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Results for “Developmental Biology”

Search indexed bioRxiv preprints in genomics, neuroscience, cell biology and bioinformatics. Read source abstracts and check manuscript versions; preprints are not peer reviewed.

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At least 379 records · Page 21Linked to original sources

bioRxiv2017

Evidence for the temporal regulation of insect segmentation by a conserved set of developmental transcription factors

Long-germ insects, such as the fruit fly Drosophila melanogaster, pattern their segments simultaneously, whereas short germ insects, such as the beetle Tribolium castaneum, pattern their segments sequentially, from anterior to posterior. While the two modes of segmentation at first appear to be very different, many details of segmentation gene expression are surprisingly similar between long-germ and short-germ species. Collectively, these observations hint that insect segmentation may involve fairly conserved patterning mechanisms, which occur within an evolutionarily malleable spatiotemporal framework. Based on genetic and comparative evidence, we now propose that, in both Drosophila and Tribolium embryos, the temporal progression of the segmentation process is regulated by a temporal sequence of Caudal, Dichaete, and Odd-paired expression. These three transcription factors are broadly expressed in segmenting tissues, providing spatiotemporal information that intersects with the information provided by periodically-expressed segmentation genes such as the pair-rule factors. However, they are deployed differently in long-germ versus short-germ insects, acting as simple timers in Drosophila, but as smooth, retracting wavefronts in Tribolium, compatible with either gap gene-based or oscillator-based generation of periodicity, respectively.

developmental biology↗

bioRxiv2017

Characterization Of NvLWamide-Like Neurons Reveals Stereotypy In Nematostella Nerve Net Development

Cnidarian nervous systems are traditionally described as diffuse nerve nets lacking true organization. However, there are examples of stereotypical structure in the nerve nets of multiple cnidarian species that suggest nerve nets are organized. We previously demonstrated that the NvashA target gene NvLWamide-like is expressed in a small subset of the Nematostella nerve net and speculated that observing a few neurons within the developing nerve net would provide a better indication of potential stereotypy. Here we document NvLWamide-like expression more systematically. NvLWamide-like is initially expressed in the typical neurogenic salt and pepper pattern within the ectoderm at the gastrula stage, and expression expands to include endodermal salt and pepper expression at the planula larval stage. Expression persists in both ectoderm and endoderm in adults. We generated an NvLWamide-like::mCherry transgenic reporter line to visualize the neural architecture. NvLWamide-like is expressed in six neural subtypes identifiable by neural morphology and location. Upon completing development the numbers of neurons in each neural subtype are minimally variable between animals and the projection patterns of each subtype are consistent. Between the juvenile polyp and adult stages the number of neurons for each subtype increases. We conclude that cnidarian nerve nets are organized, develop in a stereotyped fashion, and that one aspect of generating the adult nervous system is to modify the juvenile nervous system by increasing neural number proportionally with size.

developmental biology↗

bioRxiv2017

The Effects of Central Nervous System Stimulants on Drosophila melanogaster Reproduction

Stimulant drugs are used everyday by people around the world. The effect stimulants have on developing human fetuses is widely unknown. The fruit fly Drosophila melanogaster has become a valuable system to model the complex effects and properties of drugs in mammals. In this study, Drosophila is used to analyze the effects of stimulant exposure on reproduction to determine if stimulants cause a significant decrease in the number of offspring produced by parent generations. Caffeine, nicotine, and pseudoephedrine hydrochloride were found to significantly decrease the number of offspring in experimental populations. Further experimentation is necessary to understand the mechanisms underlying these results.

developmental biology↗

bioRxiv2017

Impairment of zebrafish reproduction upon exposure to melengestrol acetate

Synthetic progestins contamination is common in the aquatic ecosystem, which may lead to serious health problem on aquatic animals. Melengestrol acetate (MGA) has been detected in the aquatic environment; however, its potential effects on fish reproduction are largely unclear. Here, we aimed to investigate the endocrine disruption and impact of MGA on zebrafish reproduction. Six-month old reproductive zebrafish were exposed to four nominal concentrations of MGA (1,10, 100 and 200 ng/L) for 15 days. Treatment with MGA reduced the egg production with a significant decrease at 200 ng/L. The circulating concentrations of estradiol and testosterone in female zebrafish or 11-keto testosterone in male zebrafish were significantly diminished compared to the non-exposed control fish. The early embryonic development or hatching rates were unaffected during the MGA exposure. Our results indicated that MGA was a potent endocrine disruptor in fish and the fish reproduction could be impaired even during a short-term exposure to MGA.

developmental biology↗

bioRxiv2017

Stepwise approach to oocyte depletion in Sry mutated XY female mice

Little is known regarding the mechanisms underlying infertility in male-to-female sex-reversed females. To gain a better understanding of germ cell dysfunction in this condition, we produced XYsry--females via the CRISPR/Cas9 system in C57BL/6 inbred strain mice. Mutant mice showed severe attrition of germ cells during foetal development, resulting in depletion of ovarian germ cells before sexual maturation. Comprehensive transcriptome analysis of embryonic day 13.5 primordial germ cells (PGCs) and postnatal day 1 oocytes demonstrated that XYsry--PGCs had already deviated from the developmental process at the mitotic stage. In addition, XYsry--oocytes resulted in meiotic disruption and developmental failure, and the genes related to these processes were significantly changed. This grievous disruption, which caused germ cell deterioration in XYsry--females, was shown to proceed from the germ cells themselves. These results provide novel insight into the germ cell depletion of sex-reversed mice as well as into disorders of sex differentiation in human, such as Swyer syndrome, wherein patients present as typical females albeit with an XY karyotype and infertile.

developmental biology↗

bioRxiv2017

Epigenetic resetting of human pluripotency

Much attention has focussed on conversion of human pluripotent stem cells (PSC) to a more naive developmental status. Here we provide a method for resetting via transient histone deacetylase inhibition. The protocol is effective across multiple PSC lines and can proceed without karyotype change. Reset cells can be expanded without feeders with a doubling time of around 24 hours. WNT inhibition stabilises the resetting process. The transcriptome of reset cells diverges markedly from primed PSC and shares features with human inner cell mass (ICM). Reset cells activate expression of primate-specific transposable elements. DNA methylation is globally reduced to the level in the ICM but is non-random, with gain of methylation at specific loci. Methylation imprints are mostly lost, however. Reset cells can be re-primed to undergo tri-lineage differentiation and germline specification. In female reset cells, appearance of bi-allelic X-linked gene transcription indicates re-activation of the silenced X chromosome. On re-conversion to primed status, XIST-induced silencing restores monoallelic gene expression. The facile and robust conversion routine with accompanying data resources will enable widespread utilisation, interrogation, and refinement of candidate naive cells.

developmental biology↗

bioRxiv2017

Cell code: the evolving arrangement of molecules that perpetuates life

Life is perpetuated through a single-cell bottleneck between generations in many organisms. Here, I highlight that this cell holds information in two distinct forms: in the linear DNA sequence that is replicated during cell divisions, and in the three-dimensional arrangement of molecules that can change during development but that is recreated at the start of each generation. These two interdependent stores of information - one replicating with each cell division and the other cycling with a period of one generation - coevolve while perpetuating an organism. Unlike the genome, the cycling arrangement of molecules, which could include RNAs, proteins, sugars, lipids, etc., is not well understood. Because this arrangement and the genome are together transmitted from one generation to the next, analysis of both is necessary to understand evolution, origins of inherited diseases, and consequences of genome engineering. Recent developments suggest that tools are in place to examine how all the information to build an organism is encoded within a single cell, and how this cell code is reproduced in every generation.

developmental biology↗

bioRxiv2017

An optimized pipeline for parallel image-based quantification of gene expression and genotyping after in situ hybridization

Advances in genome engineering have resulted in the generation of numerous zebrafish mutant lines. A commonly used method to assess gene expression in the mutants is in situ hybridization (ISH). Because the embryos can be distinguished by genotype after ISH, comparing gene expression between wild type and mutant siblings can be done blinded and in parallel. Such experimental design reduces the technical variation between samples and minimises the risk of bias. This approach, however, requires an efficient method of genomic DNA extraction from post-ISH fixed zebrafish samples to ascribe phenotype to genotype. Here we describe a method to obtain PCR-quality DNA from 95-100% of zebrafish embryos, suitable for genotyping after ISH. In addition, we provide an image analysis protocol for quantifying gene expression of ISH-probed embryos, adaptable for the analysis of different expression patterns. Finally, we show that intensity-based image analysis enables accurate representation of the variability of gene expression detected by ISH and that it can complement quantitative methods like qRT-PCR. By combining genotyping after ISH and computer-based image analysis, we have established a high-confidence, unbiased methodology to assign gene expression levels to specific genotypes, and applied it to the analysis of molecular phenotypes of newly generated lmo4a mutants.\n\nSUMMARY STATEMENTOur optimized protocol to genotype zebrafish mutant embryos after in situ hybridization and digitally quantify the in situ signal will help to standardize existing experimental designs and methods of analysis.

developmental biology↗

bioRxiv2017

Neural stem cells induce the formation of their physical niche during organogenesis

Most organs rely on stem cells to maintain homeostasis during post-embryonic life. Typically, stem cells of independent lineages work coordinately within mature organs to ensure proper ratios of cell types. Little is known, however, on how these different stem cells locate to forming organs during development. Here we show that neuromasts of the posterior lateral line in medaka are composed of two independent life-long lineages with different embryonic origins. Clonal analysis and 4D imaging revealed a hierarchical organisation with instructing and responding roles: an inner, neural lineage induces the formation of an outer, border cell lineage (nBC) from the skin epithelium. Our results demonstrate that the neural lineage is necessary and sufficient to generate nBCs highlighting self-organisation principles at the level of the entire embryo. We hypothesise that transformation of surrounding tissues plays a major role during the establishment of vertebrate stem cell niches.

developmental biology↗

bioRxiv2017

Egr1 deficiency induces browning of inguinal subcutaneous white adipose tissue in mice

Beige adipocyte differentiation within white adipose tissue, referred to as browning, is seen as a possible mechanism for increasing energy expenditure. The molecular regulation underlying the thermogenic browning process has not been entirely elucidated. Here, we identify the zinc finger transcription factor EGR1 as a negative regulator of the beige fat program. Loss of Egr1 in mice promotes browning in the absence of external stimulation and activates Ucp1 that encodes the key thermogenic mitochondrial uncoupling protein-1. Moreover, EGR1 is recruited to the proximal region of the Ucp1 promoter in subcutaneous inguinal white adipose tissue. Transcriptomic analysis of subcutaneous inguinal white adipose tissue in the absence of Egr1 identifies the molecular signature of white adipocyte browning downstream of Egr1 deletion and highlights a concomitant increase of beige differentiation marker and decrease in extracellular matrix gene expression. Conversely, Egr1 overexpression in mesenchymal stem cells decreases beige adipocyte differentiation, while increasing extracellular matrix production. These results uncover the role of Egr1 in blocking energy expenditure via direct Ucp1 transcription regulation and highlight Egr1 as a therapeutic target for counteracting obesity.

developmental biology↗

bioRxiv2017

A single cell transcriptional roadmap for cardiopharyngeal fate diversification

In vertebrates, multipotent progenitors located in the pharyngeal mesoderm form cardiomyocytes and branchiomeric head muscles, but the dynamic gene expression programs and mechanisms underlying cardiopharyngeal multipotency and heart vs. head muscle fate choices remain elusive. Here, we used single cell genomics in the simple chordate model Ciona, to reconstruct developmental trajectories forming first and second heart lineages, and pharyngeal muscle precursors, and characterize the molecular underpinnings of cardiopharyngeal fate choices. We show that FGF-MAPK signaling maintains multipotency and promotes the pharyngeal muscle fate, whereas signal termination permits the deployment of a pan-cardiac program, shared by the first and second lineages, to define heart identity. In the second heart lineage, a Tbx1/10-Dach pathway actively suppresses the first heart lineage program, conditioning later cell diversity in the beating heart. Finally, cross-species comparisons between Ciona and the mouse evoke the deep evolutionary origins of cardiopharyngeal networks in chordates.

developmental biology↗

bioRxiv2017

Inter-organ regulation of Drosophila intestinal stem cell proliferation by a hybrid organ boundary zone

SUMMARY STATEMENTGene expression at the Drosophila midgut-hindgut boundary is a hybrid of both organs. Hybrid cells repress stem cell division, but boundary injury activates stem cell division through inter-organ JAK-STAT signaling.\n\nABSTRACTThe molecular identities and regulation of cells at inter-organ boundaries are often unclear, despite the increasingly appreciated role of organ boundaries in disease. Using Drosophila as a model, here we show that a specific population of adult midgut organ boundary intestinal stem cells (OB-ISCs) is regulated by the neighboring hindgut, a developmentally distinct organ. This distinct OB-ISCs control is due to proximity to a specialized transition zone between the endodermal midgut and ectodermal hindgut that shares molecular signatures of both organs, which we term the hybrid zone (HZ). During homeostasis, proximity to the HZ restrains OB-ISC proliferation. However, injury to the adult HZ/hindgut drives up-regulation of upaired-3 cytokine and OB-ISC hyperplasia. If HZ disruption is severe, hyperplastic OB-ISCs expand across the inter-organ boundary. Our data suggest that inter-organ signaling plays an important role in controlling OB-ISCs in homeostasis and injury repair, which is likely critical in prevention of disease.

developmental biology↗

bioRxiv2017

A Positive Feedback Loop Ensures Propagation of ROS Production and JNK Signaling Throughout Drosophila Tissue Regeneration

Regenerating tissue must initiate the signaling that drives regenerative growth, and sustain that signaling long enough for regeneration to complete. How these key signals are sustained is unclear. To gain a comprehensive view of the changes in gene expression that occur during regeneration, we performed wholegenome mRNAseq of actively regenerating tissue from damaged Drosophila wing imaginal discs. We used genetic tools to ablate the wing primordium to induce regeneration, and carried out transcriptional profiling of the regeneration blastema by fluorescent labeling and sorting the blastema cells, thus identifying differentially expressed genes. Importantly, by using genetic mutants of several of these differentially expressed genes we have confirmed that they have roles in regeneration. Using this approach, we show that high expression of the gene moladietz (mol), which encodes the Duox-maturation factor NIP, is required during regeneration to produce reactive oxygen species (ROS), which in turn sustain JNK signaling during regeneration. We also show that JNK signaling upregulates mol expression, thereby activating a positive feedback signal that ensures the prolonged JNK activation required for regenerative growth. Thus, by wholegenome transcriptional profiling of regenerating tissue we have identified a positive feedback loop that regulates the extent of regenerative growth.\n\nAuthor summaryRegenerating tissue must initiate the signaling that drives regenerative growth, and then sustain that signaling long enough for regeneration to complete. Drosophila imaginal discs, the epithelial structures in the larva that will form the adult animal during metamorphosis, have been an important model system for tissue repair and regeneration for over 60 years. Here we show that damage-induced JNK signaling leads to the upregulation of a gene called moladietz, which encodes a co-factor for an enzyme, NADPH dual oxidase (DUOX), that generates reactive oxygen species (ROS), a key tissue-damage signal. High expression of moladietz induces continuous production of ROS in the regenerating tissue. The sustained production of ROS then continues to activate JNK signaling throughout the course of regeneration, ensuring maximal tissue regrowth.

developmental biology↗

bioRxiv2017

RAPGEF5 regulates nuclear transport of β-catenin

Canonical Wnt signaling coordinates many critical aspects of embryonic development, while dysregulated Wnt signaling contributes to common diseases, including congenital malformations and cancer. The nuclear localization of {beta}-catenin is the defining step in pathway activation. However, despite intensive investigation, the mechanisms regulating {beta}-catenin nuclear transport remain undefined. In a patient with congenital heart disease and heterotaxy, a disorder of left-right patterning, we previously identified the guanine nucleotide exchange factor, RAPGEF5. Here, we demonstrate that RAPGEF5 regulates left-right patterning via Wnt signaling. In particular, RAPGEF5, regulates the nuclear translocation of {beta}-catenin independently of both {beta}-catenin cytoplasmic stabilization and the importin {beta}1/Ran mediated transport system. We propose a model whereby RAPGEF5 activates the nuclear GTPases, Rap1/2, to facilitate the nuclear transport of {beta}-catenin, defining a parallel nuclear transport pathway to Ran. Our results suggest new targets for modulating Wnt signaling in disease states.

developmental biology↗

bioRxiv2017

WDR5 stabilizes actin architecture to promote multiciliated cell formation

HIGHLIGHTS O_LIWDR5 has an H3K4 independent role in the formation of multiciliated cells.\nC_LIO_LIWDR5 controls apical cell expansion, basal body patterning, and ciliogenesis in multiciliated cells.\nC_LIO_LIWDR5 localizes near the ciliary base where it connects basal bodies to F-actin.\nC_LIO_LIWDR5 stabilizes the apical actin network in multiciliated cells.\nC_LI\n\nSUMMARYThe actin cytoskeleton is critical to shape cells and pattern intracellular organelles to drive tissue morphogenesis. In multiciliated cells (MCCs), apical actin forms a lattice that drives expansion of the cell surface necessary to host hundreds of cilia. The actin lattice also uniformly distributes basal bodies across this surface. This apical actin network is dynamically remodeled, but the molecules that regulate its architecture remain poorly understood. We identify the chromatin modifier, WDR5, as a regulator of apical F-actin in multiciliated cells. Unexpectedly, WDR5 functions independently of chromatin modification in MCCs. Instead, we discover a scaffolding role for WDR5 between the basal body and F-actin. Specifically, WDR5 binds to basal bodies and migrates apically, where F-actin organizes around WDR5. Using a monomer trap for G-actin, we show that WDR5 stabilizes F-actin to maintain apical lattice architecture. In summary, we identify a novel, non-chromatin role for WDR5 in stabilizing F-actin in multiciliated cells.\n\nIN BRIEFKulkarni et al discover a chromatin independent function for WDR5 in multiciliated cell formation. WDR5 localizes to the base of cilia and functions as a scaffold between the basal bodies and the apical actin lattice. There, WDR5 stabilizes the actin lattice that allows multiciliated cells to expand their apical surface, pattern basal bodies, and generate hundreds of cilia.

developmental biology↗

bioRxiv2017

Natural variation in stochastic photoreceptor specification and color preference in Drosophila

Each individual perceives the world in a unique way, but little is known about the genetic basis of variation in sensory perception. Here we investigated natural variation in the development and function of the color vision system of Drosophila. In the fly eye, the random mosaic of color-detecting R7 photoreceptor subtypes is determined by stochastic expression of the transcription factor Spineless (Ss). Individual R7s randomly choose between SsON or SsOFF fates at a ratio of 65:35, resulting in unique patterns but consistent proportions of cell types across genetically identical retinas. In a genome wide association study, we identified a naturally occurring insertion in a regulatory DNA element in the ss gene that lowers the ratio of SsON to SsOFF cells. This change in photoreceptor fates shifts the innate color preference of flies from green to blue. The genetic variant increases the binding affinity for Klumpfuss (Klu), a zinc finger transcriptional repressor that regulates ss expression. Klu is expressed at intermediate levels to determine the normal ratio of SsON to SsOFF cells. Thus, binding site affinity and transcription factor levels are finely tuned to regulate stochastic on/off gene expression, setting the ratio of alternative cell fates and ultimately determining color preference.

developmental biology↗

bioRxiv2017

The chromatin factor Gon4l regulates embryonic axis extension by promoting mediolateral cell polarity and notochord boundary formation through negative regulation of cell adhesion

Anteroposterior axis extension during vertebrate gastrulation requires cell proliferation, embryonic patterning, and morphogenesis to be spatiotemporally coordinated, but the underlying genetic mechanisms remain poorly understood. Here we define a role for the conserved chromatin factor Gon4l, encoded by ugly duckling (udu), in coordinating tissue patterning and axis extension during zebrafish gastrulation. Although identified as a recessive enhancer of short axis phenotypes in planar cell polarity (PCP) mutants, we found that Gon4l functions in a genetically independent, partially overlapping fashion with PCP signaling to regulate mediolateral cell polarity underlying axis extension in part by promoting notochord boundary formation. We identified direct genomic targets of Gon4l and found that it acts as both a positive and negative regulator of gene expression, including limiting expression of the cell-cell and cell-matrix adhesion molecules EpCAM and Integrin3b. Excess epcam or itga3b in wild-type gastrulae phenocopied notochord boundary defects of udu mutants, while downregulation of itga3b suppressed them. By promoting formation of this anteroposteriorly aligned boundary and associated cell polarity, Gon4l cooperates with PCP signaling to coordinate morphogenesis with the anteroposterior embryonic axis.

developmental biology↗

bioRxiv2017

Sonic hedgehog is required for neural crest-dependent patterning of the intrinsic tongue musculature

The tongue is a highly specialized muscular organ important for breathing, speech, taste and swallowing. The secreted signaling molecule Sonic hedgehog (Shh) is expressed in dorsal tongue epithelium from the initial developmental stage. In this study, we utilized a series of genetic approaches to investigate the role of Shh during mouse tongue formation. Temporal-specific global deletion of Shh demonstrated a functional requirement for normal patterning of the intrinsic tongue muscles and establishment of the lingual tendon. These defects were reproduced in the mutant with a specific loss of signaling in oropharyngeal epithelium by a Shh cis-enhancer. In these mutants, Ptch1 was lost in the underlying cranial neural crest (CNC)-derived mesenchymal lineage. The importance of Shh was confirmed by generating tissue-specific deletions in the ciliopathy gene Ofd1, which transduces Shh signaling. These results revealed Shh roles in patterning of the mesodermal intrinsic tongue muscles through CNC-derived mesenchyme, including the lingual tendon.

developmental biology↗