bluuebunny/crossref_metadata_2025 huggingface dataset:
Collagens in avian neural crest development: distribution in vivo and migration-promoting ability in vitro
Authors: Perris Roberto , Krotoski Danuta , Bronner-Fraser Marianne
Published: April 2021

This study examines the spatiotemporal distribution of collagen (Col) types I-V and IX during neural crest development in vivo and their ability to support neural crest cell movement in vitro. Col I, HI and IV were widespread throughout the embryo, including the neural crest migratory pathways, whereas Col H, V and IX preferentially localized to regions from which migrating neural crest cells were absent. Col I-IV and IX occurred both in association with basement membranes and within interstitial matrices, whereas Col V only was detected in juxtaposition to basement membranes. Although initially distributed throughout the rostrocaudal extent of the somitic sclerotome, Col I and HI rearranged to the caudal portion with progressive neural crest cell migration through the rostral portion of the sclerotome. This rearrangement does not occur in neural crest-ablated embryos, suggesting that it is a direct consequence of neural crest cell migration. The perinotochordal matrix, avoided by neural crest cells, contained a metameric Col H/IX immunoreactivity along the rostrocaudal axis which alternated with that of Col I and HI. In contrast, Col IV and V were not observed in this matrix, but lined the basement membranes of the notochord and ventrolateral neural tube. To determine their functional significance for neural crest cell migration in vivo, purified collagens were tested for their ability to promote neural crest cell motility in vitro.

Neural crest cell migration on isolated collagens was most pronounced on Col I and IV, whereas Col H, V and the triple-helical fragment of Col VH were unable to support cell motility. Substrata created by copolymerization of Col I and fibronectin, or Col I and laminin-nidogen, supported cell motility better than Col I alone, whereas both Col V and a cartilage-type chondroitin sulfate proteoglycan reduced cell movement on Col I. Fibronectin bound to pre-immobilized monomeric Col I, H or V had a reduced ability to support neural crest cell movement when compared to fibronectin alone. A similar reduction was seen for Col IV bound to the low density heparan sulfate proteoglycan from the EHS mouse tumor. The results demonstrate that Col I-IX are differentially distributed in the early avian embryo. During neural crest development several of these collagens undergo dynamic reorganizations that correlate with the migration of neural crest cells. Furthermore, various collagens possess distinct abilities to support neural crest cell migration in vitro, and their migration-promoting activity can be modulated by their conformation and/or association with other matrix components.
Contiguous patterns of c-kit and steel expression: analysis of mutations at the W and Sl loci
Authors: Motro Benny , Kooy Derek Van Der , Rossant Janet , Reith Alastair , Bernstein Alan
Published: April 2021

Mutations in either the dominant white-spotting (W) or Steel (Sl) loci of the mouse lead to coat color, primordial germ cell and hematopoietic defects. Consistent with the cell autonomous and microenvironmental nature of W and Sl mutations, respectively, it has recently been shown that W encodes the c-kit receptor tyrosine kinase while Sl encodes a ligand for this receptor. Previous in situ hybridization analysis has shown that both c-kit and steel are expressed in the embryo in anatomical sites known to be affected by W and Sl mutations and in various tissues in which no corresponding phenotype has been described. To investigate the possible involvement of the Kit transduction pathway in developmental processes, we compared the patterns of expression of c-kit and steel in wild-type embryos and in embryos homozygous for severe (lethal) and mild (viable) alleles at the W and Sl loci. In addition, we analyzed the patterns of expression of both genes in adult wild-type and mutant gonads and brain. Both c-kit and steel are contiguously expressed in a wide variety of anatomical locations in both the developing embryo and in the adult. In adult gonads, steel is expressed in the follicular cells of the ovary and in Sertoli cells of the testis, the layers that immediately surround the c-kit expressing germ cells. In adult brain, the complementary patterns are particularly striking in the olfactory bulb, cerebral cortex, hippocampus region and cerebellum, steel expression in brain is probably restricted to neurons in certain areas, while c-kit is expressed in neurons and in some glial cells. Severe mutations in the W or Sl loci result in dramatic reduction or absence of c-kit positive cells in lineages known to be affected by these mutations. In contrast, these mutations do not affect the number or histological organization of c-kit positive cells in the embryonic peripheral or central nervous systems, nor is the number or organization of c-kit positive cells detectably altered in Wv/Wv or Sf’/S adult brain. Taken together, these results suggest that the Kit signaling pathway is not obligatory for the viability and/or migration of most c-kit expressing cells either because of functional redundancy with another signaling pathway or because the Kit pathway is involved in post-developmental processes of mature cells.
Notochord morphogenesis in Xenopus laevis: simulation of cell behavior underlying tissue convergence and extension
Authors: Weliky Michael , Minsuk Steve , Keller Ray , Oster George
Published: April 2021

Cell intercalation and cell shape changes drive notochord morphogenesis in the African frog, Xenopus laevis. Experimental observations show that cells elongate mediolaterally and intercalate between one another, causing the notochord to lengthen and narrow. Descriptive observations provide few clues as to the mechanisms that coordinate and drive these cell movements. It is possible that a few rules governing cell behavior could orchestrate the shaping of the entire tissue. We test this hypothesis by constructing a computer model of the tissue to investigate how rules governing cell motility and cell-cell interactions can account for the major features of notochord morphogenesis. These rules are drawn from the literature on in vitro cell studies and experimental observations of notochord cell behavior. The following types of motility rules are investigated: (1) refractory tissue boundaries that inhibit cell motility, (2) statistical persistence of motion, (3) contact inhibition of protrusion between cells, and (4) polarized and nonpolarized protrusive activity. We show that only the combination of refractory boundaries, contact inhibition and polarized protrusive activity reproduces normal notochord development. Guided by these rules, cells spontaneously align into a parallel array of elongating cells. Self alignment optimizes the geometric conditions for polarized protrusive activity by progressively minimizing contact inhibition between cells. Cell polarization, initiated at refractory tissue boundaries, spreads along successive cell rows into the tissue interior as cells restrict and constrain their neighbors’ directional bias. The model demonstrates that several experimentally observed intrinsic cell behaviors, operating simultaneously, may underlie the generation of coordinated cell movements within the developing notochord.
The prospero gene encodes a divergent homeodomain protein that controls neuronal identity in Drosophila
Authors: Chu-Lagraff Quynh , Wright Dorothy M. , McNeil Leslie Klis , Doe Chris Q.
Published: April 2021

The Drosophila central nervous system (CNS) develops from a population of stem cells called neuroblasts; each neuroblast goes through an invariant cell lineage to produce a characteristic family of neurons or glia. We are interested in the molecular mechanisms controlling neuroblast cell lineage. Recently we identified the prospero (pros) gene, which is expressed in embryonic neuroblasts. Loss of pros function results in aberrant expression of the homeobox genes fushi tarazu, evenskipped and engrailed in a subset of neuroblast progeny, suggesting that pros plays an early and fundamental role in the specification of neuronal fate (Doe et al. 1991). Here we show that the pros gene encodes a highly divergent homeodomain. The homeodomain contains several of the most conserved amino acids characteristic of known homeodomains, yet it is considerably less basic than previously identified homeodomains. These data are consistent with a model in which pros controls neuroblast cell lineages by regulating gene expression.
The early development of the frog retinotectal projection
Authors: Taylor Jeremy S. H.
Published: April 2021

The guidance of retinal ganglion cell axons has been investigated in embryos of the frog Xenopus. During the initial development of the brain a series of axon tracts are laid down forming a basic ‘scaffold’ or framework. Retinal axons grow through one of these tracts, the tract of the post-optic commissure (tPOC). This is the only tract that extends through the rostral part of the brain at these early stages of development. The origin and development of the tPOC has been studied using antibodies which label neurons at their earliest stages of differentiation. The first sign of the tPOC is a chain of neurons which differentiate simultaneously in the caudolateral part of the diencephalon. Axons from these neurons grow the short distance between adjacent cells interlinking the chain to form a descending tract. A series of other axon projections are then added to the tPOC, each of which is segregated into a particular subregion of the tract. Retinal axons are added to the tract approximately 18 h after its formation. They grow in the sub-pial part of the tract and always occupy the rostral-most edge. Retinal axons follow the tract to the region of the developing tectum where they leave, turn dorsally, and terminate.

The reliance of retinal axons on this pre-existing pathway has been demonstrated by experimentally altering the course of the tPOC during its early development. The caudo-lateral wall of the diencephalon has been rotated through 90° at a stage just before the tPOC neurons differentiate. Confirmation of the predicted alteration in the course of the tPOC has been made using immunocytochemistry. In such manipulated brains, retinal axons maintain their strong affinity for the rostral edge of the tPOC, following its altered course through the diencephalon.
Expression patterns of c-myb and of v-myb induced myeloid-1 (mim-1) gene during the development of the chick embryo
Authors: Quéva Christophe , Ness Scott A. , Grässer Frederick A. , Graf Thomas , Vandenbunder Bernard , Stéhelin Dominique
Published: April 2021

The v-myb oncogene of the acute avian leukemia virus E26 encodes a transcription factor that directly regulates the promyelocyte-specific mim-1 gene (Ness, S.A., Marknell, A. and Graf, T. Cell, 59,1115–1125). We have investigated the relationship between the c-myb proto-oncogene and the transcription of the mim-1 gene both in vitro and in vivo. We demonstrate that the c-myb protein can transactivate the transcription of mim-1 in a transient transfection assay.

In the chick embryo, we confirm that mim-1 is specifically expressed during granulopoiesis and we show that the expression of c-myb and mim-1 are perfectly correlated in the granulocytic spleen and pancreas. However we suggest that mim-1 is efficiently transcribed in the absence of c-myb in the yolk sac and in the promyelocytes at the onset of the colonization of the bursa of Fabricius. On the other hand c-myb transcripts detected in the early hemopoietic progenitor cells, in lymphoid cells and in proliferative epithelia are never associated with mim-1 transcription.

We conclude that the granylocyte-specific mim-1 gene is regulated by c-myZ>-dependent and c-myh-independent mechanisms depending upon the environment in which granulocytic precursor cells differentiate.
The involvement of adherens junction components in myofibrillogenesis in cultured cardiac myocytes
Authors: Goncharova Ekaterina J. , Kam Zvi , Geiger Benjamin
Published: April 2021

The distribution of adherens junction (AJ) components was investigated in cultured heart myocytes. These cells, derived from either newborn rats or chick embryos, develop elaborate arrays of myofibrils which become extensive and laterally aligned following several days in culture. The Z-disks in these cells, visualized by immunolabeling with antibodies to muscle-specific a- actinin, exhibit a characteristic periodicity of about 2 fan and are in register with those of neighboring myofibrils throughout the sarcoplasm. Vinculin, in these cells, associates with intercellular AJ and cell-matrix adhesions. In addition, this protein is detected in periodic bands located along the lateral cell membranes corresponding to “costamers” previously described by Pardo, J.V., Siliciano, J.D. and Craig, S.W. (Proc. Natn. Acad. Sci. USA, 80,1008). Similarly, N-cadherin, which is predominantly associated with intercellular junctions, is also detected in periodic striations located mainly on the dorsal and lateral cell surfaces. Using computer-aided three-dimensional microscopy confirmed that these vinculin- and N-cadherin-containing structures are located in extrajunctional sites, appar-ently associated with Z-disks of peripheral myofibrils. Based on these findings an alternative pathway is proposed for the assembly of vinculin and N-cadherin, which is not triggered by adhesive interactions with extracellular surfaces but rather by interactions at the membrane-cytoplasm interphase with the periphery of the pre-assembled myofibrills. Moreover, we present evidence that antibodies to N-cadherin, which are capable of blocking AJ formation in culture, have an inhibitory effect also on the development and alignment of myofibrils. We discuss the functional significance of the “costameric” organization of vinculin and N-cadherin and consider its involvement both in the lateral alignment of neighboring muscle cells and in the stabilization of developing myofibrils.
Mutually exclusive expression of the Strongylocentrotus purpuratus Spc1 gene and its Lytechinus pictus homologue in cells of hybrid embryos
Authors: Nisson Paul E. , Gaudette Michelle F. , Brandhorst Bruce P. , Crain William R.
Published: April 2021

The expression of the Specl gene of Strongylocentrotus purpuratus and its Lytechinus pictus homologue LpSl was analyzed in reciprocal hybrid embryos of these two species of sea urchin. While the time course of accumulation of Specl mRNA was nearly normal in hybrid embryo populations, the accumulation of LpSl mRNA was not. This was particularly evident in plutei, where the level of LpSl mRNA was less than 5% that in normal L. pictus plutei. In situ hybridization analysis of serial sections indicated that LpSl mRNA was detectable in only about 2% of hybrid plutei in either cross, whereas Specl mRNA was present in nearly all hybrid plutei; expression of either homologue was appropriately restricted to the aboral ectoderm. In crosses of L. pictus eggs with S. purpuratus sperm (LpSp), about 1% of hybrid plutei expressed LpSl RNA in most or all aboral ectoderm cells at normal levels, and did not express Specl RNA; in another 1% of the LpSp hybrid plutei the Specl and LpSl transcripts were present at normal levels in complementary, non-overlapping patches of contiguous aboral ectoderm cells. In the reciprocal SpLp cross, each hybrid pluteus expressed either only the LpSl gene (about 2%) or only the Specl gene through-out the aboral ectoderm. In SpLp hybrid gastrulae the level of LpSl mRNA was less restricted; about 2% of the embryos contained only LpSl RNA, and about half expressed only Specl transcripts, but in the remaining embryos Specl and LpSl transcripts were coexpressed in the same aboral ectoderm cells. Thus, the predominant expression of the Specl gene in hybrid embryos results from the developmentally progressive restriction of transcription of the LpSl gene to aboral ectoderm cells not expressing the Specl gene in a few hybrid embryos, while most hybrid plutei express the Specl gene exclusively.
Erythroid colony formation by fetal rat liver and spleen cells in vitro: inhibition by a low relative molecular mass component of fetal spleen
Authors: Nagel M.-D. , Nagel J.
Published: April 2021

L1ver and spleen hematopoietic cell suspensions from 20-day-old-fetal rats were fractionated on Percoll gradients. A granulocyte-rich splenic fraction inhibited CFUe production by cultures of a CFUe-enriched liver fraction, and by cultures of unfractionated liver and spleen hematopoietic cells. Conditioned medium from the spleen cell fraction contained an inhibitor of relative molecular mass, Mr, 25–35 × 103. The sensitivity of spleen cells to the inhibitor varied with the age of the fetus from which they were derived (20-day-old <18- and 19-day-old). No such age-dependence was found for liver cells. The inhibitor affects cycling CFUe, blocks the lethal effect of AraC, does not appear to be lineage-specific and its influence can be reversed by washing.
Two FGF receptor genes are differentially expressed in epithelial and mesenchymal tissues during limb formation and organogenesis in the mouse
Authors: Peters K. G. , Werner S. , Chen G. , Williams L. T.
Published: April 2021

Fibroblast growth factors (FGFs) can influence the growth and differentiation of cultured cells derived from neuroectoderm, ectoderm or mesenchyme. The FGFs interact with a family of at least four closely related receptor tyrosine kinases that are products of individual genes. To investigate the role of FGFs in the growth and differentiation of embryonic tissues and to determine whether the individual FGF receptor genes might have specific functions, we compared the localization of mRNA for two FGF receptor genes, FGFR1 (the fig gene product) and FGFR2 (the bek gene product), during limb formation and organogenesis in mouse embryos (E9.5-E16.5). Although the two genes were coexpressed in some tissues, the differential expression of FGFR1 and FGFR2 in most embryonic tissues was striking. FGFR1 was expressed diffusely in mesenchyme of limb buds, somites and organ rudiments. In contrast, FGFR2 was expressed predominantly in the epithelial cells of embryonic skin and of developing organs. The differential expression of FGFR1 and FGFR2 in mesenchyme and epithelium respectively, suggests the receptor genes are independently regulated and that they mediate different functions of FGFs during development.
Combined action of c-Kit and erythropoietin on erythroid progenitor cells
Authors: Ohneda Osamu , Yanai Nobuaki , Obinata Masuo
Published: April 2021

Mutations at the murine dominant-white spotting locus (W) (c-Jtir) affect various aspects of hematopoiesis. We have made antibodies against c-Kit with the synthetic peptides deduced from the murine c-kit gene and examined the role of c-Kit in erythropoiesis. The antibody inhibited the stromal cell-dependent large colony formation of the erythroid progenitors. In the culture of erythropoietin-responsive erythroid progenitors of the anemia-inducing Friend virus-infected mouse spleen, the antibody inhibited only proliferation, but not differentiation of the progenitor cells. The inhibition was effective only at the early phase (within 6 hours after erythropoietin addition) before the cells start to proliferate induced by erythropoietin. During the early phase, erythropoietin down-regulated c-kit gene expression. These results suggest a mechanism of combined action of c-Kit with erythropoietin on the lineage-restricted erythroid progenitor cells.
Induction of dorsal and ventral mesoderm by ectopically expressed Xenopus basic fibroblast growth factor
Authors: Kimelman David , Maas Adolf
Published: April 2021

Peptide growth factors from the fibroblast growth factor (FGF) and transforming growth factor-β families are likely regulators of mesoderm formation in the early Xenopus embryo. Although basic FGF is found in the Xenopus embryo at the correct time and at sufficient concentrations to suggest that it is the FGF-type inducer, the lack of a secretory signal sequence in the basic FGF peptide has raised questions as to its role in the inductive process. We show here that Xenopus basic FGF can ectopically induce mesoderm when translated from injected synthetic RNA within the cells of a Xenopus embryo. Basic FGF produced in this manner is able to induce the formation of both dorsal and ventral mesoderm with the type of mesoderm formed dependent on the inherent dorsal-ventral polarity of the animal hemisphere. Surprisingly, although Xenopus basic FGF produced from the injected mRNA has a potent mesodermalizing effect on animal hemisphere cells, virtually no phenotypic effect is observed with intact embryos. These results suggest that the role of Xenopus basic FGF is to specify the size of the marginal zone, and synergistically with a dorsally localized prepatterning signal, to initially establish the dorsal-ventral axis of the mesoderm.
Expression of XMyoD protein in early Xenopus laevis embryos
Authors: Hopwood N. D. , Pluck A. , Gurdon J. B. , Dilworth S. M.
Published: April 2021

A monoclonal antibody specific for Xenopus MyoD (XMyoD) has been characterized and used to describe the pattern of expression of this myogenic factor in early frog development. The antibody recognizes an epitope close to the N terminus of the products of both XMyoD genes, but does not bind XMyfS or XMRF4, the other two myogenic factors that have been described in Xenopus. It reacts in embryo extracts only with XMyoD, which is extensively phosphorylated in the embryo. The distribution of XMyoD protein, seen in sections and whole-mounts, and by immunoblotting, closely follows that of XMyoD mRNA. XMyoD protein accumulates in nuclei of the future somitic mesoderm from the middle of gastrulation. In neurulae and tailbud embryos it is expressed specifically in the myotomal cells of the somites. XMyoD is in the nucleus of apparently every cell in the myotomes. It accumulates first in the anterior somitic mesoderm, and its concentration then declines in anterior somites from the tailbud stage onwards.
Distribution analysis of transferred donor cells in avian blastodermal chimeras
Authors: Watanabe M. , Kinutani M. , Naito M. , Ochi O. , Takashima Y.
Published: April 2021

Blastodermal chimeras were constructed by transferring quail cells to chick blastoderm. Contribution of donor cells to host were histologically analyzed utilizing an in situ cell marker. Of the embryos produced by Injection of stage Xl-XIII quail cells into stage XI-2 chick blastoderm, more than 50 percent were definite chimeras. The restriction on the spatial arrangement of donor cells was induced by varying the stage of host. Ectodermal chimerism was limited to the head region and no mesodermal chimerism was shown when the quail cells were injected into stage XI-XIII blastoderm. Mesodermal and ectodermal chimerisms were limited to the trunk, not to the head region, when the quail cells were injected into the stage XIV-2 blastoderm. In these chimeras, however, some of the injected quail cells formed ectopic epidermal cysts. Consequently, the stage XIV-2 blastoderm may become intolerant of the injected cells. Our results suggest that it is possible to obtain chimeras that have chimerism limited to a particular germ layer and region by varying the stage of donor cell injection. Injected quail cells contributed to endodermal tissues and primordial germ cells regardless of the injection site. The quail-chick blastodermal chimeras could be useful in the production of a transgenic chicken and in the investigation of immunological tolerance.
Evidence for a highly selective RNA transport system and its role in establishing the dorsoventral axis of theDrosophilaegg
Authors: Cheung Hung-Kam , Serano Thomas L. , Cohen Robert S.
Published: April 2021

The specification of cell fates along the dorsoventral axis of the Drosophila embryo is dependent on the asymmetric distribution of proteins within the egg and within the egg’s outer membranes. Such asymmetries arise during oogenesis and are dependent on multiple cell-cell interactions between the developing oocyte and its neighboring somatic follicle cells. The earliest known such interaction involves the generation of a signal in the oocyte and its reception in the follicle cells lying on the dorsal surface of the oocyte at ∼ stage 10 of oogenesis. Several independent lines of investigation indicate that the fs(1)K10 (K10) gene negatively regulates the synthesis of the signal in the oocyte nucleus. Here we present data that indicate that the accumulation of K10 protein in the oocyte nucleus is a multistep process involving: (1) the synthesis of K10 RNA in nurse cells, (2) the rapid transport of K10 RNA from nurse cells into the oocyte, (3) the localization of K10 RNA to the anterior margin of the oocyte, and (4) K10 protein synthesis and localization. K10 RNA is transported into the oocyte continuously beginning at ∼ stage 2. This indicates a high degree of selectivity in transport, since most RNAs synthesized in stage 2 and older nurse cells are stored there until stage 11, when nurse cells donate their entire cytoplasm to the oocyte. The sequences responsible for the early (pre-stage 11) and selective transport of K10 RNA into the oocyte map to the 3′ transcribed non-translated region of the gene. None of the other identified genes involved in dorsoventral axis formation are required for K10 RNA transport. Two such other genes, cappuccino (capu) and spire (spir) are, however, required for the subsequent localization of K10 RNA to the oocyte’s anterior end.
Inducible cell ablation inDrosophilaby cold-sensitive ricin A chain
Authors: Moffat Kevin G. , Gould Jane H. , Smith Hazel K. , O’Kane Cahir J.
Published: April 2021

We have developed a system for temperature-inducible killing of specific cells in the fruitfly Drosophila melanogaster. The system overcomes many of the limitations of existing cell ablation methods and is in principle applicable to any non-homeothermic eukaryote. Temperature-sensitive and cold-sensitive mutations in the ricin toxin A chain (RTA) of castor bean were generated in yeast. One cold-sensitive mutation, RAcs2, produced temperature-dependent ablation of eye cells in Drosophila when expressed under control of the eye-specific sev enhancer. At 29°C, cell death was observed within 7 hours in the developing eye and no obvious toxic effects were observed elsewhere; at 18°C, extremely low toxicity was observed. DNA sequencing of RAcs2 revealed a single amino acid substitution in the RTA active site cleft.
A quantitative and interspecific test for biological activity of anti-Müllerian hormone: the fetal ovary aromatase assay
Authors: Clemente Nathalie Di , Ghaffari Sarah , Pepinsky R. B. , Pieau C. , Josso Nathalie , Cate R. L. , Vigier B.
Published: April 2021

Anti-Müllerian hormone (AMH), also known as Müllerian-inhibiting substance or factor, has previously been shown to sex-reverse the steroidogenic pattern of fetal mammalian ovaries through repression of aromatase biosynthesis. Study of the ontogeny of the response of cyclic AMP-stimulated aromatase activity of rat fetal ovaries to AMH has allowed us to develop a quantitative biossay for the hormone. Linear responses as a function of the logarithm of AMH concentration were observed over ranges of 0.2 -7.5 μg/ml for the bovine protein and 0.15 -2 μg/ml for the human protein, with a maximal decrease in aromatase activity of 90% for both proteins. Under the same in vitro conditions, AMH treatment did not affect cyclic AMP-stimulated fetal rat testicular aromatase activity. Partially purified chick AMH also decreased rat ovarian aromatase activity, allowing us to use this test to study AMH ontogeny in chick gonads. Analysis of the species specificity of AMH repression of ovarian aromatase activity indicated that turtle and rat fetal ovaries responded to AMH of other vertebrate classes, whereas aromatase activity of chick embryo ovaries could be repressed only by the homospecific hormone.
Fgf-4expression during gastrulation, myogenesis, limb and tooth development in the mouse
Authors: Niswander Lee , Martin Gail R.
Published: April 2021

Fgf-4, initially isolated as a transforming gene from human tumors, is a member of the Fibroblast Growth Factor (FGF) family. It has previously been shown by northern blot hybridization analysis to be expressed in teratocarcinoma and embryonic stem cells, suggesting that it plays a role in embryonic development. We have carried out an RNA in situ hybridization analysis of Fgf-4 expression in the developing mouse embryo, from fertilization through the 14th day of gestation (E14.5). Our results show that Fgf-4 RNA is first detected at the late blastocyst stage in cells that give rise to all of the embryonic lineages (inner cell mass cells). During the early stages of gastrulation, expression becomes restricted to the primitive streak where mesoderm and definitive endoderm are formed. Expression continues in the distal (rostral) two-thirds of the streak through approx. E10, and then is detected in the tail bud, which replaces the streak as the primary source of mesoderm. Additional sites of expression are found after the three primary germ layers are established and organogenesis begins. Fgf-4 RNA is detected transiently in the branchial arch units, the somitic myotome, the apical ectodermal ridge of the developing limb bud and the tooth bud, suggesting that the gene has multiple roles during embryogenesis. These results are compared with the expression patterns of other FGF genes. Taken together, the data suggest that individual members of the gene family are expressed sequentially in developmental pathways such as mesoderm formation and myogenesis, and play a role in specific epithelial-mesenchymal interactions.
Altered mitotic domains reveal fate map changes in Drosophila embryos mutant for zygotic dorsoventral patterning genes
Authors: Arora Kavita , Nüsslein-Volhard Christiane
Published: April 2021

The spatial and temporal pattern of mitoses during the fourteenth nuclear cycle in a Drosophila embryo reflects differences in cell identities. We have analysed the domains of mitotic division in zygotic mutants that exhibit defects in larval cuticular pattern along the dorsoventral axis. This is a powerful means of fate mapping mutant embryos, as the altered position of mitotic domains in the dorsoventral pattern mutants correlate with their late cuticular phenotypes.

In the mutants twist and snail, which fail to differentiate the ventrally derived mesoderm, mitoses specific to the mesoderm are absent. The lateral mesectodermal domain shows a partial ventral shift in twist mutants but a proportion of ventral cells do not behave characteristically, suggesting that twist has a positive role in the establishment of the mesoderm. In contrast, snail is required to repress mesectodermal fates in cells of the presumptive mesoderm. In the absence of both genes, the mesodermal and the mesectodermal anlage are deleted.

Mutations at five loci delete specific pattern elements in the dorsal half of the embryo and cause partial ventralization. Mutations in the genes zerkniillt and shrew affect cell division only in the dorsalmost cells corresponding to the amnioserosa, while the genes tolloid, screw and decapentaplegic (dpp) affect divisions in both the prospective amnioserosa and the dorsal epidermis. We demonstrate that in each of these mutants dorsally placed mitotic domains are absent and this effect is correlated with an expansion and dorsal shift in the position of more ventral domains. The loss of activity in each of the five genes results in qualitatively similar alterations in the mitotic pattern; mutations with stronger ventralizing phenotypes affect increasingly greater subsets of the dorsal cells. Double mutant analysis indicates that these genes act in a concerted manner to specify dorsal fates. The correlation between phenotypic strength and the progressive loss of dorsal pattern elements in the ventralized mutants, suggests that one of these gene products, perhaps dpp, may provide positional information in a graded manner.
An early marker of axial pattern in the chick embryo and its respecification by retinoic acid
Authors: Sundin Olof , Eichele Gregor
Published: April 2021

Chick Ghox 2.9 protein, a homeodomain-contaming polypeptide, is first detected in the mid-gastrula stage embryo and its levels increase rapidly in the late gastrula. At this time, the initially narrow band of expression along the primitive streak expands laterally to form a shield-like domain that encompasses almost the entire posterior region of the embryo and extends anteriorly as far as Hensen’s node. We have found that this expression domain co-localizes with a morphological feature that consists of a stratum of refractile, thickened mesoderm. Antibody-staining indicates that Ghox 2.9 protein is present in all cells of this mesodermal region. In contrast, expression within the ectoderm overlying the region of refractile mesoderm varies considerably. The highest levels of expression are found in ectoderm near the streak and surrounding Hensen’s node, regions that recent fate mapping studies suggest are primarily destined to give rise to neurectoderm.

At the definitive streak stage (Hamburger and Hamilton stage 4) the chick embryo is especially sensitive to the induction of axial malformations by retinoic acid. Four hours after the treatment of definitive streak embryos with a pulse of retinoic acid the expression of Ghox 2.9 protein is greatly elevated. This ectopic expression occurs in tissues anterior to Hensen’s node, including floor plate, notochord, presumptive neural plate and lateral plate mesoderm, but does not occur in the anteriormost region of the embryo. The ectopic induction of Ghox 2.9 is strongest in ectoderm, and weaker in the underlying mesoderm. Endoderm throughout the embryo is unresponsive. At stage 11, Ghox 2.9 is normally expressed at high levels within rhombomere 4 of the developing hindbrain. In retinoic-acid-treated embryos which have developed to this stage, typical rhombomere boundaries are largely absent. Nevertheless, Ghox 2.9 is still expressed as a discrete band, but one that is widened and displaced to a more anterior position.
N-cadherin expression in developing, adult and denervated chicken neuromuscular system: accumulations at both the neuromuscular junction and the node of Ranvier
Authors: Cifuentes-Diaz Carmen , Nicolet Marc , Goudou Danièle , Rieger François , Mege René Marc
Published: April 2021

N-cadherin, a member of the Ca2+-dependent cell adhesion molecule family plays essential roles in morphogenesis and histogenesis. N-cadherin has been shown in vitro to promote myoblast fusion and neurite outgrowth. We report here the cellular localization of N-cadherin during development and regeneration of the chick neuromuscular system. N-cadherin was uniformly expressed along the surface of myoblasts and myotubes of E6 limb muscles. Later, as synaptogenesis and secondary myogenesis proceeded, N-cadherin expression was down-regulated and restricted to some large-diameter fibres, then to the areas of contact between few myofibres and subsequently disappeared by embryonic day 17, suggesting that this cadherin may be implicated predominantly in fusion of primary myoblasts and, at lower degree, of secondary myoblasts. The presence of N-cadherin in muscle during the period of nerve trunk ingrowth and its down-regulation after synaptogenesis suggests that this molecule might be implicated in both processes. N-cadherin became accumulated at the neuromuscular junction only a few days after the first synaptic contacts were established and remained at the adult neuromuscular junction, suggesting a role of this molecule in the stabilization of the mature neuromuscular junction. In sciatic nerve, the level of N-cadherin expression remained unchanged from hatching to adult life. N-cadherin was widely distributed on the surface of myelinated fibres and on myelinating Schwann cells: in addition, it was concentrated at the node of Ranvier. At the ultrastructural level, the molecule was detected inside, at the surface and in the basal lamina of Schwann cells and also associated with endoneurial collagen. These observations suggest a role of N-cadherin in the structuring and stabilization of the myelin sheaths. After nerve injury, N-cadherin continued to be expressed by proliferating Schwann cells in the distal stump providing a substratum for regenerating axons. N-cadherin reappeared at the surface of denervated muscle fibres without disappearing from the former synaptic sites. It was detected not only in the sarcoplasm and on sarcolemma of denervated muscle fibres, but also in the basal lamina and in the extracellular matrix. The reexpression of N-cadherin at the surface of denervated muscle fibres suggests a role for this molecule in muscle reinnervation. The presence of N-cadherin in basal lamina and its association with collagen fibres raise questions about the release of N-cadherin in the extracellular space and the existence of a putative heterophilic ligand for N-cadherin.
Developmental regulation of the orphan receptor COUP-TF II gene in spinal motor neurons
Authors: Lutz Beat , Kuratani Shigeru , Cooney Austin J. , Wawersik Stefan , Tsai Sophia Y. , Eichele Gregor , Tsai Ming-Jer
Published: April 2021

Members of the steroid/thyroid hormone receptor super- family are involved in the control of cell identity and of pattern formation during embryonic development. Chicken ovalbumin upstream promoter-transcription factors (COUP-TFs) can act as regulators of various steroid/thyroid hormone receptor pathways. To begin to study the role of COUP-TFs during embryogenesis, we cloned a chicken COUP-TF (cCOUP-TF II) which is highly homologous to human COUP-TF II. Northern analysis revealed high levels of cCOUP-TF II transcripts during organogenesis. Nuclear extracts from whole embryos and from embryonic spinal cords were used in electrophoretic mobility shift assays. These assays showed that COUP-TF protein is present in these tissues and is capable of binding to a COUP element (a direct repeat of AGGTCA with one base pair spacing). Analysis of cCOUP-TF expression by in situ hybridization revealed high levels of cCOUP-TF II mRNA in the developing spinal motor neurons. Since the ventral properties of the spinal cord, including the development of motor neurons, is in part established by inductive signals from the notochord, we transplanted an additional notochord next to the dorsal region of the neural tube in order to induce ectopic motor neurons. We observed that an ectopic notochord induced cCOUP-TF II gene expression in the dorsal spinal cord in a region coextensive with ectopic domains of SC1 and Islet-1, two pre-viously identified motor neuron markers. Collectively, our studies raise the possibility that cCOUP-TF II is involved in motor neuron development.
Function of Drosophila ovo+ in germ-line sex determination depends on X-chromosome number
Authors: Oliver Brian , Singer Jason , Laget Valérie , Pennetta Giuseppa , Pauli Daniel
Published: April 2021

Germ-line sex determination in Drosophila melanogaster requires an assessment of the number of X chromosomes as measured against autosomal standards (XX =; female, X =; male) and signaling from the soma. Both of these sex determination cues are required for female-specific Sex-lethal+ function in germ cells. The ovo+ locus encodes zinc finger protein(s) required for female-specific splicing of Sex-lethal+ pre-mRNA, making ovo+ a candidate function acting between the two principal cues and Sex-lethal+. We have made ovo reporter genes and find that they show high activity in the germ line of females and low activity in the germ line of males. XY flies transformed into somatic females do not show high levels of reporter activity, while XX flies transformed into somatic males do. This shows that high level ovo+ expression depends on the number of X chromosomes, not the somatic sexual signals. The requirement for ovo+ function is restricted to XX flies. Mutations in ovo have no effect on XY males, X0 males or XY females, but have pronounced effects on germ cell viability in XX females, XX females with sex transformed germ lines, and XX males indicating that ovo+ gene products are required for events occurring only in flies with two X chromosomes.
GP330 is specifically expressed in outer cells during epithelial differentiation in the preimplantation mouse embryo
Authors: Gueth-Hallonet Catherine , Santa-Maria Angelica , Verroust Pierre , Maro Bernard
Published: April 2021

During preimplantation development of the mouse embryo, a layer of outer cells differentiates into a perfect epithelium, the trophectoderm. The divergence between the trophectoderm and the inner cell mass takes place from the 8-cell stage to the 64-cell stage and precedes their commitment at the blastocyst stage. In this work, we have investigated the expression of gp330, a 330×103Mr glycoprotein found in clathrin-coated areas of the plasma membrane of some epithelial cells characterized by a high level of endocytic activity. Our results show that gp330 is first synthesized in 16-cell stage embryos and that its appearance is restricted to outer cells until the blastocyst stage. Furthermore, its expression is repressed in inner cells at a post-transcriptional level, probably through the development of extensive cell-cell contacts.
Apoptosis during macrophage-dependent ocular tissue remodelling
Authors: Lang Richard , Lustig Marc , Francois Fritz , Sellinger Meredith , Plesken Heide
Published: April 2021

We have characterized the nature and pattern of cell death during regression of the pupillary membrane, a developmentally transient capillary network found in the anterior chamber of the eye. This analysis has revealed that the cellular components of the pupillary membrane include vascular endothelial cells in an intricate network of fine capillaries as well as attendant macrophages. The capillaries are situated on the anterior surface of the lens and held in relative position by a cobweb-like meshwork of extracellular matrix fibres that regress along with the cellular components of this structure. Cell death during regression of the pupillary membrane is characteristic of apoptosis. Specifically, apoptotic bodies containing condensed chromatin can be observed in vascular endothelial cells and genomic DNA isolated from the pupillary membrane shows the nucleosomal fragmentation pattern typical of apoptotic cells. Using a method for labelling fragmented DNA in tissue preparations (TUNEL), we have assessed the overall pattern of apoptotic cell death during pupillary membrane regression. We find that apoptosis occurs either in single cells in healthy vessels or synchronously along the entire length of a capillary segment. Both morphological and TUNEL analysis indicate that capillary regression occurs from junction to junction one segment at a time. We propose a model to explain the pattern of capillary regression observed and conclude from these and previous experiments (Lang and Bishop (1993)Cell 74, 453-462), that during regression of the pupillary membrane, the macrophage elicits target cell death by inducing apoptosis.
Strain-specific differences in mouse oocytes and their contributions to epigenetic inheritance
Authors: Latham Keith E.
Published: April 2021

Previous experiments revealed a strain-dependent effect of egg cytoplasm on the developmental potential of androgenetic (two paternal genomes) mouse embryos. Eggs obtained from C57BL/6 mice supported androgenone development to the blastocyst stage at a much higher frequency than eggs from DBA/2 mice. Transient exposure of paternal pronuclei to DBA/2 egg cytoplasm also compromised development, indicating that the DBA/2 egg cytoplasm negatively affected the ability of paternal pronuclei to support blastocyst formation. An essential first step toward understanding the molecular mechanism by which egg modifier factors influence gene expression is to determine the number of loci that are responsible for the strain difference. To do this, (B6D2)F1 hybrid females were backcrossed to DBA/2 males and the eggs from individual female progeny assayed for their ability to support androgenetic development. Approximately one fourth of the backcross females produced eggs that failed to support androgenone development, indicating that two independently segregating genetic loci are most likely responsible for the difference between DBA/2 and C57BL/6 egg phenotypes. Comparison of DBA/2 and C57BL/6 oocytes by two-dimensional protein gel electrophoresis revealed at least 17 proteins that exhibited significant, reproducible, quantitative differences in rates of synthesis. All of these proteins were synthesized in (B6D2)F1 oocytes. These data, combined with the previous observation that the C57BL/6 egg phenotype is dominant, are consistent with a model in which a C57BL/6 allele at either locus provides a protective function, either by antagonizing the actions of the DBA/2 alleles or by providing, through partial or complete redundancy, a function not provided by the DBA/2 alleles.
TGF-β1 influences the relative development of the exocrine and endocrine pancreas in vitro
Authors: Sanvito F. , Herrera P.-L. , Huarte J. , Nichols A. , Montesano R. , Orci L. , Vassalli J.-D.
Published: April 2021

Pancreatic rudiments from E12.5 mouse embryos undergo extensive development and differentiation when cultured in three-dimensional gels of extracellular matrix proteins for up to 12 days. Whereas collagen gels promote the formation of numerous exocrine acini and relatively small clusters of endocrine cells, in basement membrane (EHS) matrices the development of endocrine cells is dramatically favoured over that of acinar tissue. Buds embedded in a collagen gel contiguous to an EHS gel also fail to develop acini, suggesting the involvement of diffusible factor(s). Addition of cytokines to cultures of pancreatic buds in collagen gels modifies the relative proportions of the epithe-lial components of the gland. In the presence of EGF the proportion of the tissue occupied by ducts overrides that of acinar structures, whereas the endocrine portion of the tissue is not significantly modified. TGF-β1 partially mimicks the effect of EHS matrix in inhibiting the devel-opment of acinar tissue without decreasing the amount of ducts and mesenchyme; TGF-β1 also promotes the devel-opment of endocrine cells, in particular of insulin-contain-ing β cells and of cells expressing genes of the PP-fold family. These results show that cytokines can modulate the development of the pancreas and suggest a role for TGF-β1 in regulating the balance between the acinar and endocrine portions of the gland in vivo. More generally, they are compatible with the notion that, during organo-genesis, cytokines act as paracrine factors responsible for the development and maintenance of appropriate proportions of different tissue constituents.