Nkx6+ motor neurons remains unclear. METHODS: We use a neuroblast specific Gal4 line, sparse labelling and molecular markers to identify an Nkx6+ VO motor neuron produced by the NB7-1 lineage. We use lineage analysis to birth-date the VO motor neuron to the Kr+ Pdm+ neuroblast temporal identity window. We use gain- and loss-of-function strategies to test the role of Kr+ Pdm+ temporal identity and the Nkx6 transcription factor in specifying VO neuron identity. RESULTS: Lineage analysis identifies an Nkx6+ neuron born from the Kr+ Pdm+ temporal identity window in the NB7-1 lineage, resulting in alternation of cardinal motor neuron subtypes within this lineage (Eve>Nkx6 > Eve). Co-overexpression of Kr/Pdm generates ectopic VO motor neurons within the NB7-1 lineage - the first evidence that this TTF combination specifies neuronal identity. Moreover, the Kr/Pdm combination promotes Nkx6 expression, which itself is necessary and sufficient for motor neuron targeting to ventral oblique muscles, thereby revealing a molecular specification pathway from temporal patterning to cardinal transcription factor expression to motor neuron target selection. CONCLUSIONS: We show that one neuroblast lineage generates interleaved cardinal motor neurons fates; that the Kr/Pdm TTFs form a novel temporal identity window that promotes expression of Nkx6; and that the Kr/Pdm > Nkx6 pathway is necessary and sufficient to promote VO motor neuron targeting to the correct ventral muscle group.

Organization and postembryonic development of glial cells in the adult central brain of Drosophila.

Glial cells exist throughout the nervous system, and play essential roles in various aspects of neural development and function. Distinct types of glia may govern diverse glial functions. To determine the roles of glia requires systematic characterization of glia diversity and development. In the adult Drosophila central brain, we identify five different types of glia based on its location, morphology, marker expression, and development. Perineurial and subperineurial glia reside in two separate single-cell layers on the brain surface, cortex glia form a glial mesh in the brain cortex where neuronal cell bodies reside, while ensheathing and astrocyte-like glia enwrap and infiltrate into neuropils, respectively. Clonal analysis reveals that distinct glial types derive from different precursors, and that most adult perineurial, ensheathing, and astrocyte-like glia are produced after embryogenesis. Notably, perineurial glial cells are made locally on the brain surface without the involvement of gcm (glial cell missing). In contrast, the widespread ensheathing and astrocyte-like glia derive from specific brain regions in a gcm-dependent manner. This study documents glia diversity in the adult fly brain and demonstrates involvement of different developmental programs in the derivation of distinct types of glia. It lays an essential foundation for studying glia development and function in the Drosophila brain.

A repressor-decay timer for robust temporal patterning in embryonic Drosophila neuroblast lineages.

Biological timers synchronize patterning processes during embryonic development. In the Drosophila embryo, neural progenitors (neuroblasts; NBs) produce a sequence of unique neurons whose identities depend on the sequential expression of temporal transcription factors (TTFs). The stereotypy and precision of NB lineages indicate reproducible TTF timer progression. We combine theory and experiments to define the timer mechanism. The TTF timer is commonly described as a relay of activators, but its regulatory circuit is also consistent with a repressor-decay timer, where TTF expression begins when its repressor decays. Theory shows that repressor-decay timers are more robust to parameter variations than activator-relay timers. This motivated us to experimentally compare the relative importance of the relay and decay interactions in vivo. Comparing WT and mutant NBs at high temporal resolution, we show that the TTF sequence progresses primarily by repressor-decay. We suggest that need for robust performance shapes the evolutionary-selected designs of biological circuits.

Characterizing ionic species in PM2.5 and PM10 in four Pearl River Delta cities, south China.

PM2.5 and PM10 samples were collected at four major cities in the Pearl River Delta (PRD), South China, during winter and summer in 2002. Six water-soluble ions, Na+, NH4+, K+, Cl-, NO3- and SO4(2-) were measured using ion chromatography. On average, ionic species accounted for 53.3% and 40.5% for PM2.5 and PM10, respectively in winter and 39.4% and 35.2%, respectively in summer. Secondary ions such as sulfate, nitrate and ammonium accounted for the major part of the total ionic species. Sulfate was the most abundant species followed by nitrate. Overall, a regional pollution tendency was shown that there were higher concentrations of sulfate, nitrate and ammonium in Guangzhou City than those in the other PRD cities. Significant seasonal variations were also observed with higher levels of species in winter but lower in summer. The Asian monsoon system was favorable for removal and diffusion of air pollutants in PRD in summer while highly loading of local industrial emissions tended to deteriorate the air quality as well. NO3-/SO4(2-) ratio indicated that mobile sources have considerably contribution to the urban aerosol, and stationary sources should not be neglected. Besides the primary emissions, complex atmospheric reactions under favorable weather conditions should be paid more attention for the control of primary emission in the PRD region.

Enhanced Microwave Absorption Properties by Tuning Cation Deficiency of Perovskite Oxides of Two-Dimensional LaFeO3/C Composite in X-Band.

Development of microwave absorption materials with tunable thickness and bandwidth is particularly urgent for practical applications but remains a great challenge. Here, two-dimensional nanocomposites consisting of perovskite oxides (LaFeO3) and amorphous carbon were successfully obtained through a one pot with heating treatment using sodium chloride as a hard template. The tunable absorption properties were realized by introducing A-site cation deficiency in LaFeO3 perovskite. Among the A-site cation-deficient perovskites, La0.62FeO3/C (L0.62FOC) has the best microwave absorption properties in which the maximum absorption is -26.6 dB at 9.8 GHz with a thickness of 2.94 mm and the bandwidth range almost covers all X-band. The main reason affecting the microwave absorption performance was derived from the A-site cation deficiency which induced more dipoles polarization loss. This work proposes a promising method to tune the microwave absorption performance via introducing deficiency in a crystal lattice.

Automatic 3-D grayscale volume matching and shape analysis.

Recently, shape matching in three dimensions (3-D) has been gaining importance in a wide variety of fields such as computer graphics, computer vision, medicine, and biology, with applications such as object recognition, medical diagnosis, and quantitative morphological analysis of biological operations. Automatic shape matching techniques developed in the field of computer graphics handle object surfaces, but ignore intensities of inner voxels. In biology and medical imaging, voxel intensities obtained by computed tomography (CT), magnetic resonance imagery (MRI), and confocal microscopes are important to determine point correspondences. Nevertheless, most biomedical volume matching techniques require human interactions, and automatic methods assume matched objects to have very similar shapes so as to avoid combinatorial explosions of point. This article is aimed at decreasing the gap between the two fields. The proposed method automatically finds dense point correspondences between two grayscale volumes; i.e., finds a correspondent in the second volume for every voxel in the first volume, based on the voxel intensities. Mutiresolutional pyramids are introduced to reduce computational load and handle highly plastic objects. We calculate the average shape of a set of similar objects and give a measure of plasticity to compare them. Matching results can also be used to generate intermediate volumes for morphing. We use various data to validate the effectiveness of our method: we calculate the average shape and plasticity of a set of fly brain cells, and we also match a human skull and an orangutan skull.

miR-124 modulates gefitinib resistance through SNAI2 and STAT3 in non-small cell lung cancer.

Gefitinib is used as a first-line treatment for advanced non-small cell lung cancer (NSCLC). Unfortunately, most NSCLC patients inevitably develop gefitinib resistance during treatment. In addition to EGFR mutation status, the mechanisms involved are largely unknown. In this study, we showed that miR-124, a tumor suppressor, was significantly down-regulated in gefitinib-resistant NSCLC patients and cell lines compared with gefitinib-sensitive patients and cell lines. In addition, the miR-124 depletion induced gefitinib resistance, and miR-124 overexpression sensitized gefitinib-resistant cells to gefitinib. Mechanistic analysis revealed that miR-124 decreased SNAI2 and STAT3 expression by directly targeting their 3'UTRs and that knocking down SNAI2 or STAT3 partly reversed the gefitinib resistance induced by miR-124 depletion. Our data demonstrate that the miR-124 plays a new critical role in acquired resistance to gefitinib and that the manipulation of miR-124 might provide a therapeutic strategy for reversing acquired gefitinib resistance.

Transient nuclear Prospero induces neural progenitor quiescence.

Stem cells can self-renew, differentiate, or enter quiescence. Understanding how stem cells switch between these states is highly relevant for stem cell-based therapeutics. Drosophila neural progenitors (neuroblasts) have been an excellent model for studying self-renewal and differentiation, but quiescence remains poorly understood. In this study, we show that when neuroblasts enter quiescence, the differentiation factor Prospero is transiently detected in the neuroblast nucleus, followed by the establishment of a unique molecular profile lacking most progenitor and differentiation markers. The pulse of low level nuclear Prospero precedes entry into neuroblast quiescence even when the timing of quiescence is advanced or delayed by changing temporal identity factors. Furthermore, loss of Prospero prevents entry into quiescence, whereas a pulse of low level nuclear Prospero can drive proliferating larval neuroblasts into quiescence. We propose that Prospero levels distinguish three progenitor fates: absent for self-renewal, low for quiescence, and high for differentiation.

The Snail family member Worniu is continuously required in neuroblasts to prevent Elav-induced premature differentiation.

Snail family transcription factors are best known for regulating epithelial-mesenchymal transition (EMT). The Drosophila Snail family member Worniu is specifically transcribed in neural progenitors (neuroblasts) throughout their lifespan, and worniu mutants show defects in neuroblast delamination (a form of EMT). However, the role of Worniu in neuroblasts beyond their formation is unknown. We performed RNA-seq on worniu mutant larval neuroblasts and observed reduced cell-cycle transcripts and increased neural differentiation transcripts. Consistent with these genomic data, worniu mutant neuroblasts showed a striking delay in prophase/metaphase transition by live imaging and increased levels of the conserved neuronal differentiation splicing factor Elav. Reducing Elav levels significantly suppressed the worniu mutant phenotype. We conclude that Worniu is continuously required in neuroblasts to maintain self-renewal by promoting cell-cycle progression and inhibiting premature differentiation.

A resource for manipulating gene expression and analyzing cis-regulatory modules in the Drosophila CNS.

Here, we describe the embryonic central nervous system expression of 5,000 GAL4 lines made using molecularly defined cis-regulatory DNA inserted into a single attP genomic location. We document and annotate the patterns in early embryos when neurogenesis is at its peak, and in older embryos where there is maximal neuronal diversity and the first neural circuits are established. We note expression in other tissues, such as the lateral body wall (muscle, sensory neurons, and trachea) and viscera. Companion papers report on the adult brain and larval imaginal discs, and the integrated data sets are available online (http://www.janelia.org/gal4-gen1). This collection of embryonically expressed GAL4 lines will be valuable for determining neuronal morphology and function. The 1,862 lines expressed in small subsets of neurons (<20/segment) will be especially valuable for characterizing interneuronal diversity and function, because although interneurons comprise the majority of all central nervous system neurons, their gene expression profile and function remain virtually unexplored.

A pair of inhibitory neurons are required to sustain labile memory in the Drosophila mushroom body.

Labile memory is thought to be held in the brain as persistent neural network activity. However, it is not known how biologically relevant memory circuits are organized and operate. Labile and persistent appetitive memory in Drosophila requires output after training from the α'ß' subset of mushroom body (MB) neurons and from a pair of modulatory dorsal paired medial (DPM) neurons. DPM neurons innervate the entire MB lobe region and appear to be pre- and postsynaptic to the MB, consistent with a recurrent network model. Here we identify a role after training for synaptic output from the GABAergic anterior paired lateral (APL) neurons. Blocking synaptic output from APL neurons after training disrupts labile memory but does not affect long-term memory. APL neurons contact DPM neurons most densely in the α'ß' lobes, although their processes are intertwined and contact throughout all of the lobes. Furthermore, APL contacts MB neurons in the α' lobe but makes little direct contact with those in the distal α lobe. We propose that APL neurons provide widespread inhibition to stabilize and maintain synaptic specificity of a labile memory trace in a recurrent DPM and MB α'ß' neuron circuit.