SEARCH · Search bioRxiv

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 Read source abstracts and check manuscript versions; preprints are not peer reviewed.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 631 records · Page 35Linked to original sources

bioRxiv2018

Tbx1 regulates extracellular matrix- and cell-cell interactions in the second heart field.

Tbx1, the major candidate gene for DiGeorge or 22q11.2 deletion syndrome, is required for efficient incorporation of cardiac progenitors (CPs) of the second heart field (SHF) into the heart. However, the mechanisms by which TBX1 regulates this process are still unclear. Here, we have used two independent models, mouse embryos and cultured cells, to define the role of TBX1 in establishing morphological and dynamic characteristics of SHF in the mouse. We found that loss of TBX1 impairs extra cellular matrix (ECM)-integrin-focal adhesion (FA) signaling in both models. Mosaic analysis in embryos showed that this function is non-cell autonomous and, in cultured cells, loss of TBX1 impairs cell migration and focal adhesions. Additionally, we found that ECM-mediated outside-in integrin signaling is disrupted upon loss of TBX1. Finally, we show that interfering with the ECM-integrin-FA axis between E8.5 and E9.5 in mouse embryos, corresponding to the time window within which TBX1 is required in the SHF, causes outflow tract dysmorphogenesis. Our results demonstrate that TBX1 is required to maintain the integrity of ECM-cell interactions in the SHF, and that this interaction is critical for cardiac outflow tract development. More broadly, our data identifies a novel TBX1 downstream pathway as an important player in SHF tissue architecture and cardiac morphogenesis.

developmental biology↗

bioRxiv2018

Week-long imaging of cell divisions in the Arabidopsis root meristem

Characterizing the behaviors of dynamic systems requires capturing them with high temporal and spatial resolution. Owing to its transparency and genetic tractability, the Arabidopsis thaliana root lends itself well to live imaging when combined with cell and tissue-specific fluorescent reporters. We developed a novel 4D imaging method that utilizes simple confocal microscopy and readily available components to track cell divisions in the root stem cell niche and surrounding region for up to one week. This new setup allows us to finely analyze meristematic cell division rates that lead to patterning. Using this method, we performed a direct measurement of cell division intervals within and around the root stem cell niche. The results reveal a short, steep gradient of cell division in proximal stem cells, with progressively more rapid cell division rates from QC, to cells in direct contact with the QC (initials), to their immediate daughters, after which division rates appear to become more homogeneous. These results provide a baseline to study how perturbations in signaling could affect cell division patterns in the root meristem.

developmental biology↗

bioRxiv2018

From spiral cleavage to bilateral symmetry: The developmental cell lineage of the annelid brain

The spiral cleavage pattern is characteristic for Spiralia (Lophotrochozoa), a large assembly of marine invertebrates. In most cases, spiral cleavage produces freely swimming, trochophora-type larvae with a simple nervous system that controls ciliary locomotion. These larvae acquire bilateral symmetry, as manifested for example in the larval brain. The transition from the rotational symmetry of spiral cleavage into the bilateral adult body has not yet been understood. Here, we present the developmental cell lineage of the brain of the annelid Platynereis dumerilii from the zygote until the mid-trochophore stage (~30 hpf), in combination with a gene expression atlas for several embryonic and larval stages. Comparison of multiple embryos reveals a highly stereotypical development and an invariant cell lineage of the differentiated cell types. In addition, we observe a fundamental subdivision of the larval brain into a highly proliferative dorsolateral region and an early differentiating ventromedial region that gives rise to the apical nervous system. The transition from rotational to bilateral symmetry progresses gradually from the lateral to the central regions. Strikingly, the spiral-to-bilateral transition does not involve extensive cell migration. Rather, corresponding cells in different spiral quadrants acquire highly divergent identities in line with their bilateral position.

developmental biology↗

bioRxiv2018

Early lethality of embryos derived from transgenic Xenopus females is associated with reduced ovarian grem1 expression

The grem1 gene codes a protein that inhibits the action of multiple members of a growth factor family known as bone morphogenetic proteins (BMPs). Certain members of this BMP family can regulate both fecundity and fertility in mammals via their action on oocyte (egg) development, and grem1 has been identified as a marker of oocyte quality in humans. The model amphibian Xenopus laevis is far more fecund than mammals, producing thousands of eggs in a clutch. However, female transgenic frogs carrying grem1 under the control of a stress inducible hsp70 promoter (\"G\" frogs) produce very few viable offspring. Here, we show that this is not due to reduced fecundity or fertilization rate, but results from a significant reduction in subsequent survival over the first day of development. Embryos that successfully survive for the first day were found to go on to develop normally when compared to their peers. Both the morphology and stage distribution of oocytes from G females appears normal, and oocytes develop at expected rates, although stage VI oocytes were found to have a lower response to in vitro progesterone treatment. Unexpectedly, levels of grem1 mRNA were found to be consistently lower in the female ovaries from four independent G transgenic lines than in wild type ovaries. Both transgenic and wild type offspring were equally affected, confirming a maternal effect. Our study shows that transgenic females with the lowest levels of grem1 transcripts in the ovary have the lowest rates of survival past the first day of amphibian embryogenesis, equivalent to pre-implantation staged mammalian embryos. The reduced expression of grem1 in the oocytes of transgenic females suggests transgene supression of an endogenous locus may occur in the Xenopus female germline, an unexpected finding.

developmental biology↗

bioRxiv2018

An unconventional myosin, myosin 1d regulates Kupffer’s vesicle morphogenesis and laterality

INTRODUCTION INTRODUCTION RESULTS MATERIALS AND METHODS AUTHOR CONTRIBUTIONS REFERENCES Establishing left-right (LR) asymmetry is a fundamental process essential for arrangement of visceral organs during development. In vertebrates, motile cilia driven fluid flow in the left-right organizer (LRO) is essential for initiating symmetry breaking event1-3. Without a definite LRO structure in invertebrates, LR asymmetry is initiated at a cellular level by actin-myosin driven chirality4, 5. In Drosophila, myosin1D drives tissue-specific chirality in hind-gut looping6, 7. Here, we show that myosin 1d (myo1d) is essential for establishing LR asymmetry in zebrafish. Using super-r ...

developmental biology↗

bioRxiv2018

ADAR1 A-to-I RNA alters codon usage

BackgroundFully grown mammalian oocytes and eggs are transcriptionally quiescent, and therefore have a unique RNA environment in which cellular processes depend on post-transcriptional regulation. RNA editing of adenosines into inosines (A-to-I) by adenosine deaminases acting on RNA (ADARs) is a common post-transcriptional gene regulatory mechanism, yet it has not been systematically studied in oocytes.\n\nResultsA genome-wide RNA editing analysis of transcriptionally active growing oocytes from postnatal day 12 (PND12) mice, fully grown germinal vesicle (GV) oocytes, and transcriptionally quiescent metaphase II (MII) eggs indicates an abundant amount of A-to-I editing of mRNA transcripts. Editing of mRNA was greatest in GV oocyte and MII eggs compared to the PND12 immature oocytes, this was consistent with ADAR1 levels within these cells. Compared to somatic tissues, oocytes exhibited a different pattern of RNA editing, with a high proportion of RNA edits occurring in the coding regions. These edits resulted in nucleotide substitutions that were enriched at the third nucleotide of the codon (wobble position). Codon usage can affect mRNA stability and translation efficiency.\n\nConclusionsRNA editing in mouse oocytes is distinct from RNA editing in somatic cells due to increased frequencies of coding sequence RNA edits. We provide evidence in support of a previously unreported phenomenon of selective ADAR1 editing of the codon wobble position. Editing of the wobble position has the potential to fine tune post-transcriptional gene regulation through altering codon usage. This important observation advances our current understanding of RNA editing in mammalian cells.

developmental biology↗

bioRxiv2018

SOX2 Regulation by Hedgehog Signaling Controls Adult Lingual Epithelium Homeostasis

The adult tongue epithelium is continuously renewed from epithelial progenitor cells, and this process relies on intact Hedgehog (HH) signaling. In mice, inhibition of the HH pathway using Smoothened antagonists (HH pathway inhibitors or HPIs) leads to taste bud loss over a span of several weeks. Previously, we demonstrated that overexpression of Sonic Hedgehog (SHH) in lingual epithelial progenitors induces formation of ectopic taste buds accompanied by locally increased SOX2 expression, consistent with the hypothesis that taste bud differentiation depends on SOX2 downstream of HH. To test this idea, we inhibited HH signaling by treating SOX2-GFP mice with HPI and found a rapid and drastic decline in SOX2-GFP expression in taste progenitors and taste buds. Using a conditional Cre-lox system to delete Sox2, we found that loss of SOX2 blocks differentiation of both taste buds and non-taste epithelium that comprises the majority of the tongue surface; progenitor cells increase in number at the expense of differentiated taste cells and lingual keratinocytes. In contrast to the normal pattern of basally restricted proliferation, dividing cells are overabundant, disorganized and present in suprabasal epithelial layers in Sox2 deleted tongues. Additionally, SOX2 loss in taste progenitors leads non-cell autonomously to rapid loss of taste bud cells via apoptosis, dramatically shortening taste cell lifespans. Finally, when Sox2 is conditionally deleted in mice with constitutive overexpression of SHH, ectopic taste buds fail to form and endogenous taste buds disappear; instead, robust hyperproliferation takes over the entire lingual epithelium. In sum, our experiments suggest that SOX2 functions downstream of HH signaling to regulate lingual epithelium homeostasis.

developmental biology↗

bioRxiv2018

Tracing the Transitions from Pluripotency to Germ Cell Fate with CRISPR Screening

Early mammalian development entails a series of cell fate transitions that includes transit through naive pluripotency to post-implantation epiblast. This subsequently gives rise to primordial germ cells (PGC), the founding population of the germline lineage. To investigate the gene regulatory networks that control these critical cell fate decisions, we developed a compound-reporter system to track cellular identity in a model of PGC specification (PGC-like cells; PGCLC), and coupled it with unbiased genome-wide CRISPR screening. This enabled identification of key genes both for exit from pluripotency and for acquisition of PGC fate, with further characterisation revealing a central role for the transcription factors Nr5a2 and Zfp296 in germline ontogeny. Abrogation of these genes results in significantly impaired PGCLC development due to widespread activation (Nr5a2-/-) or inhibition (Zfp296-/-) of WNT pathway components. This leads to aberrant upregulation of the somatic programme or failure to appropriately activate germline genes in PGCLC, respectively, and consequently loss of germ cell identity. Overall our study places Zfp296 and Nr5a2 as key components of an expanded PGC gene regulatory network, and outlines a transferable strategy for identifying critical regulators of complex cell fate transitions.

developmental biology↗

bioRxiv2018

Sexual stage-induced long noncoding RNAs in the filamentous fungus Fusarium graminearum

Long noncoding RNA (lncRNA) plays important roles in morphological differentiation and development in eukaryotes. In filamentous fungi, however, little is known about lncRNAs and their roles in sexual development. Here we describe sexual stage-induced lncRNAs during the formation of perithecia, the sexual fruiting bodies of Fusarium graminearum. We have identified 547 lncRNAs whose expression was developmental stage-specific, with about 40% of which peaked during the development of asci, the sac-like structures containing meiospores. A large fraction of the lncRNAs were found to be antisense to mRNAs, forming 300 sense-antisense pairs. Although small RNAs (sRNAs) were produced from these overlapping loci, most of the antisense lncRNAs appeared not to be involved in gene silencing pathways. Genome-wide analysis of sRNA clusters identified many silenced loci at the meiotic stage. However, we found transcriptionally-active sRNA clusters, many of which were associated with lncRNAs. Also, we observed that many antisense lncRNAs and their respective sense transcripts were induced in parallel as the perithecia matured. To identify regulatory components for lncRNA expression, we analyzed mutants defective in the nonsense-mediated decay (NMD) pathway. A subset of the lncRNAs appeared to be targeted by the NMD before the perithecia formation, suggesting a suppressive role of the NMD in lncRNA expression during vegetative stage. This research provides fundamental genomic resources that will spur further investigations on developmental lncRNAs that may play important roles in shaping the fungal fruiting bodies.

developmental biology↗

bioRxiv2018

Integrin signaling downregulates filopodia in muscle-tendon attachment

Cells in developing tissues need to sense their environment for their accurate targeting to specific destinations. This occurs in developing muscles, which need to attach to their respective tendon cell before muscle contractions can begin. Elongating myotube tips form filopodia, which are presumed to have sensory roles, and are later suppressed upon building the attachment site. Here, we use live imaging and quantitative image analysis of lateral transverse (LT) myotubes in Drosophila to show that filopodia suppression occurs as a result of integrin signaling. Loss of the integrin subunits PS2 and {beta}PS increased filopodia number and length at stages when they are normally suppressed. Conversely, inducing integrin signaling, achieved by expression of constitutively dimerised {beta}PS cytoplasmic domain (di{beta}), prematurely suppressed filopodia. We discovered that the integrin signal is transmitted through the ArfGAP and scaffolding protein Git (G-protein receptor coupled interacting protein) and its downstream kinase Pak (p21-activated kinase). Absence of these proteins causes profuse filopodia formation and prevents filopodial inhibition by di{beta}. Thus, integrin signaling switches off the exploratory behaviour of myotubes seeking tendons, enabling the actin machinery to focus on forming a strong attachment and assembling the contractile apparatus.

developmental biology↗

bioRxiv2018

The TMEM16A Channel Mediates the fast polyspermy block in Xenopus Laevis

In externally fertilizing animals, such as sea urchins and frogs, prolonged depolarization of the egg immediately after fertilization inhibits the entry of additional sperm - a phenomenon known as the fast block to polyspermy. In the African clawed frog, Xenopus laevis, this depolarization is driven by a Ca2+-activated Cl- efflux. Although the prominent Ca2+-activated Cl- currents generated by immature X. laevis oocytes are conducted by xTMEM16A channels, little is known about which channels contribute to fertilization-competency in mature eggs. Moreover, the gamete undergoes a gross transformation as it matures from an immature oocyte into a fertilization-competent egg. Here we report the results of our approach to identify the Ca2+-activated Cl- channel that triggers the fast block. Querying published proteomics and RNA-seq data, we identified two Ca2+-activated Cl- channels expressed in fertilization-competent X. laevis eggs: xTMEM16A and xBEST2A. Furthermore, transcripts for these channels increase in abundance during gamete maturation. To determine if either of these mediates the fast block, we characterized exogenously expressed xTMEM16A and xBEST2A using pharmacologic inhibitors. None of the inhibitors tested blocked xBEST2A currents specifically. However, Ani9 and MONNA each reduced xTMEM16A currents by more than 70%, while only nominally inhibiting those generated by xBEST2A. Using whole-cell recordings during fertilization, we found that Ani9 and MONNA effectively diminished fertilization-evoked depolarizations. These results indicate that fertilization activates TMEM16A channels in X. laevis eggs and induces the earliest known event triggered by fertilization: the fast block to polyspermy.\n\nHIGHLIGHTSO_LIProtein for the channels xBEST2A and xTMEM16A is present in X. laevis eggs.\nC_LI\n\nO_LIThe inhibitors MONNA and Ani9 effectively block xTMEM16A compared to xBEST2A.\nC_LI\n\nO_LIXenopus laevis fertilization opens TMEM16A to trigger egg depolarization.\nC_LI\n\nO_LIThe TMEM16A-mediated depolarization is critical for the fast block to polyspermy.\nC_LI

developmental biology↗

bioRxiv2018

Loss of embryonic neural crest cardiomyocytes causes adult hypertrophic cardiomyopathy

Neural crest cells migrate to the embryonic heart and transform into a small number of cardiomyocytes, but their functions in the developing and adult heart are unknown. Here, we map the fates of neural crest derived cardiomyocytes (NC-Cms) and genetically ablate them in embryogenesis in zebrafish. Specific NC-Cm ablation results in aberrant trabeculation patterns and altered Notch signaling, but is not detrimental to the development of the fish or early heart function. Strikingly, embryonic NC-Cm ablation results in adult fish that show severely hypertrabeculated hearts, altered cardiomyocyte size, diminished adult heart capacity and consequently poor physiological response to cardiac stress tests. Thus, we identify a novel developmental mechanism and genetic pathway that predisposes adults to hypertrophic cardiomyopathy and provides the first zebrafish model of adult-onset heart failure.

developmental biology↗

bioRxiv2018

Elongation during segmentation shows axial variability, low mitotic rates, and synchronized cell cycle domains in the crustacean, Thamnocephalus platyurus

Segmentation in arthropods typically occurs by sequential addition of segments from a posterior growth zone, but cell behaviors producing posterior elongation are not well known. Using precisely staged larvae of the crustacean, Thamnocephalus platyurus, we systematically examined cell division patterns and morphometric changes associated with posterior elongation during segmentation. We show that cell division is required for normal elongation but that cells in the growth zone need only divide ~1.5 times to meet that requirement; correspondingly, direct measures of cell division in the growth zone are low. Morphometric measurements of the growth zone and of newly formed segments suggest tagma-specific features of segment generation. Using methods for detecting two different phases in the cell cycle, we show distinct domains of synchronized cells in the posterior. Borders of cell cycle domains correlate with domains of segmental gene expression, suggesting an intimate link between segment generation and cell cycle regulation.\n\nSummary StatementPosterior growth zone has synchronized cell cycle domains but shows little cell division during segment addition in a crustacean. Dimensions of the shrinking posterior growth zone change at tagma boundaries.

developmental biology↗

bioRxiv2018

Hedgehog signaling controls progenitor differentiation timing during heart development

Heterochrony, defined as differences in the timing of developmental processes, impacts organ development, homeostasis, and regeneration. The molecular basis of heterochrony in mammalian tissues is poorly understood. We report that Hedgehog signaling activates a heterochronic pathway that controls differentiation timing in multiple lineages. A differentiation trajectory from second heart field cardiac progenitors to first heart field cardiomyocytes was identified by single-cell transcriptional profiling in mouse embryos. A survey of developmental signaling pathways revealed specific enrichment for Hedgehog signaling targets in cardiac progenitors. Removal of Hh signaling caused loss of progenitor and precocious cardiomyocyte differentiation gene expression in the second heart field in vivo. Introduction of active Hh signaling to mESC-derived progenitors, modelled by transient expression of the Hh-dependent transcription factor GLI1, delayed differentiation in cardiac and neural lineages in vitro. A shared GLI1-dependent network in both cardiac and neural progenitors was enriched with FOX family transcription factors. FOXF1, a GLI1 target, was sufficient to delay onset of the cardiomyocyte differentiation program in progenitors, by epigenetic repression of cardiomyocyte-specific enhancers. Removal of active Hh signaling or Foxf1 expression from second heart field progenitors caused precocious cardiac differentiation in vivo, establishing a mechanism for resultant Congenital Heart Disease. Together, these studies suggest that Hedgehog signaling directly activates a gene regulatory network that functions as a heterochronic switch to control differentiation timing across developmental lineages.

developmental biology↗

bioRxiv2018

A microtubule-based mechanism predicts cell division orientation in plant embryogenesis

Oriented cell divisions are significant in plant morphogenesis because plant cells are embedded in cell walls and cannot relocate. Cell divisions follow various regular orientations, but the underlying mechanisms have not been clarified. We show that cell-shape dependent self-organisation of cortical microtubule arrays is crucial for determining planes of early tissue-generating divisions and forms the basis for robust control of cell division orientation in the embryo. To achieve this, we simulate microtubules on actual cell surface shapes from which we derive a minimal set of three rules for proper array orientation. The first rule captures the effects of cell shape alone on microtubule organisation, the second rule describes the regulation of microtubule stability at cell edges and the third rule includes the differential effect of auxin on local microtubule stability. These rules explain early embryonic division plane orientations and offer a framework for understanding patterned cell divisions in plant morphogenesis.

developmental biology↗

bioRxiv2018

Characterization of convergent thickening, a major convergence force producing morphogenic movement in amphibians

We characterize the morphogenic process of convergent thickening (CT), which occurs in the involuting marginal zone (IMZ) during gastrulation of Xenopus, the African clawed frog. CT was described previously as the tendency of explants of the ventral IMZ of Xenopus to converge their circumblastoporal dimension and thicken their radial dimension (Keller and Danilchik 1988). Here we show that CT occurs from the onset of gastrulation, initially throughout the pre-involution IMZ. We suggest that CT is driven by an increase in the interfacial tension between the deep IMZ and its epithelium, resulting in cells of the deep IMZ tending to minimize their surface area. In explants, this results in a progressive shortening (convergence) of the IMZ along its longer mediolateral axis and thickening in the orthogonal planes, and can generate tensile force (Shook et al. 2018). In vivo, convergence of the annular IMZ generates circumferential tension, closing the blastopore. These results provide the first clear example of a tensile morphogenic force from a Holtfreterian/Steinbergian change in tissue affinity.

developmental biology↗

bioRxiv2018

p53 deletion rescues apoptosis and microcephaly in a Kif20b mouse mutant

Building a cerebral cortex of the proper size involves balancing rates and timing of neural stem cell (NSC) proliferation, neurogenesis, and cell death. The cellular mechanisms connecting genetic mutations to brain malformation phenotypes are still poorly understood. Microcephaly may result when NSC divisions are too slow, produce neurons too early, or undergo apoptosis, but the relative contributions of these cellular mechanisms to various types of microcephaly are not understood. We previously showed that mouse mutants in Kif20b (formerly called Mphosph1, Mpp1, or KRMP1) have small cortices that show elevated apoptosis, and defects in maturation of NSC midbodies, which mediate cytokinetic abscission. Here we test the contribution of intrinsic NSC apoptosis to brain size reduction in this lethal microcephaly model. By making double mutants with the pro-apoptotic genes Bax and Trp53 (p53), we find that apoptosis of cortical NSCs accounts for most of the microcephaly, but that there is a significant apoptosis-independent contribution as well. Remarkably, heterozygous p53 deletion is sufficient to fully rescue survival of the Kif20b mutant into adulthood. In addition, the NSC midbody maturation defects are not rescued by p53 deletion, showing that they are either upstream of p53 activation, or in a parallel pathway. Thus, this work potentially identifies a novel midbody-mediated pathway for p53 activation, and elucidates both NSC apoptosis and abscission mechanisms that could underlie human microcephaly or other brain malformations.

developmental biology↗

bioRxiv2018

Verified the effectiveness of AsCpf1 system in a variety of vertebrate species

CRISPR/Cpf1 system is a novel genomic editing tool. Because of its more sophisticated components, and lower off-target rate, it has the potential to become a better gene-editing tool. Previous reports showed that CRISPR/Cpf1 could work effectively in multiple species. But our data show that AsCpf1 activity has a big difference in different vertebrates. Using in vitro experiments, we finally learned that the difference between species is due to temperature.

developmental biology↗