bluuebunny/crossref_metadata_2025 huggingface dataset:
Changes in mRNA length accompany translational regulation of the somatic and testis-specific cytochrome c genes during spermatogenesis in the mouse
Authors: Hake Laura E. , Alcivar Acacia A. , Hecht Norman B.
Published: April 2021

The mouse testis contains two isotypes of cytochrome c, which differ in 14 of 104 amino acids: cytochrome cs is present in all somatic tissues and cytochrome cT is testis specific. The regulation of cytochrome cS and cytochrome cT gene expression during spermatogenesis was examined by Northern blot analysis using specific cDNA probes. Total RNA was isolated from adult tissues, enriched germinal cell populations and polysomal gradients of total testis and isolated germinal cells. Three cytochrome CS mRNAs were detected averaging 1.3 kb, 1.1 kb and 0.7 kb in all tissues examined; an additional 1.7 kb mRNA was observed in testis. Isolated germinal cells through prepuberal pachytene spermatocytes contained only the three smaller mRNAs; the 1.7 kb mRNA was enriched in round spermatids. All three smaller cytochrome cs mRNAs were present on polysomes; the 1.7 kb mRNA was non-polysomal. Cytochrome cT mRNA of 0.6 – 0.9 kb was detected in testis; mRNA levels were low in early spermatogonia and peaked in prepuberal pachytene spermatocytes. In adult pachytene spermatocytes, a subset of the cytochrome cT mRNAs, 0.7 – 0.9kb, was present on polysomes; a shortened size class, 0.6 – 0.75 kb, was non-polysomal. A distinct, primarily non-polysomal, cytochrome cT 0.7 kb mRNA was present in round spermatids. These results indicate that (1) both cytochrome c.s and cytochrome cT mRNAs are present in early meiotic cells, (2) a 1.7 kb cytochrome cS mRNA is post-meiotically expressed and non-polysomal and (3) cytochrome cS and cytochrome cT mRNAs are each developmentally and translationally regulated during spermatogenesis.
Storage protein gene expression is localised to regions lacking mitotic activity in developing pea embryos. An analysis of seed development in Pisum sativum XIV
Authors: Hauxwell Angela J. , Corke Fiona M. K. , Hedley Cliff L. , Wang Trevor L.
Published: April 2021

Storage protein gene expression has been studied in relation to mitotic activity to ascertain whether these processes are linked during embryo development in pea. Sections from immature pea embryos were probed by in situ hybridisation to show the pattern of vicilin storage protein gene expression. In addition, the location of mitotic cells was identified using fluorescence microscopy. Vicilin mRNA was first localized in the parenchyma cells of the upper adaxial region of the cotyledons. As the embryos increased in fresh weight, gene expression spread from this region, in a wave-like manner, down and across the cotyledons. The gene was only expressed in those regions of the embryo that lacked mitotic activity.
Cell shape changes during gastrulation in Drosophila
Authors: Leptin Maria , Grunewald Barbara
Published: April 2021

The first morphogenetic movement during Drosophila development is the invagination of the mesoderm, an event that folds a one-layered epithelium into a multilayered structure. In this paper, we describe the shape changes and behaviour of the cells participating in this process and show how mutations that change cell fate affect this behaviour.

We divide the formation of the mesodermal germ layer into two phases. During the first phase, the ventral epithelium folds into a tube by a series of concerted cell shape changes (ventral furrow formation). Based on the behaviour of cells in this phase, we conclude that the prospective mesoderm is not a homogeneous cell population, but consists of two subpopulations. Each subpopulation goes through a distinctive sequence of specific cell shape changes which together mediate the invagination of the ventral furrow. In the second phase, the invaginated tube of mesoderm loses its epithelial character, the mesoderm cells disperse, divide and then spread out along the ectoderm to form a single cell layer.

To test how ventral furrow formation depends on cell fates in the mesoderm and in neighbouring cells we alter these fates genetically using maternal and zygotic mutations. These experiments show that some of the aspects of cell behaviour specific for ventral furrow cells are part of an autonomous differentiation programme. The force driving the invagination is generated within the region of the ventral furrow, with the lateral and dorsal cell populations contributing little or none of the force. Two known zygotic genes that are required for the formation of the mesoderm, twist and snail, are expressed in ventral furrow cells, and the correct execution of cell shape changes in the mesoderm depends on both.

Finally, we show that the region where the ventral furrow forms is determined by the expression of mesoderm-specific genes, and not by mechanical or other epigenetic properties of the egg.
Microenvironment created by stromal cells is essential for a rapid expansion of erythroid cells in mouse fetal liver
Authors: Ohneda Osamu , Yanai Nobuaki , Obinata Masuo
Published: April 2021

Mouse stromal cell lines (FLS lines), established from the livers of 13-day gestation mouse fetus, supported the proliferation and differentiation of the erythroid progenitor cells from mouse fetal livers and bone marrow in a semisolid medium in the presence of erythropoietin. A large erythroid colony of over 1000 benzidine-positive erythroid cells was developed from a single erythroid progenitor cell on the FLS cell layer after 4 days of culture. When in close contact with the layer, the erythroid progenitor cells divided rapidly with an average generation time of 9.6 h and mature erythroid cells, including enucleated erythrocytes, were produced. The present studies demonstrate that the microenvironment created by the stromal cells can support the rapid expansion of erythropoietic cell population in the fetal liver of mice.
DNA sequence-specific binding activity of the heat-shock transcription factor is heat-inducible before the midblastula transition of early Xenopus development
Authors: Ovsenek Nick , Heikkila John J.
Published: April 2021

We have examined the activity of the Xenopus heat-shock transcription factor (HSF) in extracts from stressed and unstressed embryos at various stages of development using DNA mobility shift analysis. A specific interaction between HSF and a synthetic oligonucleotide corresponding to the proximal heat-shock element (HSE) of the Xenopus HSP70B gene was greatly enhanced in heat-shocked embryos compared to controls. HSF binding was inducible at all developmental stages examined including pre-midblastula transition (MBT) stages which are incapable of expressing HSP genes. In time-course experiments with both cleavage and neurula stage embryos, the activation of HSF binding was rapid and transient. Removal of cleavage and neurula stage embryos from heat stress resulted in a rapid loss of binding activity. The molecular mass of HSF, as determined by comparative gel electrophoresis of photoaffinity-labeled factor was 88X103 in both heat-shocked cleavage and neurula stage embryos. These experiments suggest that maternally derived HSF is stored in pre-MBT embryos in a heat-activatable form and may function in the regulation of heat-shock genes immediately after the MBT.
Phenotypic rescue of mutant brown melanocytes by a retrovirus carrying a wild-type tyrosinase-related protein gene
Authors: Bennett Dorothy C. , Huszar Dennis , Laipis Philip J. , Jaenisch Rudolf , Jackson Ian J.
Published: April 2021

A mouse cDNA for the developmentally controlled, melanocyte-specific protein, tyrosinase-related protein 1 (TRP-1), was previously cloned and reported to show genetic linkage with the coat-colour locus brown (b) on mouse chromosome 4. The cDNA has been inserted into a retroviral vector derived from Moloney murine leukaemia virus, under the control of the human histone H4 promoter. This vector was used to infect melanocytes of the immortal line melan-b, which are homozygous for the b mutation and which display light brown pigmentation in culture. Infected cultures containing between 0.2 and 2 copies of provirus per cell displayed an altered phenotype: 20-50 % of cells now had the black to dark brown colour characteristic of cultured wild-type (Black, B/B) mouse melanocytes. Thus the TRP-1 gene complements the brown mutation. We conclude that TRP-1 is the product of the wild-type-b-locus.
Embryonic stem cells alone are able to support fetal development in the mouse
Authors: Nagy András , Gócza Elen , Diaz Elizabeth Merentes , Prideaux Valerie R. , Iványi Eszter , Markula Merja , Rossant Janet
Published: April 2021

The developmental potential of embryonic stem (ES) cells versus 3.5 day inner cell mass (ICM) was compared after aggregation with normal diploid embry os and with developmentally compromised tetraploid embryos. ES cells were capable of colonizing somatic tissues in diploid aggregation chimeras but less efficiently than ICMs of the same genotype. When ICM"-"tetraploid and IS-tetraploid chimeras were made, the newborns were almost all completely ICM-or ES-derived. as judged by GPI isozyme analysis, but tetraploid cells were found in the yolk sac endoderm and trophectoderm lineage. Investigation of ES contribution in 13.5 day ES-tetraploid chimeras by DNA in situ hy bridization confirmed the complete tetraploid origin of the placenta (except the fetal blood and blood vessels) and the yolk sac endoderm. However, the yolk sac mesoderm, amnion and fetus contained only ES-derived cells.

ES-derived newborns failed to survive after birth, although they had normal birthweight and anatomically they appeared normal. This phenomenon remains unexplained at the moment.

The present results prove that ES cells are able to support complete fetal development, resulting in ES-derived newborns, and suggest a useful route for studying the development of genetically manipulated ES cells in all fetal lineages.
Differential expression of laminin A and B chains during development of embryonic mouse organs
Authors: Klein Gerd , Ekblom Marja , Fecker Lothar , Timpl Rupert , Ekblom Peter
Published: April 2021

Laminin is a large glycoprotein of basement membranes. The best described laminin from a mouse tumor contains three polypeptide chains (A, Bl and B2), but there is recent evidence that some cell types produce, laminin isoforms lacking the A chain. We have here studied the occurrence of the isoforms during mouse organogenesis. In all tissues studied, the A chain mRNA and polypeptide were more weakly expressed than those of the B chains. Laminin A chain polypeptides showed a much more restricted tissue distribution than the B chains. Laminin A chain polypeptide was mainly detected in basement membranes of epithelial cells, suggesting that this chain is important for morphogenesis of epithelial sheets. Most endothelial basement membranes and all embryonic mesenchyme matrices studied seemed to lack the A chain even though they contained B chains. Several of the cells producing laminin devoid of A chain seem to produce other polypeptides that become complexed to the B chains. With an anti-laminin antiserum, which in immunblots reacts only with A and B polypeptide chains, additional polypeptides of 160 and 190×l03Mr were co-precipitated from all tissues studied. In developing heart, a polypeptide of 300×l03Mr was co-precipitated in addition. Our data suggest that these laminin-associated polypeptides are not formed by a differential splicing of the known A chain mRNA. Northern blotting of poly(A)+ RNA showed only 10 kb A chain transcripts but no truncated forms. We conclude that several cell types in the mouse embryo produce laminin variants that lack the 400×l03Mr A chain. Since a major cell binding site of laminin contains parts of the A chain, the variants should differ in biological function from laminin containing this A chain.
Retinoic acid modulation of the early development of the inner ear is associated with the control of c-fos expression
Authors: Represa Juan , Sanchez Alberto , Miner Cristina , Lewis John , Giraldez Fernando
Published: April 2021

The effects of retinoic acid (RA) on the early development of the inner ear were studied in vitro using isolated chick embryo vesicles. Low concentrations of RA (1–50 nM) inhibited vesicular growth in stage 18 otic vesicles that were made quiescent and then reactivated by either serum or bombesin. Growth inhibition was concentration-dependent and was paralleled by a reduction in the rate of DNA synthesis as measured by [3H]thymidine incorporation. Half-inhibition occurred between 1 and 10 nM RA, and the full effect at 20 nM. Retinoic acid, in the presence of serum, induced the precocious differentiation of (1) secretory epithelium, the tegmentum vasculosum and endolymphatic sac and (2) early sensory and supporting epithelia. These structures were positioned in their corresponding normal presumptive areas. The overall direction of growth was reversed by RA and the ratio of the internal to the external vesicular surface area increased with RA concentration. The expression of the nuclear protooncogene c-fos in the developing otic vesicle was transient and stage-dependent. High levels of c-fos mRNA were positively correlated with cell proliferation. Incubation of growth-arrested otic vesicles with bombesin plus insulin at concentrations that induced cell proliferation produced a strong induction of c-fos. This mitogen-induced expression was suppressed by 25 nM RA. The results suggest (1) a role for retinoic acid in controlling the early development of the inner ear and (2) that this control is effected through the regulation of the proto-oncogene c-fos.
Retinoic acid receptors and cellular retinoid binding proteins: I. A systematic study of their differential pattern of transcription during mouse organogenesis
Authors: Dollé Pascal , Ruberte Esther , Leroy Pierre , Morriss-Kay Gillian , Chambon Pierre
Published: April 2021

We report here the gene expression patterns, as revealed by in situ hybridisation, of the retinoic acid receptors alpha, beta and gamma (RAR-α, -β and -γ), and the cellular binding proteins for retinol and retinoic acid (CRBP, CRABP) in non-neural tissues of mouse embryos during the period of organogenesis. At all stages, RAR-α transcripts were almost ubiquitous, whereas the distribution of transcripts of the other four genes was distinctive in all systems. At early stages in the formation of an organ, the expression patterns were different in the epithelium, the adjacent mesenchyme, and in mesenchyme more distant from the epithelium, suggesting a role for RA and RA receptors in epithelial–mesenchymal tissue interactions. In the developing face, limb bud and genital tubercle, where large expanses of mesenchyme are present, differential patterns of expression were established before the onset of overt tissue differentiation, suggesting some significance for pattern formation in these regions. The distribution of RAR-β transcripts in tracheobronchial, intestinal and genital tract epithelia is consistent with the possibility that RAR-β plays a role in mediating retinoid effects on the differentiated stage of these epithelia. Possible developmental roles of RARs in relation to the expression patterns of other genes are discussed. CRBP expression domains showed a high degree of overlap with RAR-β and RAR-γ, and a mutual exclusivity with CRABP expression domains. Correlation of these expression patterns with the morphogenetic effects of vitamin A deficiency and retinoid excess lead us to propose that the function of CRBP is to store and release retinol where high levels of RA are required for specific morphogenetic processes, while CRABP serves to sequester RA in regions where normal developmental functions require RA levels to be low. Where both binding protein genes are expressed in a non-overlapping pattern within a large area of mesenchyme, a gradient of free RA may be created between them by release of retinol-derived RA from CRBP-expressing cells, with binding to CRABP enhancing the steepness of the decline in concentration distant to the source.
Nucleolar distribution of proteins B23 and nucleolin in mouse preimplantation embryos as visualized by immunoelectron microscopy
Authors: Biggiogera M. , Bürki K. , Kaufmann S. H. , Shaper J. H. , Gas N. , Amalric F. , Fakan S.
Published: April 2021

The ultrastructural distribution of proteins B23 and nucleolin in the nucleolus of mouse embryos from the zygote to the early blastocyst has been analyzed by means of specific antibodies and immunocytochemistry using colloidal gold complexes as markers. In parallel, silver staining of nucleoli was carried out on ultrathin sections. Our results show that the compact prenucleolar bodies at 1- and 2-cell stage as well as the compact residual fibrillar masses observed up to the morula stage, are labelled with the two antibodies. These masses, however, are not stained with silver up to the 4-cell stage. In well-developed nucleoli, the two antibodies co-localize in the dense fibrillar component (DFC) and the granular component (GC) while fibrillar centers (FCs) are devoid of label. On the contrary, silver staining occurs in the FCs and DFC but not in the GC. Our observations suggest that there is no direct relationship between the occurrence of silver staining and the distribution of protein B23 or nucleolin. Moreover, neither the localization of the two above proteins nor silver staining are unequivocally related to the nucleolar activity.
Developmental loss of laminin from the interstitial extracellular matrix correlates with decreased laminin gene expression
Authors: Kücherer-Ehret Andrea , Pottgiesser Jutta , Kreutzberg Georg W. , Thoenen Hans , Edgar David
Published: April 2021

The expression of the polypeptide subunits of the glycoprotein laminin in developing mouse tissues was analysed by immunoblots and Northern blots, and by immunohistochemistry at the ultrastructural level. In the neonate, almost all the laminin of the sciatic nerve was freely extractable and was located mainly in the mesenchymal interstitial extracellular matrix, rather than in basement membranes. During the first two postnatal weeks, the distribution of laminin shifted to assume the adult pattern, most being located in basement membranes and insoluble under physiological conditions. Analysis of laminin subunit expression showed that both the mRNA for the laminin B chains and the corresponding polypeptides are widely expressed in nerve and other tissues, the mRNA levels decreasing during the first two postnatal weeks as the amount of laminin in the tissue increased. In contrast, the A chain mRNA and polypeptide were undetectable in nerve at any age studied, although they were present in perinatal kidney and placenta. It is proposed that the large amount of soluble laminin present in the developing interstitial extracellular matrix is a consequence of the high levels of expression of laminin mRNA, the subsequent decrease in expression resulting in the adult distribution where most laminin is insoluble within the basement membrane.
Early C. elegans embryos are transcriptionally active
Authors: Schauer Irene E. , Wood William B.
Published: April 2021

We have developed a nucleotide incorporation assay for run-on transcription in C. elegans embryonic extracts as an approach to characterizing early transcription. The incorporation is primarily polymerase II-catalyzed RNA synthesis, producing transcripts of the expected size range for mRNAs. Incorporation is insensitive to inhibitors of reinitiation, indicating that the activity represents primarily elongation of nascent chains initiated prior to extract preparation. The transcripts produced appear to be unprocessed pre-mRNAs. Hy bridization of labeled transcripts from extracts of staged embryos to a set of cloned genes suggests that the specificity of the in vitro reaction accurately reflects developmentally regulated in vivo transcription. Com parative analyses of transcription in extracts from various stages indicate that pregastrulation embryos are active transcriptionally and that the level of transcrip tion per nucleus is approximately constant throughout embryogenesis. Furthermore, most embryonically ex pressed genes are already being transcribed in pregas trulation embryos. We also demonstrate that the labeled embryonic run-on transcripts can be used as probes to screen for sequences transcribed preferentially in pregastrulation embryos. There appears to be only a small set of such sequences, which could represent a previously unsuspected class of embryonically tran scribed genes important for early embryogenesis.
Expression of IGF-I and -II mRNA in the brain and craniofacial region of the rat fetus
Authors: Lievre C. Ayer-Le , Ståhlbom P.A. , Sara V. R.
Published: April 2021

Insulin-like growth factors (IGF-I and -II) are present in the brain during development, with high levels of both being also found in the periphery particularly in the embryo. IGFs in the brain are believed to stimulate the proliferation of neuronal and glial precursors and their phenotypic differentiation. Using in situ hybridization, we have investigated the distribution of cells producing IGF-I and -II in the rat fetus during the second half of prenatal development with special emphasis on the peripheral and central nervous system. High levels of IGF-I mRNA were found in the olfactory bulb and in discrete neurons of the cranial sensory ganglia, notably in the trigeminal ganglion, as early as 13 days of gestation, in the pineal primordium of 18 day old fetuses, and in discrete groups of cells in the cochlear epithelium located laterally outside the forming spiral organ, in day 13 to 21 fetuses. High levels of IGF-II mRNA in the brain, besides the choroid plexus and the leptomeninges, were detected in hypothalamus, in the floor of the 3rd ventricle at all stages studied, in the pineal primordium at 18 days and in the pars intermedia of the pituitary or in the Rathke’s pouch epithelium from which it is derived, with progressive fading towards the end of the gestation. In the peripheral nervous system the IGF-II mRNA was only found in association with the vascular endothelia of the ganglia.

IGF-II mRNA in the nervous system was found in highly vascularized areas, meninges, blood vessels and choroid plexuses. It is thus associated with structures involved in the production of extracellular fluids and/or substrate transport and supply in the nervous tissues. A more specific role in the differentiation or fetal endocrine function should be considered for IGF-II in cells producing melatonin and melanocyte stimulating hormone (MSH) in the pineal and pituitary glands, respectively.

The presence of IGF-I mRNA in the nervous system could be associated with fiber outgrowth and synapto-genesis in the cases of olfactory bulb and developing iris. The role of IGF-I in restricted populations of cells of the cochlear epithelium and in the pineal gland is unclear and requires further investigations including a search for IGF-I receptors in possible target cells.

In the sensory ganglia, the presence of high levels of IGF-I mRNA eventually corresponds to the production, by post-translational processing, of the amino-terminal tripeptide of IGF-I, which might represent a neurotransmitter for these sensory neurons.
Differential expression of TGF β1, β2 and β3 genes during mouse embryogenesis
Authors: Schmid Peter , Cox David , Bilbe Graeme , Maier Rainer , Mcmaster Gary K.
Published: April 2021

We have examined by Northern analysis and in situ hybridisation the expression of TGF β1 β2and β3 during mouse embryogenesis. TGF β1 is expressed predominantly in the mesodermal components of the embryo e.g. the hematopoietic cells of both fetal liver and the hemopoietic islands of the yolk sac, the mesenchymal tissues of several internal organs and in ossifying bone tissues. The strongest TGF β2 signals were found in early facial mesenchyme and in some endodermal and ectodermal epithelial cell layers e.g., lung and cochlea epithelia. TGF β3 was strongest in prevertebral tissue, in some mesothelia and in lung epithelia. All three isoforms were expressed in bone tissues but showed distinct patterns of expression both spatially and temporally. In the root sheath of the whisker follicle, TGF β1 β2 and β3 were expressed simultaneously. We discuss the implication of these results in regard to known regulatory elements of the TGF β genes and their receptors.
Embryonic gene expression patterns of TGF β1, β2 and β3 suggest different developmental functions in vivo
Authors: Millan Fergus A. , Denhez Fabienne , Kondaiah Paturu , Akhurst Rosemary J.
Published: April 2021

We have compared the expression of the genes encoding transforming growth factors β1, β2 and β3 during mouse embryogenesis from 9.5 to 16.5 days p.c. using in situ hybridisation to cellular RNAs. Each gene has a different expression pattern, which gives some indication of possible biological function in vivo.

All three genes appear to be involved in chondro ossification, though each is expressed in a different cell type. Transcripts of each gene are also present in embryonic epithelia. Epithelial expression of TGF β1, β2 and β3 RNA is associated with regions of active morphogenesis involving epithelial-mesenchymal inter actions. In addition, widespread epithelial expression of TGF β2 RNA can be correlated with epithelial differentiation per se. The localisation of TGF β2 RNA in neuronal tissue might also be correlated with differentiation. Finally both TGF β1 and β2 transcripts are seen in regions actively undergoing cardiac septation and valve formation, suggesting some interaction of these growth factors in this developmental process.
T-cadherin expression alternates with migrating neural crest cells in the trunk of the avian embryo
Authors: Ranscht Barbara , Bronner-Fraser Marianne
Published: April 2021

Trunk neural crest cells and motor axons move in a segmental fashion through the rostral (anterior) half of each somitic sclerotome, avoiding the caudal (posterior) half. This metameric migration pattern is thought to be caused by molecular differences between the rostral and caudal portions of the somite. Here, we describe the distribution of T-cadherin (truncated-cadherin) during trunk neural crest cell migration. T-cadherin, a novel member of the cadherin family of cell adhesion molecules was selectively expressed in the caudal half of each sclerotome at all times examined. T-cadherin immunostaining appeared graded along the rostrocau-dal axis, with increasing levels of reactivity in the caudal halves of progressively more mature (rostral) somites. The earliest T-cadherin expression was detected in a small population of cells in the caudal portion of the somite three segments rostral to last-formed somite. This initial T-cadherin expression was observed concomitant with the invasion of the first neural crest cells into the rostral portion of the same somite in stage 16 embryos. When neural crest cells were ablated surgically prior to their emigration from the neural tube, the pattern of T-cadherin immunoreactivity was unchanged compared to unoperated embryos, suggesting that the metameric T-cadherin distribution occurs independent of neural crest cell signals. This expression pattern is consistent with the possibility that T-cadherin plays a role in influencing the pattern of neural crest cell migration and in maintaining somite polarity.
Nuclear–cytoplasmic interactions during ovine oocyte maturation
Authors: Sun F. Z. , Moor R. M.
Published: April 2021

The present studies have been undertaken to investigate the interactions that occur between the nucleus and cytoplasm of ovine oocytes at various stages during meiotic maturation. We report that the nucleus of ovine fully grown dictyate stage oocytes can be efficiently removed by a microsurgical enucleation procedure. It is demonstrated that between the initiation of maturation and germinal vesicle breakdown certain newly syn-thesized polypeptides are selectively sequestered in the oocyte nucleus and the major sequestered polypeptide has a relative molecular mass of 28 000, which represent at least 9 % of the total labelled polypeptides transferred to the oocyte nucleus during the first 4h of maturation. The experiments provide evidence that the removal of the oocyte nucleus at various times before germinal vesicle breakdown (GVBD) does not prevent the major series of changes in protein synthesis that occurs after entry into a metaphase. We conclude therefore that the mixing of the nucleoplasm and cytoplasm is not essential for the initiation or progression of the protein repro-gramming process during maturation. In addition, the experiments show that the development of the ability to condense chromatin during ovine oocyte maturation is independent of the oocyte nucleus. The combined results strongly support the hypothesis that the extensive series of translational changes that occur in oocytes during maturation are controlled by cytoplasmic rather than nuclear factors.
Interactions of the Drosophila gap gene giant with maternal and zygotic pattern-forming genes
Authors: Eldon Elizabeth D. , Pirrotta Vincenzo
Published: April 2021

The Drosophila gene giant (gt) is a segmentation gene that affects anterior head structures and abdominal segments A5–A7. Immunolocalization of the gt product shows that it is a nuclear protein whose expression is initially activated in an anterior and a posterior domain. Activation of the anterior domain is dependent on the maternal bicoid gradient while activation of the posterior domain requires maternal nanos gene product. Initial expression is not abolished by mutations in any of the zygotic gap genes. By cellular blastoderm, the initial pattern of expression has evolved into one posterior and three anterior stripes of expression. The evolution, position and width of these stripes are dependent on interactions between gt and the other gap genes. In turn, gt activity in these domains affects the expression of the other gap genes. These interactions, typical of the crossregulation previously observed among gap genes, confirm that gt is a member of the gap gene class whose function is necessary to establish the overall pattern of gap gene expression. After cellular blastoderm, gt protein continues to be expressed in the head region in parts of the maxillary and mandibular segments as well as in the labrum. Expression is never detected in the labial or thoracic segment primordia but persists in certain head structures, including the ring gland, until the end of embryonic development.
Independent roles of centrosomes and DNA in organizing theDrosophilacytoskeleton
Authors: Yasuda Glenn K. , Baker Jayne , Schubiger Gerold
Published: April 2021

The early embryonic divisions of Drosophila melanogaster are characterized by rapid, synchronized changes of the nuclei and surrounding cytoskeleton. We report evidence that these changes are carried out by two separately organized systems. DNA was sufficient to cause assembly of nuclear lamina and the formation of nuclear membrane with pore structures. Free centrosomes were correlated with the formation of microtubule, microfilament and spectrin networks in the absence of nuclei. In addition, we found that the morphology of the cytoskeleton associated with the free centrosomes cycled in response to the embryonic cell cycle cues. These observations suggest that the centrosomes may be responsible for the organization of this extensive cytoskeleton. The early divisions may therefore result from the independent cycling of two systems, the nucleus and the surrounding cytoskeleton, that respond separately to the mitotic cues in the embryo and function together to give the synchronized early divisions. The Drosophila embryo has an ‘intermediate’ mitotic system in which the nuclear membrane does not break down completely during mitosis. We speculate that the principles of cytoskeleton organization in this system may be different from those of the Xenopus ‘open’ mitotic system.
Analysis of distribution patterns of gap junctions during development of embryonic chick facial primordia and brain
Authors: Minkoff Robert , Parker Susan B. , Hertzberg Elliot L.
Published: April 2021

Gap junction distribution in the facial primordia of chick embryos at the time of primary palate formation was studied employing indirect immunofluorescence localization with antibodies to gap junction proteins initially identified in rat liver (27×103Mr, connexin 32) and heart (43×103Mr, connexin 43). Immunolocalization with antibodies to the rat liver gap junction protein (27 ×103Mr) demonstrated a ubiquitous and uniform distribution in all regions of the epithelium and mesenchyme except the nasal placode. In the placodal epithelium, a unique non-random distribution was found characterized by two zones: a very heavy concentration of signal in the superficial layer of cells adjacent to the exterior surface and a region devoid of detectable signal in the interior cell layer adjacent to the mesenchyme. This pattern was seen during all stages of placode invagination that were examined. The separation of gap junctions in distinct cell layers was unique to the nasal placode, and was not found in any other region of the developing primary palate. One other tissue was found that exhibited this pattern -the developing neural epithelium of the brain and retina. These observations suggest the presence of region-specific signaling mechanisms and, possibly, an impedance of cell communication among subpopulations of cells in these structures at critical stages of development. Immunolocalization with antibodies to the ‘heart’ 43 × 103Mr gap junction protein also revealed the presence of gap junction protein in facial primordia and neural epithelium. A non-uniform distribution of immunoreactivity was also observed for connexin 43.
The relationship between cell proliferation and the transcription of the nuclear oncogenes c-myc, c-myb and c-ets-1 during feather morphogenesis in the chick embryo
Authors: Desbiens Xavier , Quéva Christophe , Jaffredo Thierry , Stéhelin Dominique , Vandenbunder Bernard
Published: April 2021

We have described the expression of three nuclear protooncogenes, c-myc, c-myb and c-ets-1 during feather morphogenesis in the chick embryo. In parallel with the expression patterns obtained by in situ hybridization, we have mapped the spatial distribution of S-phase cells by monitoring the incorporation of 5-bromodeoxyuridine.

We do not detect c-myc or c-myb transcripts during the early stages when S-phase cells are scattered in the dermis and in the epidermis. Rather c-ets-1 transcripts are abundant in the dermal cells which divide and accumulate under the uniform epidermis. At the onset of the formation of the feather bud, cells within each rudiment cease DNA replicative activities and c-myc transcripts are detected both in the epidermis and in the underlying dermis. This expression precedes the reentry into the S phase. The transcription of c-myb, which has been previously tightly linked to hemopoietic cells is also detected in the developing skin. This expression is essentially located in proliferating epidermal cells on and after the beginning of feather outgrowth. As feather outgrowth proceeds, the distribution of c-myc and c-myb transcripts is restricted to the highly proliferating epidermis. In contrast c-ets-1 transcripts are never detected in the epidermis. During the later stages of skin morphogenesis, the transcription of c-ets-1 is restricted to the endothelial cells of blood vessels, as previously described.

We suggest that the differential expression of these nuclear oncogenes reflects the activation of different mitotic controlling pathways during the development of the skin.
Genetically identical parthenogenetic mouse embryos produced by inhibition of the first meiotic cleavage with cytochalasin D
Authors: Kubiak Jacek , Paldi Andras , Weber Michèle , Maro Bernard
Published: April 2021

The microfilament inhibitor cytochalasin D inhibits extrusion of the first polar body when present during the first meiotic division of mouse oocytes; however, it does not interfere with anaphase movement of chromosomes, and thus induces the formation of tetraploid oocytes. After the separation of chromosomes in anaphase, two spindles start to assemble. However, they merge rapidly and a single meiotic spindle forms. During the transition between metaphase I and metaphase II, in the presence of cytochalasin D, a drop in histone kinase activity takes place demonstrating a transitional decrease in the activity of the maturation promoting factor. These oocytes can be activated parthenogenetically a few hours after washing out the inhibitor. After completion of the second meiotic division and extrusion of a polar body, they contain a diploid number of chromosomes. They are genetically identical to each other and to their mother. Such eggs develop to the blastocyst stage and can implant in the uteri of foster mothers. Most of these fetuses die before the 9th day of gestation, as do diploid control fetuses treated with cytochalasin D during the second meiotic division. The heterozygous state of the experimental embryos obtained after activation of eggs recovered from heterozygous females and treated with cytochalasin D during the first meiotic .division was confirmed using a glucose-phosphate isomerase assay. This technique allows the production of genetic clones of parthenogenetic embryos by simple means.
In situ analysis of fetal, prepuberal and adult XX ↔ XY chimaeric mouse testes: Sertoli cells are predominantly, but not exclusively, XY
Authors: Palmer Stephen J. , Burgoyne Paul S.
Published: April 2021

The testes of fetal, prepuberal and adult XX ↔ XY chimaeras were examined using in situ hybridisation to identify the β -globin transgenic marker contained in one component of each chimaera. This enabled the proportion of XX and XY cells contributing to the major cell lineages of the testis to be estimated from sectioned and air-dried material. A few XX Sertoli cells were found in all three age groups, but the XX contribution was always much lower than in other somatic cell types. Significantly, in fetal XX↔ XY testes, Sertoli cells were the only cell type to show a bias in favour of the XY component. This strengthens the view that Tdy acts solely in the lineage that gives rise to Sertoli cells. However, the finding of some fetal XX Sertoli cells means that one of the steps in the Trfy-initiated process of Sertoli cell determination is capable of locally recruiting XX cells.
Developmental expression of fibrillarin and U3 snRNA inXenopus laevis
Authors: Caizergues-Ferrer Michèle , Mathieu Colette , Mariottini Paolo , Amalric Francois , Amaldi Francesco
Published: April 2021

Fibrillarin is one of the protein components that together with U3 snRNA constitute the U3 snRNP, a small nuclear ribonucleoprotein particle involved in ribosomal RNA processing in eucaryotic cells. Using an antifibrillarin antiserum for protein detection and a fibrillarin cDNA and a synthetic oligonucleotide complementary to U3 snRNA as hybridization probes, the expression of these two components has been studied during Xenopus development. Fibrillarin mRNA is accumulated early in oogenesis, like many other messengers, and translated during oocyte growth. Fibrillarin protein is thus progressively accumulated throughout oogenesis to be assembled with U3 snRNA and used for ribosome production in the amplified nucleoli. After fertilization, the amount of U3 snRNA decreases while the maternally accumulated fibrillarin mRNA is maintained and utilized to produce more protein. After the mid-blastula transition, stored fibrillarin is assembled with newly synthesized U3 snRNA and becomes localized in the prenucleolar bodies and reforming nucleoli.
Drosophila gastrulation: analysis of cell shape changes in living embryos by three-dimensional fluorescence microscopy
Authors: Kam Zvi , Minden Jonathan S. , Agard David A. , Sedat John W. , Leptin Maria
Published: April 2021

The first event of Drosophila gastrulation is the formation of the ventral furrow. This process, which leads to the invagination of the mesoderm, is a classical example of epithelial folding. To understand better the cellular changes and dynamics of furrow formation, we examined living Drosophila embryos using three-dimensional time-lapse microscopy. By injecting fluorescent markers that visualize cell outlines and nuclei, we monitored changes in cell shapes and nuclear positions. We find that the ventral furrow invaginates in two phases. During the first ‘preparatory’ phase, many prospective furrow cells in apparently random positions gradually begin to change shape, but the curvature of the epithelium hardly changes. In the second phase, when a critical number of cells have begun to change shape, the furrow suddenly invaginates. Our results suggest that furrow formation does not result from an ordered wave of cell shape changes, contrary to a model for epithelial invagination in which a wave of apical contractions causes invagination. Instead, it appears that cells change their shape independently, in a stochastic manner, and the sum of these individual changes alters the curvature of the whole epithelium.
mRNA localization studies suggest that murine FGF-5 plays a role in gastrulation
Authors: Hébert Jean M. , Boyle Monica , Martin Gail R.
Published: April 2021

During gastrulation in the mouse, the pluripotent embryonic ectoderm cells form the three primary germ layers, ectoderm, mesoderm and endoderm. Little is known about the mechanisms responsible for these processes, but evidence from previous studies in amphibians, as well as expression studies in mammals, suggest that signalling molecules of the Fibroblast Growth Factor (FGF) family may play a role in gastrulation. To determine whether this might be the case for FGF-5 in the mouse embryo, we carried out RNA in situ hybridization studies to determine when and where in the early postimplantation embryo the Fgf-5 gene is expressed. We chose to study this particular member of the FGF gene family because we had previously observed that its pattern of expression in cultures of teratocarcinoma cell aggregates is consistent with the proposal that Fgf-5 plays a role in gastrulation in vivo. The results reported here show that Fgf-5 expression increases dramatically in the pluripotent embryonic ectoderm just prior to gastrulation, is restricted to the cells forming the three primary germ layers during gastrulation, and is not detectable in any cells in the embryo once formation of the primary germ layers is virtually complete. Based on this provocative expression pattern and in light of what is known about the functions in vitro of other members of the FGF family, we hypothesize that in the mouse embryo Fgf-5 functions in an autocrine manner to stimulate the mobility of the cells that contribute to the embryonic germ layers or to render them competent to respond to other inductive or positional signals.
Neuroanatomical and functional analysis of neural tube formation in notochordless Xenopus embryos; laterality of the ventral spinal cord is lost
Authors: Clarke J. D. W. , Holder Nigel , Soffe S. R. , Storm-Mathisen J.
Published: April 2021

Notochordless Xenopus embryos were produced by u.v. irradiation of the uncleaved fertilized egg. The spinal cords were examined using intermediate filament staining for glial cells, retrograde HRP staining for neuronal morphology and an anti-glycinergic antibody to reveal commissural cells and axons. The floorplate cells of the normal cord appear to be absent and their position along the ventral midline of the cord is occupied by motor neurones, Kolmer-Agduhr cells, radial glial cells and a ventrally placed marginal zone containing the longitudinal axons. Motor neurone number is reduced to 15 % of control values, and the sensory extramedullary cell number is increased twentyfold. Commissural axons are still able to cross the ventral cord but do so at abnormal angles and some commissural axons continue to grow circumferentially up the contralateral side of the cord rather than turning to grow longitudinally. Extracellular electrophysiological recordings from motor axons reveal that the normal alternation of locomotor activity on the left and right side of the embryo is lost in notochordless animals. These results suggest that the notochord and/or the normal floor plate structure are important for the development of the laterality of spinal cord connections and may influence motor neurone proliferation or differentiation.
Two C. elegans genes control the programmed deaths of specific cells in the pharynx
Authors: Ellis Ronald E. , Horvitz H. Robert
Published: April 2021

The genes ces-1 and ces-2 control the decisions of two cells in the nematode Caenorhabditis elegans to undergo programmed cell death. Mutations that cause a gain of ces-1 function or a reduction of ces-2 function prevent these cells, the sisters of the two pharyngeal NSM neurons, from dying. These mutations do not affect most other cell deaths. Genetic studies indicate that ces-1 and ces-2 affect the fates of the NSM sisters by regulating the genes required for all programmed cell deaths to occur.
Nerve-dependent and -independent tenascin expression in the developing chick limb bud
Authors: Wehrle-Haller Bernhard , Koch Manuel , Baumgartner Stefan , Spring Jürg , Chiquet Matthias
Published: April 2021

The extracellular matrix protein, tenascin, appears in a restricted pattern during organ morphogenesis. Tenascin accumulates along developing peripheral nerves as they leave the spinal cord and enter the limb mesenchyme (Wehrle and Chiquet, Development 110, 401– 415, 1990). Here we found that most but not all tenascin deposited along growing nerves is of glial origin. By in situ hybridization with a tenascin cDNA probe, we determined the site of tenascin mRNA accumulation both in normal and nerve-free limbs. In normal wing buds, tenascin mRNA was first detected within the developing limb nerves. Vinculin-positive glial precursor cells, which comigrate with the axons, are the likely source of this tenascin message. In nerveless wing grafts, tenascin was first expressed in tendon primordia in the absence, and thus independently, from innervation. In contrast to normal limbs, grafted wing buds neither contained vinculin-positive glial precursor cells, nor expressed tenascin in regions proximal to tendon primordia. In normal wing buds, tenascin deposited by tendon primordia transiently parallels and surrounds certain developing nerves. After the major nerve pattern is established, tenascin mRNA disappears from nerves in the upper limb, but is expressed in perichondrium and tendons. We propose that glial tenascin facilitates the penetration of axons into the limb bud and is important for nerve fasciculation. In some places, early tendon primordia might help to guide the migration of axons and glial precursor cells towards their target. Abbreviations: ECM, extracellular matrix; TBS, Tris buffered saline; mAb, monoclonal antibody, IgG, immunoglobulin gamma; SDS, sodium dodecyl sulfate; TGF-β, transforming growth factor-β; FCS, fetal calf serum.