bluuebunny/crossref_metadata_2025 huggingface dataset:
Anterior mesendoderm induces mouse Engrailed genes in explant cultures
Authors: Ang Siew-Lan , Rossant Janet
Published: April 2021

We have developed germ layer explant culture assays to study the role of mesoderm in anterior-posterior (A-P) patterning of the mouse neural plate. Using isolated explants of ectodermal tissue alone, we have demonstrated that the expression of Engrailed-1 (En-1) and En-2 genes in ectoderm is independent of mesoderm by the midto late streak stage, at least 12 hours before their onset of expression in the neural tube in vivo at the early somite stage. In recombination explants, anterior mesendoderm from headfold stage embryos induces the expression of En-1 and En-2 in preto early streak ectoderm and in posterior ectoderm from headfold stage embryos. In contrast, posterior mesendoderm from embryos of the same stage does not induce En genes in preto early streak ectoderm but is able to induce expression of a general neural marker, neurofilament 160×103Mr. These results provide the first direct evidence for a role of mesendoderm in induction and regionalization of neural tissue in mouse.
Non-neuronal cells inhibit catecholaminergic differentiation of primary sensory neurons: role of leukemia inhibitory factor
Authors: Fan Guoping , Katz David M.
Published: April 2021

Although some sensory ganglion cells in mature animals are catecholaminergic, most mammalian sensory neurons that express the catecholamine-synthesizing enzyme tyrosine hydroxylase (TH) do so only transiently during early gangliogenesis in vivo. The lack of TH expression at later stages appears to be due to modulation of this catecholaminergic potential. A previous study showed that the phenotype reappears, for example, when E16.5 and older sensory ganglia are dissociated in culture into single cells, suggesting that extracellular influences can modulate TH expression. Moreover, TH expression in dissociate cultures is celldensity dependent, as a four-fold increase in plating density led to a 30% decrease in the percentage of TH neurons. The present study demonstrates that inhibition of TH expression in high density cultures is mediated by ganglionic non-neuronal cells (NNC), as removal of NNC abolished density-dependent inhibition. Moreover, plating E16.5 trigeminal neurons at low density on top of NNC monolayers resulted in an 85% decrease in the percentage of TH neurons. Treatment of cultures with non-neuronal cell conditioned medium (NNC–CM) reproduced the effect of coculture with NNC, suggesting that diffusible factors from NNC were involved in the inhibition of TH. The inhibitory effect of NNC–CM was mimicked by treatment of dissociate cultures with ciliary neurotrophic factor (CNTF) and leukemia inhibitory factor (LIF). However, immunoprecipitation of NNC–CM with antibodies against LIF or CNTF showed that only anti-LIF antibodies were able partially to remove the TH inhibitory activity of NNC–CM. Therefore, LIF is one, but not the only, factor mediating NNC inhibition of TH expression in cultured sensory neurons. In summary, these data indicate that ganglionic NNC can regulate sensory transmitter phenotype in culture by inhibiting expression of specific molecular traits. The finding that LIF can partially account for the inhibitory effect of ganglionic NNC on TH expression suggests a novel role for this cytokine in regulating differentiation of catecholaminergic properties in sensory neurons.
Evolutionary change in neural development within the arthropods: axonogenesis in the embryos of two crustaceans
Authors: Whitington Paul M. , Leach David , Sandeman Renate
Published: April 2021

It has been previously suggested that there is a conservative program for neural development amongst the arthropods, on the basis that a stereotyped set of cells involved in establishing the axon tracts in the CNS of insect embryos is also present in crayfish embryos. We have examined the spatiotemporal pattern of axon growth from a set of early differentiating central neurons in the embryo of two crustaceans, the woodlouse Porcellio scaber and the freshwater crayfish Cherax destructor, and drawn comparisons with insect neurons whose somata lie in corresponding positions within the CNS. While many of the woodlouse and crayfish neurons show a similar pattern of axon growth to their insect counterparts, the axon trajectories taken by others differ from those seen in insects. We conclude that this aspect of early neural development has not been rigidly conserved during the evolution of the crustaceans and insects. However, the extent of similarity between the insects and the crustaceans is consistent with the idea that these groups of arthropods share a common evolutionary ‘Bauplan’ for the construction of their nervous systems. While the pattern of early axon growth in the woodlouse and crayfish embryos is sufficiently similar that many neurons could be confidently recognised as homologues, several differences were noted in both the relative order of axon outgrowth and axon morphologies of individual neurons.
Xenopus axis formation: induction of goosecoid by injected Xwnt-8 and activin mRNAs
Authors: Steinbeisser Herbert , Robertis Eddy M. De , Ku Min , Kessler Daniel S. , Melton Douglas A.
Published: April 2021

In this study, we compare the effects of three mRNAs goosecoid, activin and Xwnt-8 that are able to induce partial or complete secondary axes when injected into Xenopus embryos. Xwnt-8 injection produces complete secondary axes including head structures whereas activin and goosecoid injection produce partial secondary axes at high frequency that lack head structures anterior to the auditory vesicle and often lack notochord. Xwnt-8 can activate goosecoid only in the deep marginal zone, i.e., in the region in which this organizerspecific homeobox gene is normally expressed on the dorsal side. Activin B mRNA, however, can turn on goosecoid in all regions of the embryo. We also tested the capacity of these gene products to restore axis formation in embryos in which the cortical rotation was blocked by UV irradiation. Whereas Xwnt-8 gives complete rescue of anterior structures, both goosecoid and activin give partial rescue. Rescued axes including hindbrain structures up to level of the auditory vesicle can be obtained at high frequency even in the absence of notochord structures. The possible functions of Wnt-like and activin-like signals and of the goosecoid homeobox gene, and their order of action in the formation of Spemann’s organizer are discussed.
Puffs and PCR: the in vivo dynamics of early gene expression during ecdysone responses in Drosophila
Authors: Huet François , Ruiz Claude , Richards Geoff
Published: April 2021

The steroid hormone ecdysone orchestrates insect development by regulating gene networks. In Drosophila the most detailed description of ecdysone action is the sequential activation of early and late puffs in the polytene chromosomes of the late larval salivary gland. A number of these early puffs (2B5, 74EF and 75B) contain complex transcription units (Broad-Complex, E74 and E75 respectively) encoding families of regulatory proteins which are expressed in most if not all tissues. In vitro, transcripts of the different isoforms of these early genes as well as the ecdysone receptor (EcR) present varying dose response characteristics (Karim and Thummel, 1992, EMBO J. 11, 4083–4093). We have developed an in vivo approach using a reverse transcription-polymerase chain reaction assay (RT-PCR) so as to visualise these transcripts in the RNA extracted from a single salivary gland. Using one salivary gland lobe for developmental puff staging and the sister lobe for RT-PCR, we have obtained precise developmental profiles for these transcripts and have extended our study to other tissues and stages where puffing studies were not possible. In the salivary gland we have characterised three distinct ecdysone responses. For the mid and late third larval instar responses our results confirm and extend the conclusions of the in vitro studies concerning the temporal expression of the early gene isoforms. The relatively brief prepupal response contains elements in common with each of the larval responses and all three can be explained by the profiles of the respective ecdysone peaks. Interestingly EcR transcripts respond differently during each response. The analysis of different tissues of the same animal reveals subtle differences in the timing of the ecdysone response and isoform expression and suggests that this may reflect tissue differences in the ecdysone profiles. As these molecules have homologues in vertebrates, our analysis may have general implications for the organisation of hormonal responses in vivo.
W-sash affects positive and negative elements controlling c-kit expression: ectopic c-kit expression at sites of kit-ligand expression affects melanogenesis
Authors: Duttlinger Regina , Manova Katia , Chu Tang Y. , Gyssler Corina , Zelenetz Andrew D. , Bachvarova Rosemary F. , Besmer Peter
Published: April 2021

The receptor tyrosine kinase c-kit and its cognate ligand KL are encoded at the white spotting (W) and steel (Sl) loci of the mouse, respectively. Mutations at both the W and the Sl locus cause deficiencies in gametogenesis, melanogenesis and hematopoiesis (erythrocytes and mast cells). The W-sash mutation differs from most W mutations in that it affects primarily mast cells and melanogenesis but not other cellular targets of W and Sl mutations. Thus, Wsh/Wsh mice are fertile and not anemic, but they lack mast cells in their skin and intestine and are devoid of coat pigment. Heterozygotes are black with a broad white sash/belt in the lumbar region. In order to determine the basis for the phenotypes of W-sash mice, we investigated c-kit RNA and protein expression patterns in adult Wsh/Wsh mice and during embryonic development. We show that c-kit expression is absent in bone-marrow-derived Wsh/Wsh mast cells, the fetal and the adult lung, and the digestive tract at embryonic day , tissues that normally express c-kit. Unexpectedly, in and Wsh/Wsh embryos, we found c-kit expression in the dermatome of the somites, the mesenchyme around the otic vesicle and the floorplate of the neural tube, structures known to express the c-kit ligand in wild-type embryos. The ectopic c-kit expression in Wsh homozygous embryos does not affect c-kit ligand expression. The presumed Wsh/Wsh melanoblasts appeared to be normal and, at similar numbers were found in normal and homozygous mutant embryos. At embryos had a graded distribution of melanoblasts from cranial to caudal with a minimum in the lumbar region. Whereas homozygous Wsh/Wsh embryos essentially lacked c-kit-positive cells in the skin, heterozygous Wsh/+ embryos had reduced numbers of melanoblasts compared to +/+ with few or none in the lumbar region (future sash). It is proposed that ectopic c-kit expression in the somitic dermatome affects early melanogensis in a dominant fashion. Molecular analysis of Wsh chromosomal DNA revealed a deletion or rearrangement in the vicinity of the c-kit gene. These results provide an explanation for the Wsh phenotype and have implications for the control of c-kit expression.
A novel, tissue-specific integrin subunit, βν, expressed in the midgut of Drosophila melanogaster
Authors: Yee Gene H. , Hynes Richard O.
Published: April 2021

The integrins are a family of cell surface receptors for extracellular matrix proteins and counter-receptors on other cells. We have used the polymerase chain reaction to identify a novel integrin receptor β subunit in Drosophila melanogaster. The deduced amino acid sequence of this subunit, which we have termed βν (betaneu), indicates that it has several unusual properties. The βν subunit is roughly 33% identical with each of the previously sequenced vertebrate and Drosophila β subunits and is lacking four of the 56 cysteine residues characteristic of most members of this protein family. The expression of the βν gene is strikingly restricted. It is temporally regulated, with maximal expression occurring at 12-15 hours of embryonic development. In situ hybridization analyses and antibody localization on whole-mount embryos reveal that βν expression is tissuespecific and confined to the developing midgut endoderm and its precursors during embryogenesis. Tissue specificity of expression is maintained through later stages of development as βν transcripts are found exclusively in the larval midgut. Within this structure, βν transcripts are especially concentrated in the cells of the midgut imaginal islands which give rise to the adult midgut.
Establishment of gut fate in the E lineage of C. elegans: the roles of lineage-dependent mechanisms and cell interactions
Authors: Goldstein Bob
Published: April 2021

The gut of C. elegans derives from all the progeny of the E blastomere, a cell of the eight cell stage. Previous work has shown that gut specification requires an induction during the four cell stage (Goldstein, B. (1992) Nature 357, 255–257). Blastomere isolation and recombination experiments were done to determine which parts of the embryo can respond to gut induction. Normally only the posterior side of the EMS blastomere contacts the inducing cell, P2. When P2 was instead placed in a random position on an isolated EMS, gut consistently differentiated from the daughter of EMS contacting P2, indicating that any side of EMS can respond to gut induction. Additionally, moving P2 around to the opposite side of EMS in an otherwise intact embryo caused EMS’s two daughter cells to switch lineage timings, and gut to differentiate from the descendents of what normally would be the MS blastomere. The other cells of the four cell stage, ABa, ABp, and P2, did not form gut when placed in contact with the inducer. To determine whether any other inductions are involved in gut specification, timed blastomere isolations were done at the two and eight cell stages. In the absence of cell contact at the two cell stage, segregation of gut fate proceeded normally at both the two and four cell stages. Gut fate also segregated properly in the absence of cell contact at the eight cell stage. A model is presented for the roles of lineage-dependent mechanisms and cell interactions in establishing gut fate in the E lineage.
Foetal testes control the prenatal growth and differentiation of the gubernacular cones in rabbits - a tribute to the late Professor Alfred Jost
Authors: Schoot P. van der
Published: April 2021

Gubernacular cones develop during foetal life in males of various species, including many of the common small laboratory animals. Postnatally these papilla-like organs invert and develop into the muscular cremaster sacs, providing space for testis descent. The mechanism governing male-specific development of these structures during foetal and postnatal life is unknown but foetal testicular androgens or anti-Müllerian hormone are unlikely to be involved. The present study of gubernacular cone development in 28-day-old rabbit foetuses castrated 5–9 days before questions whether foetal testis hormones play any role in these developmental processes. The study comprised an analysis of the microscopic slides in the legacy of the late Professor Alfred Jost in Paris.

Castration at an earlier (19 days) or later (23/24 days) day of foetal life interfered with gubernacular cone growth and differentiation. Unilateral castration partially inhibited ipsilateral gubernacular cone growth. Implantation of a foetal testis close to the ovary could induce male-type gubernacular cone growth in females. Together the data unequivocally support the concept of foetal testicular hormonal control of male-specific gubernacular cone development. Further study is required to unravel the nature of the active foetal testicular agent.
Mesoderm formation in Xenopus ectodermal explants overexpressing Xwnt8: evidence for a cooperating signal reaching the animal pole by gastrulation
Authors: Sokol S. Y.
Published: April 2021

It is demonstrated here that the ability of injected Xwnt8 RNA to trigger mesoderm formation in Xenopus presumptive ectoderm (animal caps) depends on the time of explantation. Animal caps isolated from Xwnt8 injected embryos at the late blastula/early gastrula stages differentiate mesodermal tissues whereas caps isolated from early blastula do not. This finding suggests that an endogenous signal reaches the animal cap by the late blastula stage and cooperates with Xwnt8 to induce mesoderm. Similarly, late animal caps isolated at st. 10 from lithium-treated embryos, but not those from control embryos, elongate and express muscle-specific actin transcripts. In addition, the data presented suggests that the cooperating signal is distributed homo-geneously with respect to the future dorsoventral axis and may require FGF- and activin-dependent signal transduction pathways. These observations support a model in which mesoderm is induced in vivo by a combined action of several different signals.
Stem cell factor induces outgrowth of c-kit-positive neurites and supports the survival of c-kit-positive neurons in dorsal root ganglia of mouse embryos
Authors: Hirata Tatsumi , Morii Eiichi , Morimoto Masahiro , Kasugai Tsutomu , Tsujimura Tohru , Hirota Seiichi , Kanakura Yuzuru , Nomura Shintaro , Kitamura Yukihiko
Published: April 2021

The c-kit receptor tyrosine kinase is highly expressed by about 10% of the neurons in the dorsal root ganglia (DRGs) of mouse embryos. We investigated the in vitro effect of stem cell factor (SCF), the ligand for c-kit receptor, on DRGs. Recombinant murine SCF (rmSCF) induced the outgrowth of c-kit-positive neurites from DRGs of normal (+/+) embryos. The effect of SCF was dose dependent and completely abolished by anti-c-kit ACK2 monoclonal antibody (mAb). Some neurites whose outgrowth was induced by nerve growth factor (NGF) were c-kit-positive, but anti-NGF mAb did not inhibit the rmSCF-induced neurite outgrowth. rmSCF did not induce neurite outgrowth from DRGs of W/W embryos that did not express c-kit receptors on the cell surface and of W42/W42 mutant embryos that expressed c-kit receptors without tyrosine kinase activity. rmSCF also had a trophic effect on c-kit-positive neurons in the culture of dissociated DRG cells. Most c-kit-positive neurons appeared to respond to NGF as well, and the SCF-responsive subpopulation represented about 10% of NGF-responsive neurons. rmSCF did not support the survival of DRG neurons from embryos of W/W and W42/W42 genotypes. These results suggest that the stimulus through the c-kit receptor tyrosine kinase has an important role in development of the peripheral nervous system.
Novel genes expressed differentially in ascidians with alternate modes of development
Authors: Swalla Billie J. , Makabe Kazuhiro W. , Satoh Noriyuki , Jeffery William R.
Published: April 2021

We have used a subtractive procedure to isolate cDNA clones encoding genes expressed differentially in ascidian species with alternate modes of development. The ascidians used in this study were Molgula oculata, which develops a tailed (urodele) larva, and Molgula occulta, which develops a tailless (anural) larva. Two of the identified clones, Uro-2 and Uro-11, are described. Southern blots show that the Uro-2 and Uro-11 genes are present in both species, but the corresponding mRNAs are expressed preferentially in the urodele species. In situ hybridization showed that Uro-2 and Uro-11 transcripts accumulate in small oocytes during oogenesis. The maternal Uro-2 and Uro-11 transcripts were distributed throughout the oocyte cytoplasm. Transcript concentrations declined during vitellogenesis, but mature eggs still contain detectable levels of Uro-2 and Uro-11 mRNA. After fertilization, the maternal Uro-2 and Uro-11 transcripts were localized in the ectoplasm of uncleaved zygotes and mostly entered the ectoderm cells during cleavage. The Uro-2 gene appears to produce only maternal transcripts. In contrast, the Uro-11 gene may also produce zygotic transcripts, which accumulate between gastrulation and neurulation in posterior epidermis, neural and tail muscle cells. Zygotic expression of the Uro-11 gene was not detected in embryos of the anural species. The deduced amino acid sequences of the Uro-2 and Uro-11 cDNAs suggest that they encode novel basic proteins with distinctive structural features. The predicted Uro-2 protein contains a leucine zipper motif, suggesting that it may dimerize with another protein. The predicted Uro-11 protein contains a nuclear localization signal, a region with similarity to part of the DNA-binding motif in the bacterial histone-like HU and IHF proteins, 12 repeats of the proposed DNA-binding motif S(T)PXX, and a potential zinc finger of the C6 or C6H2 class, suggesting that it may be a DNA-binding protein. The Uro-2 and Uro-11 proteins are candidates for regulatory factors involved in the evolutionary transition from urodele to anural development.
Nkx-2.5: a novel murine homeobox gene expressed in early heart progenitor cells and their myogenic descendants
Authors: Lints Thierry J. , Parsons Linda M. , Hartley Lynne , Lyons Ian , Harvey Richard P.
Published: April 2021

We have isolated two murine homeobox genes, Nkx-2.5 and Nkx-2.6, that are new members of a sp sub-family of homeobox genes related to Drosophila NK2, NK3 and NK4/msh-2. In this paper, we focus on the Nkx-2.5 gene and its expression pattern during post-implantation development. Nkx-2.5 transcripts are first detected at early headfold stages in myocardiogenic progenitor cells. Expression preceeds the onset of myogenic differentiation, and continues in cardiomyocytes of embryonic, foetal and adult hearts. Transcripts are also detected in future pharyngeal endoderm, the tissue believed to produce the heart inducer. Expression in endoderm is only found laterally, where it is in direct apposition to promyocardium, suggesting an interaction between the two tissues. After foregut closure, Nkx-2.5 expression in endoderm is limited to the pharyngeal floor, dorsal to the developing heart tube. The thyroid primordium, a derivative of the pharyngeal floor, continues to express Nkx-2.5 after transcript levels diminish in the rest of the pharynx. Nkx-2.5 transcripts are also detected in lingual muscle, spleen and stomach.

The expression data implicate Nkx-2.5 in commitment to and/or differentiation of the myocardial lineage. The data further demonstrate that cardiogenic progenitors can be distinguished at a molecular level by late gastrulation. Nkx-2.5 expression will therefore be a valuable marker in the analysis of mesoderm development and an early entry point for dissection of the molecular basis of myogenesis in the heart.
Segment polarity gene interactions modulate epidermal patterning in Drosophila embryos
Authors: Bejsovec Amy , Wieschaus Eric
Published: April 2021

Each segment of a Drosophila larva shows a precisely organized pattern of cuticular structures, indicating diverse cellular identities in the underlying epidermis. Mutations in the segment polarity genes alter the cuticle pattern secreted by the epidermal cells; these mutant patterns provide clues about the role that each gene product plays in the development of wild-type epidermal pattern. We have analyzed embryos that are multiply mutant for five key patterning genes: wingless, patched, engrailed, naked and hedgehog. Our results indicate that wild-type activity of these five segment polarity genes can account for most of the ventral pattern elements and that their gene products interact extensively to specify the diverse cellular identities within the epidermis. Two pattern elements can be correlated with individual gene action: wingless is required for formation of naked cuticle and engrailed is required for formation of the first row of denticles in each abdominal denticle belt. The remaining cell types can be produced by different combinations of the five gene activities. wingless activity generates the diversity of cell types within the segment, but each specific cell identity depends on the activity of patched, engrailed, naked and hedgehog. These molecules modulate the distribution and interpretation of wingless signalling activity in the ventral epidermal cells and, in addition, each can contribute to pattern through a pathway independent of the wingless signalling pathway
Expression of the chicken GATA factor family during early erythroid development and differentiation
Authors: Leonard Mark W. , Lim Kim-Chew , Engel James Douglas
Published: April 2021

The DNA motif WGATAR has been identified within transcriptional regulatory domains of globin and other erythroid-specific genes and the activator proteins that bind to this regulatory element, the GATA factors, belong to a multi-gene family that is expressed in chicken erythroid cells. Here we show that, as in chickens, multiple members of the GATA factor family are expressed in human and murine erythroid cells. During the early stages of chicken embryogenesis (well before blood island formation), each of the GATA family members is transcribed with a unique temporal and spatial pattern. In the primitive erythroid lineage, tran-scription of the embryonic -globin gene parallels GATA-1 expression while the switch to -globin transcription in definitive erythroid cells is directly preceded by a pronounced increase in GATA-3 accumulation. The timing and pattern of expression of these different mRNAs during avian erythroid development and differentiation suggests that temporally regulated changes in GATA factor expression are required for vertebrate hematopoiesis.
Prestalk cells in monolayer cultures exhibit two distinct modes of cellulose synthesis during stalk cell differentiation in Dictyostelium
Authors: Blanton R. L.
Published: April 2021

Stalk formation in Dictyostelium discoideum involves the synthesis of a stalk tube by the prestalk cell population and stalk cell walls by the individual prestalk cells. Cellulose is a major structural component of the stalk tube and stalk cell walls. The DIF-deficient strain HM44 was used to study the events of stalk formation in monolayer cultures. The induction of cellulose synthase activity was shown to require both DIF and cAMP. Microscopical observations of monolayer cultures using the cellulose-indicating fluorochrome Tinopal LPW demonstrated the presence in these cultures of two cellulose-containing materials: the stalk cell walls and an intercellular material found between cells and around cell clumps. The synthesis of intercellular material precedes that of stalk cell walls in induced cultures. Cells committed to stalk cell formation were delayed in doing so if they were switched to medium containing cAMP but no DIF. During this delay the cells synthesized large quantities of the intercellular material. The intercellular material was shown to be microfibrillar, was sensitive to cellulase, and labelled with a colloidal gold-conjugated cellulase. The intercellular material may have the same mode of cellulose synthesis as that involved in stalk tube formation. If so, that mode would be favored by DIF and cAMP in combination, whereas the cellulose synthesis involved in stalk cell wall formation would be DIF-dependent but delayed or repressed by cAMP.
Expression of Drosophila glass protein and evidence for negative regulation of its activity in non-neuronal cells by another DNA-binding protein
Authors: Ellis Michael C. , O’Neill Elizabeth M. , Rubin Gerald M.
Published: April 2021

The glass gene encodes a DNA-binding zinc-finger protein required for the development of Drosophila photoreceptor cells and which appears to regulate a number of genes specifically expressed in photoreceptors. We have generated monoclonal antibodies to Glass and used them to examine Glass distribution during development. Glass is expressed in all cell types of the developing eye and in all other organs that contain photoreceptor cells in Drosophila, including a small number of cells in the brain. We altered the normal pattern of glass expression by placing the gene under the control of the hsp70 promoter. Our results suggest that nonphotoreceptor cells are restricted in their response to Glass expression. In an effort to discover the mechanism of this restriction, we examined the expression of a number of reporter gene constructs. Our results suggest that nonsensory cells are unable to express certain reporter constructs in response to Glass expression because another DNA-binding factor represses Glass activity in nonsensory cells.
Constitutive expression of a somatic heat-inducible hsp70 gene during amphibian oogenesis
Authors: Billoud Bernard , Rodriguez-Martin Maria-Luz , Berard Line , Moreau Nicole , Laine Marie-Christine , Angelier Nicole
Published: April 2021

We isolated and characterized a sequence coding for heat-shock protein 70 (HSP70) of the amphibian Pleu-rodeles waltl. Results from S1 nuclease protection assays led us to conclude that an hsp70 gene, strictly inducible in somatic cells during heat shock, is constitutively active during oogenesis. By quantitative northern and western blot analysis, we showed that both hsp70 mRNA and HSP70-related protein levels increased in oocytes from stage II to stage VI under physiological conditions. Furthermore, by in situ hybridization to the nascent transcripts of lampbrush chromosome loops, we provided evidence for a clear-cut relationship between this increase in hsp70 mRNA and transcriptional activity during the lampbrush stage of oogenesis. These results strongly suggest that hsp70 genes are actively transcribed throughout oogenesis. HSP70-related proteins localized in the cytoplasm of young oocytes are progressively transferred to the nucleus in the course of oogenesis and preferentially accumulated in the nuclei of some stage VI oocytes.
Expression of the c-ret proto-oncogene during mouse embryogenesis
Authors: Pachnis Vassilis , Mankoo Baljinder , Costantini Frank
Published: April 2021

The c-ret proto-oncogene encodes a receptor tyrosine kinase whose normal function has yet to be determined. To begin to investigate the potential role of this gene in vertebrate development, we have isolated cDNA clones representing the murine c-ret gene, and have analyzed the pattern of expression during mouse embryogenesis, using northern blotting, in situ hybridization to histological sections and whole-mount hybridization histochemistry. c-ret transcripts were detected beginning at day 8.5 of embryogenesis, and were observed in a number of cell lineages in the developing peripheral and central nervous systems, as well as in the excretory system. In the cranial region at day 8.5-9.5, c-ret mRNA was restricted to a population of neural crest cells migrating from rhombomere 4 and forming the anlage of the facioacoustic ganglion, as well as to a closely associated domain of surface ectoderm and pharyngeal endoderm. At later stages (10.5–14.5 days), c-ret mRNA was observed in all cranial ganglia. In the peripheral nervous system of the trunk, c-ret was expressed in the autonomic ganglia and in subsets of cells in the dorsal root ganglia. In the enteric nervous system, c-ret was expressed in the presumptive enteric neuroblasts of the vagal crest (day 9.0–11.5), and in the myenteric ganglia of the gut (day 13.5–14.5). c-ret mRNA was observed in several regions of the central nervous system, including the undifferentiated neuroepithelial cells of the ventral neural tube (8.5 days), the motor neurons in the spinal cord and the hindbrain (10.5–14.5 days), the embryonic neuroretina (day 13.5) and the layers of the postnatal retina containing ganglion, amacrine and horizontal cells. Outside the nervous system, c-ret was expressed in the nephric (Wolffian) duct at day 8.5–10.5, the ureteric bud epithelium (but not the surrounding metanephric mesenchyme) at day 11.0–11.5, and the growing tips of the renal collecting ducts (but not the previously formed, subcortical portions of the collecting ducts, or the mesenchyme-derived renal vesicles) at day 13.5–17.5. Our results suggest that the c-ret gene may encode the receptor for a factor involved in the proliferation, migration, differentiation or survival of a variety of neuronal cell lineages, as well as in inductive interactions during organogenesis of the kidney.
The developing organ of Corti contains retinoic acid and forms supernumerary hair cells in response to exogenous retinoic acid in culture
Authors: Kelley Matthew W. , Xu Xiao-Mei , Wagner Michael A. , Warchol Mark E. , Corwin Jeffrey T.
Published: April 2021

The mammalian organ of Corti has one of the most highly ordered patterns of cells in any vertebrate sensory epithelium. A single row of inner hair cells and three or four rows of outer hair cells extend along its length. The factors that regulate the formation of this strict pattern are unknown. In order to determine whether retinoic acid plays a role during the development of the organ of Corti, exogenous retinoic acid was added to embryonic mouse cochleae in vitro. Exogenous retinoic acid significantly increased the number of cells that developed as hair cells and resulted in large regions of supernumerary hair cells and supporting cells containing two rows of inner hair cells and up to 11 rows of outer hair cells. The effects of retinoic acid were dependent on concentration and on the timing of its addition.

Western blot analysis indicated that cellular retinoic acid binding protein (CRABP) was present in the sensory epithelium of the embryonic cochlea. The amount of CRABP apparently increased between embryonic day 14 and postnatal day 1, but CRABP was not detectable in sensory epithelia from adults. A retinoic acid reporter cell line was used to demonstrate that retinoic acid was also present in the developing organ of Corti between embryonic day 14 and postnatal day 1, and was also present in adult cochleae at least in the vicinity of the modiolus. These results suggest that retinoic acid is involved in the normal development of the organ of Corti and that the effect of retinoic acid may be to induce a population of prosensory cells to become competent to differentiate as hair cells and supporting cells.
The formation and maintenance of the definitive endoderm lineage in the mouse: involvement of HNF3/forkhead proteins
Authors: Ang Siew-Lan , Wierda Allison , Wong David , Stevens Kimberly A. , Cascio Stephanie , Rossant Janet , Zaret Kenneth S.
Published: April 2021

Little is known about genes that govern the development of the definitive endoderm in mammals; this germ layer gives rise to the intestinal epithelium and various other cell types, such as hepatocytes, derived from the gut. The discovery that the rat hepatocyte transcription factor HNF3 is similar to the Drosophila forkhead gene, which plays a critical role in gut development in the fly, led us to isolate genes containing the HNF3/forkhead (HFH) domain that are expressed in mouse endoderm develop ment. We recovered mouse HNF3β from an embryo cDNA library and found that the gene is first expressed in the anterior portion of the primitive streak at the onset of gastrulation, in a region where definitive endoderm first arises. Its expression persists in axial structures derived from the mouse equivalent of Hensen’s node, namely definitive endoderm and notochord, and in the ventral region of the developing neural tube. Expression of the highly related gene, HNF3α, appears to initiate later than HNF3P and is first seen in midline endoderm cells. Expression. subsequently appears in notochord, ventral neural tube, and gut endoderm in patterns similar to HNF3β. Microscale DNA binding assays show that HNF3 proteins are detectable in the midgut at 9.5 days p.c. At later stages HNF3 mRNAs and protein are expressed strongly in endoderm-derived tissues such as the liver. HNF3 is also the only known hepatocyte enriched transcription factor present in a highly de-dif ferentiated liver cell line that retains the capacity to re differentiate to the hepatic phenotype. Taken together, these studies suggest that HNF3α and HNF3β are involved in both the initiation and maintenance of the endodermal lineage. We also discovered a novel HFH containing gene, HFH-ES.1, that is expressed transiently in posterior ectoderm and mesoderm at the primitive streak stage, and later predominantly in the neural tube. HFH-ES.1 is highly similar in structure and expression profile to the Drosophila HFH gene FD4, suggesting that HFH family members have different, evolutionarily conserved roles in development.
The role of the Enhancer of split complex during cell fate determination in Drosophila
Authors: Tata Fred , Hartley David A.
Published: April 2021

Molecular and genetic data predict that the Enhancer of split locus functions at the end of a pathway dictating appropriate cell fate determination in a number of developmental contexts. We have sought to dissect the role individual member genes of the complex play through a molecular analysis. Of the two principle class of genes, the first, members of the basic helix-loop-helix (bHLH) class of proteins are expressed in specific regions of the embryo in subtle, overlapping patterns in cells that will differentiate as epidermis. The second, groucho, a member of the WD40 class of proteins, is expressed more generally. Immunoprecipitation experiments do not implicate groucho in G protein mediated signal transduction, a known function of many WD40 type proteins. Instead, the nuclear localisation of the protein suggests a relationship to the bHLH members of the complex. Differences in expression of the bHLH genes between neurogenic mutants implies two pathways to their activation during epidermal determination.
Integrins and morphogenesis
Authors: Brown Nicholas H. , Bloor James W. , Dunin-Borkowski Olga , Martin-Bermudo M. Dolores
Published: April 2021

The Drosophila position specific (PS) integrins consist of two cell surface heterodimers, PS1 (αPS1βPS) and PS2 (αPS2βPS), which are expressed on complementary sides of attachments between cell layers and are essential for these attachments. Current evidence suggests that the PS integrins bind to components of the extracellular matrix, similar to the majority of vertebrate integrins, but specific Drosophila ligands have not yet been identified. In the embryo PS1 is found on the surface of the epidermis and endoderm, while PS2 is restricted to the mesoderm. The integrins are concentrated at the sites where the somatic muscles attach to the epidermis and at the interface between the visceral mesoderm and the endoderm. In myospheroid mutant embryos, which lack the βPS subunit, the adhesion between the mesoderm and the other cell layers fails. The PS integrins are also required for the adhesion of the dorsal to the ventral surface of the wing during metamorphosis. PS1 is expressed on the basal surface of the dorsal cells and PS2 is expressed on the ventral cells. Loss of PS integrin function in the wing results in balloon shaped wings because of the failure of the two surfaces of the wing blade to adhere to each other. These and other aspects of the phenotypes of mutations in the genes encoding the PS integrins indicate that integrins play an important role in the adhesion of different cell layers to each other and thus an essential role in the morphogenesis of the organism. The use of extracellular matrix receptors in this role may aid in keeping the different cell layers distinct.
Glycoconjugates mark a transient barrier to neural crest migration in the chicken embryo
Authors: Oakley R. A. , Lasky C. J. , Erickson C. A. , Tosney K. W.
Published: April 2021

We report that two molecular markers correlate with a transient inhibition of neural crest cell entry into the dorsolateral path between the ectoderm and the somite in the avian embryo. During the period when neural crest cells are excluded from the dorsolateral path, both peanut agglutinin lectin (PNA)-binding activity and chondroitin-6-sulfate (C6S) immunoreactivity are expressed within this path. Both markers decline as neural crest cells enter. Moreover, both markers are absent after an experimental manipulation that accelerates neural crest entry into this path. Specifically, dermamyotome deletions abolish expression of both markers and allow neural crest cells to enter the dorsolateral path precociously. After partial deletions, dermatome remnants remain. These remnants retain PNA and C6S labeling and impede migration locally. Local glycoconjugate expression thus correlates directly with avoidance responses. Since both PNA-binding activity and C6S expression also typify inhibitory somitic tissues, molecules indicated by these markers (or co-regulated molecules) are likely to inhibit both neural crest and axon advance.
Function of the retinoic acid receptors (RARs) during development: (I) Craniofacial and skeletal abnormalities in RAR double mutants
Authors: Lohnes David , Mark Manuel , Mendelsohn Cathy , Dollé Pascal , Dierich Andrée , Gorry Philippe , Gansmuller Anne , Chambon Pierre
Published: April 2021

Numerous congenital malformations have been observed in fetuses of vitamin A-deficient (VAD) dams [Wilson, J. G., Roth, C. B., Warkany, J., (1953), Am. J. Anat. 92, 189-217]. Previous studies of retinoic acid receptor (RAR) mutant mice have not revealed any of these malformations [Li, E., Sucov, H. M., Lee, K.-F., Evans, R. M., Jaenisch, R. (1993) Proc. Natl. Acad. Sci. USA 90, 1590-1594; Lohnes, D., Kastner, P., Dierich, A., Mark, M., LeMeur, M., Chambon, P. (1993) Cell 73, 643-658; Lufkin, T., Lohnes, D., Mark, M., Dierich, A., Gorry, P., Gaub, M. P., LeMeur, M., Chambon, P. (1993) Proc. Natl. Acad. Sci. USA 90, 7225-7229; Mendelsohn, C., Mark, M., Dollé, P., Dierich, A., Gaub, M.P., Krust, A., Lampron, C., Chambon, P. (1994a) Dev. Biol. in press], suggesting either that there is a considerable functional redundancy among members of the RAR family during ontogenesis or that the RARs are not essential transducers of the retinoid signal in vivo. In order to discriminate between these possibilities, we have generated a series of RAR compound null mutants. These RAR double mutants invariably died either in utero or shortly after birth and presented a number of congenital abnormalities, which are reported in this and in the accompanying study. We describe here multiple eye abnormalities which are found in various RAR double mutant fetuses and are similar to those previously seen in VAD fetuses. Interestingly, we found further abnormalities not previously reported in VAD fetuses. These abnormalities affect ocular glands, salivary glands and their associated ducts, the axial and limb skeleton, and all skeletal elements derived from the mesectoderm of the frontonasal mass and of the second and third pharyngeal arches. RAR double mutants also exhibit supernumerary cranial skeletal elements that are present in the ancestral reptilian skull. The role of retinoic acid (RA) and of the RARs in the ontogenesis of the affected structures, particularly of those that are derived from mesenchymal neural crest cells, is discussed.
Function of the retinoic acid receptors (RARs) during development: (II) Multiple abnormalities at various stages of organogenesis in RAR double mutants
Authors: Mendelsohn Cathy , Lohnes David , Décimo Didier , Lufkin Thomas , LeMeur Marianne , Chambon Pierre , Mark Manuel
Published: April 2021

Compound null mutations of retinoic acid receptor (RAR) genes lead to lethality in utero or shortly after birth and to numerous developmental abnormalities. In the accompanying paper (Lohnes, D., Mark, M., Mendelsohn, C., Dollé, P., Dierich, A., Gorry, Ph., Gansmuller, A. and Chambon, P. (1994). Development 120, 2723-2748), we describe malformations of the head, vertebrae and limbs which, with the notable exception of the eye defects, were not observed in the offspring of vitamin A-deficient (VAD) dams. We report here abnormalities in the neck, trunk and abdominal regions of RAR double mutant mice, which include :(i) the entire respiratory tract, (ii) the heart, its outlow tract and the great vessels located near the heart, (iii) the thymus, thyroid and parathyroid glands, (iv) the diaphragm, (v) the genito-urinary system, and (vi) the lower digestive tract. A majority of these abnormalities recapitulate those observed in the fetal VAD syndrome described by Joseph Warkany’s group more than fourty years ago [Wilson, J. G., Roth, C. B. and Warkany, J. (1953) Am. J. Anat., 92, 189-217; and refs therein]. Our results clearly demonstrate that RARs are essential for vertebrate ontogenesis and therefore that retinoic acid is the active retinoid, which is required at several stages of the development of numerous tissues and organs. We discuss several possibilities that may account for the apparent functional redundancy observed amongst retinoic acid receptors during embryogenesis.
Regulation of motor neuron dendrite growth by NMDA receptor activation
Authors: Kalb Robert G.
Published: April 2021

Spinal motor neurons undergo great changes in morphol-ogy, electrophysiology and molecular composition during development. Some of this maturation occurs postnatally when limbs are employed for locomotion, suggesting that neuronal activity may influence motor neuron develop-ment. To identify features of motor neurons that might be regulated by activity we first examined the structural devel-opment of the rat motor neuron cell body and dendritic tree labeled with cholera toxin-conjugated horseradish peroxidase. The motor neuron cell body and dendrites in the radial and rostrocaudal axes grew progressively over the first month of life. In contrast, the growth of the dendritic arbor/cell and number of dendritic branches was biphasic with overabundant growth followed by regression until the adult pattern was achieved. We next examined the influence of neurotransmission on the development of these motor neuron features. We found that antagonism of the N-methyl-D-aspartate (NMDA) subtype of glutamate receptor inhibited cell body growth and dendritic branching in early postnatal life but had no effect on the maximal extent of dendrite growth in the radial and ros-trocaudal axes. The effects of NMDA receptor antagonism on motor neurons and their dendrites was temporally restricted; all of our anatomic measures of dendrite structure were resistant to NMDA receptor antagonism in adults. These results suggest that the establishment of mature motor neuron dendritic architecture results in part from dendrite growth in response to afferent input during a sensitive period in early postnatal life.
Mesoderm induction by activin requires FGF-mediated intracellular signals
Authors: LaBonne Carole , Whitman Malcolm
Published: April 2021

We have examined the role of FGF signaling during activin-mediated mesoderm induction in Xenopus. Using dominant inhibitory mutants of FGF signal transducers to disrupt the FGF-signaling pathway at the plasma membrane or in the cytosol prevents animal cap blastomeres from expressing several mesodermal markers in response to exogenous activin. Dominant inhibitory mutants of the FGF receptor, c-ras or c-raf inhibit the ability of activin to induce molecular markers of both dorsal and ventral mesoderm including Xbra, Mix1 and Xnot. Some transcriptional responses to activin such as goosecoid and Xwnt8 are inhibited less effectively than others, however, suggesting that there may differing requirements for an FGF signal in the responses of mesoderm-specific genes to activin induction. Despite the requirement for this signaling pathway during activin induction, downstream components of this pathway are not activated in response to activin, suggesting that activin does not signal directly through this pathway.