A fundamental goal in biology is to understand the molecular basis of cell identity. Pluripotent embryonic stem (ES) cell identity is governed by a set of transcription factors centred on the triumvirate of Oct4, Sox2 and Nanog. These proteins often bind to closely localised genomic sites. Recent studies have identified additional transcriptional modulators that bind to chromatin near sites occupied by Oct4, Sox2 and Nanog. This suggests that the combinatorial control of gene transcription might be fundamental to the ES cell state. Here we discuss how these observations advance our understanding of the transcription factor network that controls pluripotent identity and highlight unresolved issues that arise from these studies.
]]>Regenerative ability varies depending on animal species and developmental stage, but the factors that determine this variability remain unclear. Although Xenopus laevis tadpole tails possess high regenerative ability, this is transiently lost during the `refractory period'. Here, we show that tail amputation evokes different immune responses in wound tail stumps between the `refractory' and `regeneration' periods: there was delayed or prolonged expression of some immune-related genes in the refractory period, whereas there was no obvious or transient expression of other immune-related genes in the regeneration periods. In addition, immune suppression induced by either immunosuppressant treatment or immune cell depletion by knockdown of PU.1 significantly restored regenerative ability during the refractory period. These findings indicate that immune responses have a crucial role in determining regenerative ability in Xenopus tadpole tails.
]]>Obtaining the diversity of interneuron subtypes in their appropriate numbers requires the orchestrated integration of progenitor proliferation with the regulation of differentiation. Here we demonstrate through loss-of-function studies in mice that the Cut homeodomain transcription factor Cux2 (Cutl2) plays an important role in regulating the formation of dorsal spinal cord interneurons. Furthermore, we show that Notch regulates Cux2 expression. Although Notch signaling can be inhibitory to the expression of proneural genes, it is also required for interneuron formation during spinal cord development. Our findings suggest that Cux2 might mediate some of the effects of Notch signaling on interneuron formation. Together with the requirement for Cux2 in cell cycle progression, our work highlights the mechanistic complexity in balancing neural progenitor maintenance and differentiation during spinal cord neurogenesis.
]]>The Immunoglobulin superfamily (IgSF) proteins Neph1 and Nephrin are co-expressed within podocytes in the kidney glomerulus, where they localize to the slit diaphragm (SD) and contribute to filtration between blood and urine. Herein, we demonstrate that their Drosophila orthologs Kirre (Duf) and Sns are co-expressed within binucleate garland cell nephrocytes (GCNs) that contribute to detoxification of the insect hemolymph by uptake of molecules through an SD-like nephrocyte diaphragm (ND) into labyrinthine channels that are active sites of endocytosis. The functions of Kirre and Sns in the embryonic musculature, to mediate adhesion and fusion between myoblasts to form multinucleate muscle fibers, have been conserved in the GCNs, where they contribute to adhesion of GCNs in the `garland' and to their fusion into binucleate cells. Sns and Kirre proteins localize to the ND at the entry point into the labyrinthine channels and, like their vertebrate counterparts, are essential for its formation. Knockdown of Kirre or Sns drastically reduces the number of NDs at the cell surface....]]>
In holometabolous insects, a species-specific size, known as critical weight, needs to be reached for metamorphosis to be initiated in the absence of further nutritional input. Previously, we found that reaching critical weight depends on the insulin-dependent growth of the prothoracic glands (PGs) in Drosophila larvae. Because the PGs produce the molting hormone ecdysone, we hypothesized that ecdysone signaling switches the larva to a nutrition-independent mode of development post-critical weight. Wing discs from pre-critical weight larvae [5 hours after third instar ecdysis (AL3E)] fed on sucrose alone showed suppressed Wingless (WG), Cut (CT) and Senseless (SENS) expression. Post-critical weight, a sucrose-only diet no longer suppressed the expression of these proteins. Feeding larvae that exhibit enhanced insulin signaling in their PGs at 5 hours AL3E on sucrose alone produced wing discs with precocious WG, CT and SENS expression. In addition, knocking down the Ecdysone receptor (EcR) selectively in the discs also promoted premature WG, CUT and SENS expression in the wing discs of sucrose-fed pre-critical weight...]]>
TRIC channels function as monovalent cation-specific channels that mediate counter ion movements coupled with ryanodine receptor-mediated Ca2+ release from intracellular stores in muscle cells. Mammalian tissues differentially contain two TRIC channel subtypes: TRIC-A is abundantly expressed in excitable cells, whereas TRIC-B is ubiquitously expressed throughout tissues. Here, we report the physiological role of TRIC-B channels in mouse perinatal development. TRIC-B-knockout neonates were cyanotic owing to respiratory failure and died shortly after birth. In the mutant neonates, the deflated lungs exhibited severe histological defects, and alveolar type II epithelial cells displayed ultrastructural abnormalities. The metabolic conversion of glycogen into phospholipids was severely interrupted in the mutant type II cells, and surfactant phospholipids secreted into the alveolar space were insufficient in the mutant neonates. Moreover, the mutant type II cells were compromised for Ca2+ release mediated by inositol-trisphosphate receptors, despite Ca2+ overloading in intracellular stores. Our results indicate that TRIC-B channels take an active part in Ca2+ signalling to establish specialised functions in type II cells and are thus...]]>
The orphan nuclear receptor Nurr1 is essential for the development of meso-diencephalic dopamine (mdDA) neurons and is required, together with the homeobox transcription factor Pitx3, for the expression of genes involved in dopamine metabolism. In order to elucidate the molecular mechanisms that underlie the neuronal deficits in Nurr1-/- mice, we performed combined gene expression microarrays and ChIP-on-chip analysis and thereby identified Dlk1, Ptpru and Klhl1 as novel Nurr1 target genes in vivo. In line with the previously described cooperativity between Nurr1 and Pitx3, we show that the expression of Ptpru and Klhl1 in mdDA neurons is also dependent on Pitx3. Furthermore, we demonstrate that Nurr1 interacts with the Ptpru promoter directly and requires Pitx3 for full expression of Ptpru in mdDA neurons. By contrast, the expression of Dlk1 is maintained in Pitx3-/- embryos and is even expanded into the rostral part of the mdDA area, suggesting a unique position of Dlk1 in the Nurr1 and Pitx3 transcriptional cascades. Expression analysis in Dlk1-/- embryos reveals that Dlk1...]]>
Rho-dependent amoeboid cell movement is a crucial mechanism in both tumor cell invasion and morphogenetic cell movements during fish gastrulation. Amoeboid movement is characterized by relatively non-polarized cells displaying a high level of bleb-like protrusions. During gastrulation, zebrafish mesodermal cells undergo a series of conversions from amoeboid cell behaviors to more mesenchymal and finally highly polarized and intercalative cell behaviors. We demonstrate that Myosin phosphatase, a complex of Protein phosphatase 1 and the scaffolding protein Mypt1, functions to maintain the precise balance between amoeboid and mesenchymal cell behaviors required for cells to undergo convergence and extension. Importantly, Mypt1 has different cell-autonomous and non-cell-autonomous roles. Loss of Mypt1 throughout the embryo causes severe convergence defects, demonstrating that Mypt1 is required for the cell-cell interactions involved in dorsal convergence. By contrast, mesodermal Mypt1 morphant cells transplanted into wild-type hosts undergo dorsally directed cell migration, but they fail to shut down their protrusive behavior and undergo the normal intercalation required for extension. We further show that Mypt1 activity is...]]>
Developmental defects caused by targeted gene inactivation in mice are commonly subject to strain-specific modifiers that modulate the severity of the phenotype. Although several genetic modifier loci have been mapped in mice, the gene(s) residing at these loci are mostly unidentified, and the molecular mechanisms of modifier action remain poorly understood. Mutations in Sox18 cause a variable phenotype in the human congenital syndrome hypotrichosis-lymphedema-telangiectasia, and the phenotype of Sox18-null mice varies from essentially normal to completely devoid of lymphatic vasculature and lethal, depending on the strain of the mice, suggesting a crucial role for strain-specific modifiers in this system. Here we show that two closely related Group F Sox factors, SOX7 and SOX17, are able to functionally substitute for SOX18 in vitro and in vivo. SOX7 and SOX17 are not normally expressed during lymphatic development, excluding a conventional redundancy mechanism. Instead, these genes are activated specifically in the absence of SOX18 function, and only in certain strains. Our studies identify Sox7 and Sox17 as modifiers...]]>
Thisbe (Ths) and Pyramus (Pyr), two closely related Drosophila homologues of the vertebrate fibroblast growth factor (FGF) 8/17/18 subfamily, are ligands for the FGF receptor Heartless (Htl). Both ligands are required for mesoderm development, but their differential expression patterns suggest distinct functions during development. We generated single mutants and found that ths or pyr loss-of-function mutations are semi-lethal and mutants exhibit much weaker phenotypes as compared with loss of both ligands or htl. Thus, pyr and ths display partial redundancy in their requirement in embryogenesis and viability. Nevertheless, we find that pyr and ths single mutants display defects in gastrulation and mesoderm differentiation. We show that localised expression of pyr is required for normal cell protrusions and high levels of MAPK activation in migrating mesoderm cells. The results support the model that Pyr acts as an instructive cue for mesoderm migration during gastrulation. Consistent with this function, mutations in pyr affect the normal segmental number of cardioblasts. Furthermore, Pyr is essential for the specification of even-skipped-positive...]]>
During morphogenesis, cell movements, cell divisions and cell death work together to form complex patterns and to shape organs. These events are the outcome of decisions made by many individual cells, but how these decisions are controlled and coordinated is elusive. The adult abdominal epidermis of Drosophila is formed during metamorphosis by divisions and extensive cell migrations of the diploid histoblasts, which replace the polyploid larval cells. Using in vivo 4D microscopy, we have studied the behaviour of the histoblasts and analysed in detail how they reach their final position and to what extent they rearrange during their spreading. Tracking individual cells, we show that the cells migrate in two phases that differ in speed, direction and amount of cellular rearrangement. Cells of the anterior (A) and posterior (P) compartments differ in their behaviour. Cells near the A/P border are more likely to change their neighbours during migration. The mitoses do not show any preferential orientation. After mitosis, the sisters become preferentially aligned with...]]>
Post-transcriptional control by RNA-binding proteins is a precise way to assure appropriate levels of gene expression. Here, we identify a novel mRNA regulatory system involving Mex3b (RKHD3) and demonstrate its role in FGF signaling. mex3b mRNA has a 3' long conserved UTR, named 3'LCU, which contains multiple elements for both mRNA destabilization and translational enhancement. Notably, Mex3b promotes destabilization of its own mRNA by binding to the 3'LCU, thereby forming a negative autoregulatory loop. The combination of positive regulation and negative autoregulation constitutes a fine-tuning system for post-transcriptional control. In early embryogenesis, Mex3b is involved in anteroposterior patterning of the neural plate. Consistent with this, Mex3b can attenuate FGF signaling and destabilize mRNAs for the FGF signaling components Syndecan 2 and Ets1b through their 3' UTRs. These data suggest that the 3'LCU-mediated fine-tuning system determines the appropriate level of mex3b expression, which in turn contributes to neural patterning through regulating FGF signaling.
]]>Plant organs are generated from meristems throughout development. Patterning and elaboration of organ primordia occur as a result of organized cell division and expansion, processes that are likely to be controlled, in part, by meristem-derived signals. Communication between the meristem and lateral organs is crucial for meristem maintenance and organ patterning, and organ boundaries are thought to be important for mediating this communication. Arabidopsis thaliana LATERAL ORGAN FUSION1 (LOF1) encodes a MYB-domain transcription factor that is expressed in organ boundaries. lof1 mutants display defects in organ separation as a result of abnormal cell division and expansion during early boundary formation. lof1 mutants also fail to form accessory shoot meristems. Mutations in the closely related LATERAL ORGAN FUSION2 (LOF2) gene enhance the lof1 phenotype, such that lof1 lof2 double mutants display additional fusion defects. Genetic interactions with the CUP-SHAPED COTYLEDON genes CUC2 and CUC3 revealed a role for LOF1 in both organ separation and axillary meristem formation. Expression of the meristem determinant STM was reduced in lof1 mutant...]]>
The Polycomb group (PcG) complex is involved in the epigenetic control of gene expression profiles. In flowering plants, PcG proteins regulate vegetative and reproductive programs. Epigenetically inherited states established in the gametophyte generation are maintained after fertilization in the sporophyte generation, having a profound influence on seed development. The gametophyte size and phase dominance were dramatically reduced during angiosperm evolution, and have specialized in flowering plants to support the reproductive process. The moss Physcomitrella patens is an ideal organism in which to study epigenetic processes during the gametophyte stage, as it possesses a dominant photosynthetic gametophytic haploid phase and efficient homologous recombination, allowing targeted gene replacement. We show that P. patens PcG protein FIE (PpFIE) accumulates in haploid meristematic cells and in cells that undergo fate transition during dedifferentiation programs in the gametophyte. In the absence of PpFIE, meristems overproliferate and are unable to develop leafy gametophytes or reach the reproductive phase. This aberrant phenotype might result from failure of the PcG complex to repress proliferation...]]>
During apoptosis, dying cells are quickly internalized by neighboring cells or phagocytes, and are enclosed in phagosomes that undergo a maturation process to generate the phagoslysosome, in which cell corpses are eventually degraded. It is not well understood how apoptotic cell degradation is regulated. Here we report the identification and characterization of the C. elegans tbc-2 gene, which is required for the efficient degradation of cell corpses. tbc-2 encodes a Rab GTPase activating protein (GAP) and its loss of function affects several events of phagosome maturation, including RAB-5 release, phosphatidylinositol 3-phosphate dynamics, phagosomal acidification, RAB-7 recruitment and lysosome incorporation, which leads to many persistent cell corpses at various developmental stages. Intriguingly, the persistent cell corpse phenotype of tbc-2 mutants can be suppressed by reducing gene expression of rab-5, and overexpression of a GTP-locked RAB-5 caused similar defects in phagosome maturation and cell corpse degradation. We propose that TBC-2 functions as a GAP to cycle RAB-5 from an active GTP-bound to an inactive GDP-bound state, which is...]]>
The FGF family of extracellular signaling factors has been proposed to play multiple roles in patterning the telencephalon, the precursor to the cerebrum. In this study, unlike previous ones, we effectively abolish FGF signaling in the anterior neural plate via deletion of three FGF receptor (FGFR) genes. Triple FGFR mutant mice exhibit a complete loss of the telencephalon, except the dorsal midline. Disruption of FGF signaling prior to and coincident with telencephalic induction reveals that FGFs promote telencephalic character and are strictly required to keep telencephalic cells alive. Moreover, progressively more severe truncations of the telencephalon are observed in FGFR single, double and triple mutants. Together with previous gain-of-function studies showing induction of Foxg1 expression and mirror-image duplications of the cortex by exogenous FGF8, our loss-of-function results suggest that, rather than independently patterning different areas, FGF ligands and receptors act in concert to mediate organizer activity for the whole telencephalon.
]]>Eph receptors are widely expressed during cerebral cortical development, yet a role for Eph signaling in the generation of cells during corticogenesis has not been shown. Cortical progenitor cells selectively express one receptor, EphA4, and reducing EphA4 signaling in cultured progenitors suppressed proliferation, decreasing cell number. In vivo, EphA4-/- cortex had a reduced area, fewer cells and less cell division compared with control cortex. To understand the effects of EphA4 signaling in corticogenesis, EphA4-mediated signaling was selectively depressed or elevated in cortical progenitors in vivo. Compared with control cells, cells with reduced EphA4 signaling were rare and mitotically inactive. Conversely, overexpression of EphA4 maintained cells in their progenitor states at the expense of subsequent maturation, enlarging the progenitor pool. These results support a role for EphA4 in the autonomous promotion of cell proliferation during corticogenesis. Although most ephrins were undetectable in cortical progenitors, ephrin B1 was highly expressed. Our analyses demonstrate that EphA4 and ephrin B1 bind to each other, thereby initiating signaling. Furthermore, overexpression of...]]>
Monoaminergic neurons include the physiologically important central serotonergic and noradrenergic subtypes. Here, we identify the zinc-finger transcription factor, Insm1, as a crucial mediator of the differentiation of both subtypes, and in particular the acquisition of their neurotransmitter phenotype. Insm1 is expressed in hindbrain progenitors of monoaminergic neurons as they exit the cell cycle, in a pattern that partially overlaps with the expression of the proneural factor Ascl1. Consistent with this, a conserved cis-regulatory sequence associated with Insm1 is bound by Ascl1 in the hindbrain, and Ascl1 is essential for the expression of Insm1 in the ventral hindbrain. In Insm1-null mutant mice, the expression of the serotonergic fate determinants Pet1, Lmx1b and Gata2 is markedly downregulated. Nevertheless, serotonergic precursors begin to differentiate in Insm1 mutants, but fail to produce serotonin because of a failure to activate expression of tryptophan hydroxylase 2 (Tph2), the key enzyme of serotonin biosynthesis. We find that both Insm1 and Ascl1 coordinately specify Tph2 expression. In brainstem noradrenergic centres of Insm1 mutants, expression...]]>