bluuebunny/crossref_metadata_2025 huggingface dataset:
The role of retinoid-binding proteins in the generation of pattern in the developing limb, the regenerating limb and the nervous system
Authors: Maden M. , Ong D. E. , Summerbell D. , Chytil F.
Published: April 2021

We summarise existing data and describe new information on the levels and distribution of cellular retinoic acid-binding protein (CRABP) and cellular retinol-binding protein (CRBP) in the regenerating axolotl limb, the developing chick limb bud and the nervous system of the chick embryo in the light of the known morphogenetic effects of retinoids on these systems. In the regenerating limb, levels of CRABP rise 3- to 4-fold during regeneration, peaking at the time when retinoic acid (RA) is most effective at causing pattern duplications. The levels of CRBP are low. The potency of various retinoids in causing pattern respecification correlates well with the ability of these compounds to bind to CRABP. In the chick limb bud, the levels of CRABP are high and the levels of CRBP are low. Again the binding of various retinoids to CRABP correlates well with their ability to cause pattern duplications. By immunocytochemistry, we show that CRABP is present at high levels in the progress zone of the limb bud and is distributed across the anteroposterior axis in a gradient with the high point at the anterior margin. In the chick embryo, CRABP levels are high and CRBP levels are low. By immunocytochemistry, CRABP is localised primarily to the developing nervous system, labelling cells and axons in the mantle layer of the neural tube. These become the neurons of the commissural system. Also sensory axons label intensely with CRABP whereas motor axons do not and in the mixed nerves at the brachial plexus sensory and motor components can be distinguished on this basis. In the neural tube, CRBP only stains the ventral floor plate. Since the ventral floor plate may be a source of chemoattractant for commissural axons, we suggest on the basis of these staining patterns that RA may fulfill this role and thus be involved morphogenetically in the developing nervous system.
The role of fibroblast growth factor in early Xenopus development
Authors: Slack J. M. W. , Darlington B. G. , Gillespie L. L. , Godsave S. F. , Isaacs H. V. , Paterno G. D.
Published: April 2021

In early amphibian development, the mesoderm is formed around the equator of the blastula in response to an inductive signal from the endoderm. A screen of candidate substances showed that a small group of heparin-binding growth factors (HBGFs) were active as mesoderm-inducing agents in vitro. The factors aFGF, bFGF, kFGF and ECDGF all show similar potency and can produce inductions at concentrations above about 100 pM. The product of the murine int-2 gene is also active, but with a lower specific activity. Above the induction threshold there is a progressive increase of muscle formation with dose. Single blastula ectoderm cells can be induced and will differentiate in a defined medium to form mesodermal tissues. All inner blastula cells are competent to respond to the factors but outer cells, bearing oocyte-derived membrane, are not.

Inducing activity can be extracted from Xenopus blastulae and binds to heparin like the previously described HBGFs. Antibody neutralization and Western blotting experiments identify this activity as bFGF. The amounts present are small but would be sufficient to evoke inductions in vivo. It is not yet known whether the bFGF is localized to the endoderm, although it is known that inducing activity secreted by endodermal cells can be neutralized by heparin.

The competence of ectoderm to respond to HBGFs rises from about the 128-cell stage and falls again by the onset of gastrulation. This change is paralleled by a rise and fall of binding of 125I-aFGF. Chemical cross-linking reveals that this binding is attributable to a receptor of relative molecular mass about 130 ×103. The receptor is present both in the marginal zone, which responds to the signal in vivo, and in the animal pole region, which is not induced in vivo but which will respond to HBGFs in vitro.

In the embryo, the induction in the vicinity of the dorsal meridian is much more potent than that around the remainder of the marginal zone circumference. Dorsal inductions contain notochord and will dorsalize ventral mesoderm with which they are later placed in contact. This effect might be due to a local high bFGF concentration or, more likely, to the secretion in the dorsal region of an additional, synergistic factor. It is known that TGF-β-1 and -2 can greatly increase the effect of low doses of bFGF, although it has not yet been demonstrated that they are present in the embryo. Lithium salts have a dorsalizing effect on whole embryos or on explants from the ventral marginal zone, and also show potent synergism when applied together with HBGFs.
Neural expression of the Xenopus homeobox gene Xhox3: evidence for a patterning neural signal that spreads through the ectoderm
Authors: Altaba A. Ruiz i
Published: April 2021

The Xenopus laevis homeobox gene Xhox3 is expressed in the axial mesoderm of gastrula and neurula stage embryos. By the late neurula-early tailbud stage, mesodermal expression is no longer detectable and expression appears in the growing tailbud and in neural tissue. In situ hybridization analysis of the expression of Xhox3 in neural tissue shows that it is restricted within the neural tube and the cranial neural crest during the tailbudearly tadpole stages. In late tadpole stages, Xhox3 is only expressed in the mid/hindbrain area and can therefore be considered a marker of anterior neural development. To investigate the mechanism responsible for the anterior-posterior (A-P) regionalization of the neural tissue, the expression of Xhox3 has been analysed in total exogastrula. In situ hybridization analyses of exogastrulated embryos show that Xhox3 is expressed in the apical ectoderm of total exogastrulae, a region that develops in the absence of anterior axial mesoderm. The results provide further support for the existence of a neuralizing signal, which originates from the organizer region and spreads through the ectoderm. Moreover, the data suggest that this neural signal also has a role in A-P patterning the neural ectoderm.
Expression of nerve growth factor receptor mRNA during early development of the chicken embryo: emphasis on cranial ganglia
Authors: Hallböök Finn , Ayer-Lelièvre Christiane , Ebendal Ted , Persson Häkan
Published: April 2021

In situ hybridization with β-nerve growth factor receptor (NGF-R) oligonucleotide probes was used to study NGF-R mRNA expression in early chicken embryos. Sections through the region of the visceral arches showed high levels of NGF-R mRNA in mesenchyme of the visceral arches, neural tube and myotomes. Labelling was also seen over E3 primordium of the trigeminal ganglion (V) and in the placodal thickening of the petrosal (IX) and nodose (X) ganglionic primordia. In the E5 embryo, all cranial sensory ganglia (V, VII, VIII, IX, X) expressed NGF-R mRNA although at varying levels with higher levels in the ganglia of the Vth, IXth and Xth cranial nerves than in ganglia of the VIIth and the VIIIth nerves. Within ganglia of the Vth, IXth and Xth cranial nerves, levels of NGF-R mRNA were higher in regions containing placode-derived neurons, than in regions with neural-crest-derived neurons. The placode-derived nodose ganglion (X) expressed NGF-R mRNA at all stages of development. In the E15 embryo and later in development, two thirds of the large neuron-like cells expressed high levels of NGF-R mRNA. Our results show that expression of NGF-R mRNA, in peripheral neurons, is not restricted to cells of neural crest origin. We also show a transient expression of NGF-R mRNA early in development in a wide range of non-neuronal differentiating cells. The high level of NGF-R mRNA in early differentiating tissues suggest that the NGF-R plays a wider role during development than previously anticipated.
Genetic imprinting in clinical genetics
Authors: Clarke Angus
Published: April 2021

Genetic, and indeed genomic, imprinting does occur in humans. This is manifest at the level of the genome, the individual chromosome, subchromosomal region or fragile site, or the single locus. The best evidence at the single gene level comes from a consideration of familial tumour syndromes. Chromosomal imprinting effects are revealed when uniparental disomy occurs, as in the Prader-Willi syndrome and doubtless other sporadic, congenital anomaly syndromes. Genomic imprinting is manifest in the developmental defects of hydatidiform mole, teratoma and triploidy. Fragile (X) mental retardation shows an unusual pattern of inheritance, and imprinting can account for these effects. Future work in clinical genetics may identify congenital anomalies and growth disorders caused by imprinting: the identification of imprinting effects for specific chromosomal regions in mice will allow the examination of the homologous chromosomal region in humans.
Mechanisms for the construction and developmental control of heterochromatin formation and imprinted chromosome domains
Authors: Tartof Kenneth D. , Bremer Marilyn
Published: April 2021

The study of variegating position effects in Drosophila provides a model system to explore the mechanism and material basis for the construction and developmental control of heterochromatin domains and the imprinted genomic structures that they may create. The results of our experiments in this regard have implications for a diverse assortment of long-range chromosome phenomena related to gene and chromosome inactivation. Specifically, as a consequence of our studies on position effect variegation, we propose a simple mechanism of X-chromosome inactivation, suggest a purpose for genomic imprinting, and postulate a general means for regulating the time in development at which certain genes become heterochromatically repressed.
The evolution of genomic imprinting
Authors: Chandra H. Sharat , Nanjundiah Vidyanand
Published: April 2021

We explore three possible pathways for the evolution of genomic imprinting. (1) Imprinting may be advantageous in itself when imprinted and unimprinted alleles of a locus confer different phenotypes. If a segment of DNA is imprinted in the gametes of one sex but not in those of the other, it might lead to effects correlated with sexual dimorphism. More fundamentally, in certain organisms, sex determination might have evolved because of imprinting. When imprinting leads to chromosome elimination or inactivation and occurs in some embryos but not in others, two classes of embryos, differing in the number of functional gene copies, would result. A model for sex determination based on inequality in the actual or effective copy-number of particular noncoding, regulatory sequences of DNA has been proposed (Chandra, Proc. natn. Acad. Sci. U.S.A. 82. 1165–1169 and 6947–6949, 1985). Maternal control of offspring sex is another possible consequence of imprinting; this would indicate a potential role for imprinting in sex ratio evolution. (2) Genes responsible for imprinting may have pleiotropic effects and they may have been selected for reasons other than their imprinting ability. Lack of evidence precludes further consideration of this possibility. (3) Imprinting could have co-evolved with other traits. For instance, gamete-specific imprinting could lead to a lowered fitness of androgenetic or gynogenetic diploids relative to the fitness of ‘normal’ diploids. This in turn would reinforce the evolution of anisogamy. The reversibility of imprinting raises the possibility of occassional incomplete or improper erasure. If the site of imprinting is the egg -as appears to be the case with the human X (Chandra and Brown, Nature 253. 165–168, 1975) -either improper imprinting or improper erasure could lead to unusual patterns of inheritance (as in the fragile-X syndrome) or fitness effects skipping generations.
04 and A007-sulfatide antibodies bind to embryonic Schwann cells prior to the appearance of galactocerebroside; regulation of the antigen by axon-Schwann cell signals and cyclic AMP
Authors: Mirsky Rhona , Dubois Cathérine , Morgan Louise , Jessen Kristján R.
Published: April 2021

In the rat sciatic nerve, the relationship between Schwann cells, axons, the extracellular matrix and perineurial sheath cells undergoes extensive modification between embryo day 15 and the onset of myelination during the first postnatal day. Little is known about molecular changes in Schwann cells in this important prenatal period.

In the present paper, we use immunofluorescence to study the prenatal development and postnatal regulation of the antigen(s) recognized by the 04 monoclonal antibody and a well-characterized rat monoclonal antibody to sulfatide, A007. We show that, in a series of immunochemlcal tests, the 04 antibody recognizes only sulfatide in neonatal and adult rat nerves. Both antibodies first bind to Schwann cells in the sciatic nerve at embryo day 16-17, and all Schwann cells bind both antibodies at birth. In the adult nerve, both nonmyellnforming and myelin-formlng cells are labelled with the antibodies. Schwann cells dissociated from embryo day 15 nerves and cultured in the absence of axons develop neither 04 nor A007 binding on schedule, and 04-positive and A007-positive Schwann cells from postnatal nerves lose the ability to bind these antibodies during the first few days in culture. Schwann cells in the distal stump of transected nerves also sharply down-regulate cell surface binding of 04. High numbers of 04-positive or A007-positive Schwann cells reappear in cultures treated with agents that mimic or elevate intracellular cAMP. We conclude that two anti-sulfatide antibodies 04 and A007, recognize an antigen, probably sulfatide, that appears very early in Schwann cell development (one to two days prior to galactocerebroside) but is nevertheless subject to upregulation by axonal contact or elevation of intracellular cAMP.
Curvature and spiral geometry in aggregation patterns of Dictyostelium discoideum
Authors: Foerster P. , Müller S. C. , Hess B.
Published: April 2021

Aggregation patterns of the slime mould Dictyostelium discoideum were recorded using dark-field equipment combined with video techniques. Computerized image processing allowed the analysis of wave collision structures, expanding concentric circles and rotating spirals in terms of wave velocity and front geometry, as previously done in the Belousov-Zhabotinskii reaction, a chemical system showing similar patterns. We verified the linear relationship between the normal velocity and the curvature of wave fronts predicted by a reactiondiffusion model. The proportionality factor, which in this case is the diffusion coefficient of the chemical signal transmitter cAMP establishing communication between the cells, was determined to be 0.66×10−5cm2s-1. From measurements of positively curved circular waves, we could roughly estimate the critical radius of wave propagation (∽200//m); which means that up to 500 cells are necessary to form a center of an aggregation structure. Furthermore, we analyzed the geometrical parameters of spiral wave patterns and estimated the core radius ro to be ≈300 μm.
Colocalization of TGF-beta 1 and collagen I and III, fibronectin and glycosaminoglycans during lung branching morphogenesis*
Authors: Heine Ursula I. , Munoz Eliana F. , Flanders Kathleen C. , Roberts Anita B. , Sporn Michael B.
Published: April 2021

The possible in vivo role of TGF-beta 1 in regulating various proteins of the extracellular matrix, including fibronectin, collagen I and III, and glycosaminoglycans, was examined by immunohistochemical methods during critical stages of lung morphogenesis in the 11-to 18-day-old mouse embryo. Sections of Bouin-fixed, paraf-fin-embedded whole embryos were exposed to polyclonal antibodies specific to synthetic peptides present in the precursor part of TGF-beta 1 (pro-TGF-beta 1), in the processed TGF-beta 1 (antibody CC), collagen J and HI, fibronectin, followed by the PAP or ABC technique to visualize the location of the antibody. GAG were stained with Alcian Blue 8GX. Our results indicate colocalization of TGF-beta 1 expression and that of matrix proteins in the developing lung when branching morphogenesis (cleft formation) and tissue stabilization occur. The presence of TGF-beta 1 at the epithelialmesenchymal interfaces of stalks and clefts at a time when matrix proteins can first be visualized in these areas, suggests a direct participation of the growth factor in the development of the basic architecture of the lung.
Expression of a homeobox gene product in normal and mutant zebrafish embryos: evolution of the tetrapod body plan
Authors: Molven Anders , Wright Christopher V. E. , Bremiller Ruth , Robertis Eddy M. De , Kimmel Charles B.
Published: April 2021

An antibody was used to detect antigens in zebrafish that appear to be homologous to the frog homeodomain-containing protein XIHbox 1. These antigens show a restricted expression in the anteroposterior axis and an anteroposterior gradient in the pectoral fin bud, consistent with the distribution of XIHbox 1 protein in frog and mouse embryos. In the somitic mesoderm, a sharp anterior limit of expression coincides exactly with the boundary between somites 4 and 5, and the protein level fades out posteriorly. A similar, graded expression of the antigen is seen within the series of Rohon-Beard sensory neurons of the CNS. We also immunostained the mutant spt-1 (‘spadetail’), in which the trunk mesoderm is greatly depleted and disorganized in the region of XIHbox 1 expression. The defects stem from misdirected cell movements during gastrulation, but nevertheless, newly recruited cells that partially refill the trunk mesoderm express the antigen within the normal span of the anteroposterior axis. This finding suggests that the mutation does not delete positional information required for activation of the XIHbox 1 gene.
Sex determination in loggerhead turtles: differential expression of two hnRNP proteins
Authors: Harry Jenny L. , Williams Keith L. , Briscoe David A.
Published: April 2021

Sex determination in the loggerhead turtle, Caretta caretta, is controlled by incubation temperature during a critical period of embryogenesis. As heat-shock gene expression is temperature-dependent and has been shown to be associated with early developmental regulation in several organisms, we studied the constitutive expression of hsp70 and hsp90 in embryonic brain and urinogenital tissues to see if these proteins are differentially expressed during the sex-determining period in embryos incubated at male- (26 °C) and female-(32 °C) determining temperatures. The level of expression of hsp70 and hsp90, as determined from monoclonal antibody staining, is similar in both sexes during the sexdetermining period. However, AC88, a monoclonal antibody that identifies hsp90 in several systems, recognised two additional protein bands (Mr 42 and 46xl(r), which are differentially expressed in the urinogenital tissue. of developing male and female embryos during the sex-determining period. While the 42K and 46K proteins appear in the urinogenital tissue of developing female (32 °C) embryos until stage 25, they are not expressed in the male (26 °C) urinogenital system after stage 24. Subsequent experiments have identified both turtle proteins as heterogeneous nuclear ribonucleoprotein particles (hnRNPs). As several hnRNP proteins have specific RNA-binding sites and are involved in mRNA processing reactions, the 46K protein may mediate post- transcriptional control of specific RNA transcripts required for sexual differentiation in C. caretta.
Differential growth of facial primordia in chick embryos: responses of facial mesenchyme to basic fibroblast growth factor (bFGF) and serum in micromass culture
Authors: Richman Joy M. , Crosby Zoe
Published: April 2021

Differential growth of the three major facial primordia, the frontonasal mass, maxilla and mandible, results in a characteristic face shape. Abnormal growth of any of the primordia can lead to facial defects. In order to dissect out the factors that control growth, we developed a functional assay for cell proliferation using micromass culture and defined medium. Cell number was determined over a 4 day period and BrdU incorporation was used to determine the percentage of cells in S-phase. In defined medium, cell number progressively decreases and proliferation is very reduced in cultures of cells from all three primordia. When foetal calf serum was added, frontonasal mass cell number triples, mandible doubles and maxilla increases by half. The number of cells in S-phase increased in every case but the final cell number reflects a balance between proliferation and cell loss from the culture. The addition of basic fibroblast growth factor (bFGF) to defined medium leads to an increase in cell number in the frontonasal mass, while the cell number of mandibular and maxillary cultures is relatively unaffected. The percentage of cells in S-phase is highest in frontonasal mass cultures. Serum and bFGF both increase chondrogenesis in frontonasal mass cultures when compared to defined medium. In contrast in mandibular cultures, serum does not change the amount of cartilage and with bFGF chondrogenesis is reduced. The coordination of the changes in proliferation and differentiation in frontonasal mass cultures suggest that either these two processes are independently stimulated to the same extent or a single subpopulation of cells is stimulated to divide and differentiate into chondrocytes. The different responses of the populations of individual facial primordia to growth factors may contribute to differential growth in vivo and also be linked to the generation of facial defects.
The anterior extent of dorsal development of the Xenopus embryonic axis depends on the quantity of organizer in the late blastula
Authors: Stewart Ronald M. , Gerhart John C.
Published: April 2021

In amphibian gastrulae, the cell population of the organizer region of the marginal zone (MZ) establishes morphogenesis and patterning within itself and within surrounding regions of the MZ, presumptive neurecto-derm, and archenteron roof. We have tested the effects on pattern of reducing the amount of organizer region by recombining halves of Xenopus laevis late blastulae cut at different angles from the bilateral plane. When regions within 30° of the dorsal midline are excluded from recombinants, ventralized embryos develop lacking the entire anterior-posterior sequence of dorsal structures, suggesting that the organizer is only 60° wide (centered on the dorsal midline) at the late blastula stage. As more and more dorsal MZ (organizer) is included in the recombinant, progressively more anterior dorsal structures are formed. In all cases, when any dorsal structures are missing they are deleted serially from the anterior end. Thus, we suggest that the amount (lateral width) of the organizer in the MZ determines the anterior extent of dorsal development.
Developmental potential of neural crest-derived cells migrating from segments of developing quail bowel back-grafted into younger chick host embryos
Authors: Rothman Taube P. , Douarin Nicole M. Le , Fontaine-Pérus Josiane C. , Gershon Michael D.
Published: April 2021

The technique of back-transplantation was used to investigate the developmental potential of neural crest-derived cells that have migrated to and colonized the avian bowel. Segments of quail bowel (removed at E4) were grafted between the somites and neural tube of younger (E2) chick host embryos. Grafts were placed at a truncal level, adjacent to somites 14 –24. Initial experiments, done in vitro, confirmed that crest-derived cells are capable of migrating out of segments of foregut explanted at E4. The foregut, which at E4 has been colonized by cells derived from the vagal crest, served as the donor tissue. Comparative observations were made following grafts of control tissues, which included hindgut, lung primordia, mesonephros and limb bud. Additional experiments were done with chimeric bowel in which only the crest-derived cells were of quail origin. Targets in the host embryos colonized by crest-derived cells from the foregut grafts included the neural tube, spinal roots and ganglia, peripheral nerves, sympathetic ganglia and the adrenals, but not the gut. Donor cells in these target organs were immunostained by the monoclonal antibody, NC-1, indicating that they were crest-derived and developing along neural or glial lineages. Some of the crest-derived cells (NC-l-immunoreactive) that left the bowel and reached sympathetic ganglia, but not peripheral nerves or dorsal root ganglia, co-expressed tyrosine hydroxylase immunoreactivity, a neural characteristic never expressed by crest-derived cells in the avian gut. None of the cells leaving enteric back-grafts produced pigment. Cells of mesodermal origin were also found to leave donor explants and aggregate in dermis and feather germs near the grafts. These observations indicate that crest-derived cells, having previously migrated to the bowel, retain the ability to migrate to distant sites in a younger embryo. The routes taken by these cells appear to reflect, not their previous migratory experience, but the level of the host embryo into which the graft is placed. Some of the population of crest-derived cells that leave the back-transplanted gut remain capable of expressing phenotypes that they do not express within the bowel in situ, but which are appropriate for the site in the host embryo to which they migrate.
Localization of vasa, a component of Drosophila polar granules, in maternal-effect mutants that alter embryonic anteroposterior polarity
Authors: Hay Bruce , Jan Lily Yeh , Jan Yuh Nung
Published: April 2021

Cytoplasm at the posterior pole of the early Drosophila embryo, known as polar plasm, serves as a source of information necessary for germ cell determination and for specification of the abdominal region. Likely candidates for cytoplasmic elements important in one or both of these processes are polar granules, organelles concentrated in the cortical cytoplasm of the posterior pole. Females homozygous for any one of the maternal-effect mutations, tudor, oskar, staufen, vasa, or valois give rise to embryos that lack localized polar granules, fail to form the germ cell lineage and have abdominal segment deletions. Using antibodies against a polar granule component, the vasa protein, we find that vasa synthesis or localization is affected by these mutations. In vasa mutants, synthesis of vasa protein is absent or severely restricted. In oskar and staufen mutant females, vasa synthesis appears normal, but the vasa protein is not localized. In tudor and valois mutant females, vasa is localized to the posterior pole of oocytes, but this localization is lost following egg activation. In addition to the posterior localized vasa, there is a low level of vasa distributed throughout the embryo. A function for this distributed vasa is postulated based on the observation that embryos from Bicaudal-D mothers, in which abdominal determinants are incorrectly localized to the anterior pole, do not show any ectopic vasa localization, though abdomen development at the anterior end depends on the amount of vasa protein in the embryo.
Stable lines of transgenic zebrafish exhibit reproducible patterns of transgene expression
Authors: Stuart Gary W. , V1Elkind Juergen R. , Mcmurray James V. , Westerfield Monte
Published: April 2021

To study the frequency of germ-line transformation and to examine the reproducibility of tissue-specific trans gene expression, we produced several lines of transgenic zebrafish expressing a recombinant chloramphenicol acetyltransferase (CAT) gene. Supercoiled plasmids containing both Rous sarcoma virus and SV-40 pro moter sequences upstream of the CAT coding region were injected into zebrafish embryos prior to first cleavage. CAT activity could be detected in batches of injected embryos as early as 8 h and up to at least 12 days post-fertilization. Approximately 18 % of injected fish raised to maturity exhibited CAT activity in their fins, and approximately 5 % of injected fish became stable germ-line transformants. Breeding studies indicated that although transgenic founder fish were frequently germ-line mosaics, transgenic individuals of subsequent generations were fully hemizygous for the transgene marker. The transgenes present in the F] progeny of four independent lines were relatively well expressed in fin and skin, while lower levels of expression were observed in heart, gill and muscle. Little or no CAT expression was observed in the brain, liver and gonad. A monoclonal antibody directed against the CAT gene product consistently revealed variegated patterns of CAT expression in ectodermally derived fin epidermal cells in three of these lines. These results show that it is possible to efficiently produce stable germ-line trans formants of the zebrafish and to observe reproducible tissue-specific patterns of transgene expression in this organism. Possible mechanisms for the variegated ex pression observed within tissues are also considered.
Differential expression of TGF beta isoforms in murine palatogenesis
Authors: Fitzpatrick David R. , Denhez Fabienne , Kondaiah Paturu , Akhurst Rosemary J.
Published: April 2021

We have studied the expression of genes encoding transforming growth factors (TGFs) beta1, beta2 and beta3 during development of the secondary palate in the mouse from 11.5 to 15.5 days postcoitum using in situ hybridisation. The RNA detected at the earliest developmental stage is TGF beta3, which is localised in the epithelial component of the vertical palatal shelf. This expression continues in the horizontal palatal shelf, predominantly in the medial edge epithelium, and is lost as the epithelial seam disrupts, soon after palatal shelf fusion. TGF betal RNA is expressed with the same epithelial pattern as TGF beta3, but is not detectable until the horizontal palatal shelf stage. TGF beta2 RNA is localised to the palatal mesenchyme underlying the medial edge epithelia in the horizontal shelves and in the early postfusion palate. The temporal and spatial distribution of TGF betal, beta! and beta3 RNAs in the developing palate, together with a knowledge of in vitro TGF beta biological activities, suggests an important role for TGF beta isoforms in this developmental pro cess.
Protein kinase C and progesterone-induced maturation in Xenopus oocytes
Authors: Varnold Robert L. , Smith L. Dennis
Published: April 2021

Though progesterone-induced maturation has been studied extensively in Xenopus oocytes, the mechanism whereby the prophase block arrest is released is not well understood. The current hypothesis suggests that a reduction in cAMP and subsequent inactivation of cAMP-dependent protein kinase is responsible for reentry into the cell cycle. However, several lines of evidence indicate that maturation can be induced with out a concomitant reduction in cAMP. We show that the mass of diacylglycérol in whole oocytes and plasma membranes decreases 29% and 10% respectively, within the first 15 sec after the addition of progesterone. Diacylglycerol in plasma membranes further decreased 59 % by 5 min. We also show that the protein kinase C inhibitors sphingosine and staurosporine can induce oocyte maturation. In addition, the synthetic diglycer ide, DiC8, and microinjected PKC can inhibit or delay progesterone-induced maturation. These results together suggest that a transient decrease in protein kinase C activity may regulate entry into the cell cycle. The mechanism whereby DAG is decreased in response to progesterone is unclear. Initial studies show that progesterone leads to a decrease in IP3 suggesting that progesterone may act by reducing the hydrolysis of PIP2. On the other hand, progesterone caused a de crease in the amount of [3H]arachidonate labelling in DAG during the same time suggesting that progesterone may stimulate lipase activity. The relationship between postulated changes in the PKC pathway and those hypothesized for the PKA pathway are discussed.
XY female mice resulting from a heritable mutation in the primary testis determining gene, Tdy
Authors: Lovell-Badge Robin , Robertson Elizabeth
Published: April 2021

Chimeric mice constructed with XY embryonic stem (ES) cells that had been multiply infected with a retro viral vector were used in a genetic screen to look for mutations affecting the sex determination pathway in mice. From a small number of chimeras screened one was identified that gave rise to a low proportion of XY females amongst his offspring. Analysis of the segregat ing patterns of retroviral insertions demonstrated that the mutation was found in a subset of the offspring derived from one originally infected ES cell. However, the mutation appeared to have occurred subsequent to the infection. Some of the XY females proved to be fertile, and the mutant phenotype was found to segregate exclusively with the Y chromosome. Analysis of the offspring also confirmed the absence of any retroviral insertion that could be correlated with the mutation.

Further characterisation of the Y chromosome carry ing the mutation by karyotypic analysis, and by Southern blotting with a range of Y-specific DNA probes suggested that there has been no gross deletion or rearrangement of the Y carrying the mutation. There also appeared to be no loss of Y-specific gene functions apart from that of testis determination. Moreover, the mutation is complemented by Sxr‘, the minimum portion of the mouse Y known to carry Tdy. From the phenotype and deduced location of the mutation, we conclude that it is within the Tdy locus. This is the first such mutation to be described in mice.
The marginal zone and its contribution to the hypoblast and primitive streak of the chick embryo
Authors: Stern Claudio D.
Published: April 2021

The marginal zone of the chick embryo has been shown to play an important role in the formation of the hypoblast and of the primitive streak. In this study, time-lapse filming, fate mapping, ablation and trans plantation experiments were combined to study its contribution to these structures. It was found that the deep (endodermal) portion of the posterior marginal zone contributes to the hypoblast and to the junctional endoblast, while the epiblast portion of the same region contributes to the epiblast of the primitive streak and to the definitive (gut) endoderm derived from it. Within the deep part of the posterior marginal zone, a subpopu lation of HNK-l-positive cells contributes to the hypo blast.

Removal of the deep part of the marginal zone prevents regeneration of the hypoblast but not the formation of a primitive streak. Removal of both layers of the marginal zone leads to a primitive streak of abnormal morphology but mesendodermal cells never the less differentiate.

These results show that the two main properties of the posterior marginal zone (contributing to the hypoblast and controlling the site of primitive streak formation) are separable, and reside in different germ layers. This conclusion does not support the idea that the influence of the posterior marginal zone on the development of axial structures is due to it being the source of secondary hypoblast cells.
Pollen-specific gene expression in transgenic plants: coordinate regulation of two different tomato gene promoters during microsporogenesis
Authors: Twell David , Yamaguchi Judy , Mccormick Sheila
Published: April 2021

To investigate the regulation of gene expression during male gametophyte development, we analyzed the pro moter activity of two different genes (LAT52 and LAT59) from tomato, isolated on the basis of their anther-specific expression. In transgenic tomato, tobacco and Arabidopsis plants containing the LAT52 promoter region fused to the β-glucuronidase (GUS) gene, GUS activity was restricted to pollen. Transgenic tomato, tobacco and Arabidopsis plants containing the LAT59 promoter region fused to GUS also showed very high levels of GUS activity in pollen. However, low levels of expression of the LAT59 promoter construct were also detected in seeds and roots. With both constructs, the appearance of GUS activity in developing anthers was correlated with the onset of microspore mitosis and increased progressively until anthesis (pollen shed). Our results demonstrate co-ordinate regulation of the LAT52 and LAT59 promoters in developing microspores and suggest that the mechanisms that regulate pollen-specific gene expression are evolutionarily conserved.
Sperm-egg Association and its Relationship to the Activation of the Egg in Salmonid Fishes
Authors: Ginsburg Anna S.
Published: April 2021

A study has been made of the initial stage of fertilization in Salmonids, using the lake trout Salmo trutta L. morpha lacustris Linné and the salmon S. salar L. morpha sebago Girard. Lake trout spermatozoa are activated by coelomic fluid and by Ringer solution; in these media they retain their motility and fertilizing capacity much longer than in water.At the moment when the spermatozoa kept in coelomic fluid or Ringer solution lose their fertilizing capacity, they are not reactivated by the addition of water. After a period of ‘rest’, however, a proportion of them acquires motility upon the addition of water which leads to a partial restoration of the fertilizing capacity of the sperm suspension.When trout and salmon eggs are inseminated in coelomic fluid or Ringer solution, the union of the gametes is achieved without egg activation. A cytological investigation shows that the spermatozoon reaches the cortical egg layer along the micropylar canal and the anterior part of its head gets embedded in the cytoplasm.When trout spermatozoa are placed into egg water or other media containing products of egg secretion no changes resembling the extrusion of the acrosome filament are seen with the electron microscope. This supports previous data on the absence of the acrosome in salmonid spermatozoa.The development of eggs inseminated in coelomic fluid or Ringer solution is initiated by immersion into water. The involvement of the fertilizing spermatozoon in development is possible upon egg activation for up to several days after insemination. To achieve such an effect, a brief exposure to water is necessary (this varies from 10 to 70 sec. for different eggs).Normal development of the egg may be achieved upon its activation either after or before insemination (the only limitation being the fact that the penetration of the spermatozoon in the activated egg is possible only before the expulsion of the contents of the cortical alveoli in the micropylar region).When water pretreated eggs are inseminated in calcium-free medium, no involvement of the spermatozoon in development takes place.The latent period of the cortical reaction on activation of trout eggs by water (from the moment of appearance of changes setting the process in train until the beginning of the expulsion of the contents of the cortical alveoli) in the bulk of the eggs takes 40–50 sec. at 3 ·0°C and 30–40 sec. at 8 · 9°C.The phenomena of union of the gametes and egg activation in salmonid fishes are to a considerable degree autonomous : they can easily be separated in time and combined in any sequence. This is related to the peculiarities of the mechanism of sperm-egg interaction.
bicoid mRNA localization signal: phylogenetic conservation of function and RNA secondary structure
Authors: Macdonald Paul M.
Published: April 2021

Transcripts of the bicoid (bcd) gene are localized to the anterior pole of the Drosophila oocyte, thereby allowing formation in the embryo of an anteroposterior gradient of the bcd protein morphogen. We previously showed that a 630 nucleotide portion of the 3′ noncoding region of the bcd mRNA is necessary for this localization, and is sufficient to confer anterior localization on a heterologous transcript. Here I have used a comparative analysis to begin to more precisely define the cis-acting mRNA localization signal. The bcd genes from six additional Drosophila species were cloned, and DNA of the 3′ noncoding regions sequenced. Three of these regions were tested interspecifically for mRNA localization in D. melanogaster and each functioned correctly; these regions must therefore contain the cis-acting signal. The primary sequences, which are up to 50 % divergent from the D. melanogaster gene, show patchy homology throughout most of the region. Interestingly, all seven species can potentially form a large stereotypic secondary structure. This structure is a likely candidate for the localization signal and can be used for the rational design of mutations to test that possibility.
The Drosophila transcript encoded by the β-amyloid protein precursor-like gene is restricted to the nervous system
Authors: Martin-Morris Linda E. , White Kalpana
Published: April 2021

We have molecularly delineated a Drosophila β-amyloid protein precursor-like (Appl) gene and analyzed its pattern of expression. Appl defines a new locus within the IB division of the X-chromosome, a region previously shown to be important for neural development. The genomic limits of the Appl gene were defined by mapping of the Appl cDNAs. The Appl transcript spans ∼38kb (1kb=103 base-pairs) of genomic DNA. Genomic regions surrounding the first two exons were sequenced. The first exon contains 78 nucleotides of the coding sequence and is separated from the second exon by a ∼21 kb intron. The second exon is 171 nucleotides long and is separated from the third exon by a ∼7 kb intron.

We present in situ RNA localization data that demonstrate that the Appl transcript is found in post-mitotic neurons in all developmental stages, in the central and peripheral nervous systems. Within the nervous system transcripts are not observed in neuroblasts, newly generated neurons and at least one class of presumed glial cells. The temporal and spatial specificity of Appl expression suggests that the gene product has a function that is common to most neurons.

Appl cDNA predicts an 886-amino acid polypeptide that exhibits strong sequence similarity to the human β-amyloid protein precursor (APP) (Rosen et al. 1989). In this paper, we compare the Appl gene expression with the pattern of expression of the β-amyloid protein precursor (APP) gene in mammals. Furthermore, we suggest that during evolution, a neural-specific function encoded by the APP gene has been selectively maintained.
Distribution and migration pathways of HNK-1-immunoreactive neural crest cells in teleost fish embryos
Authors: Sadaghiani Bahram , Vielkind Juergen R.
Published: April 2021

Whole mounts and cross-sections of embryos from three species of teleost fish were immunostained with the HNK-1 monoclonal antibody, which recognizes an epitope on migrating neural crest cells. A similar distribution and migration was found in all three species. The crest cells in the head express the HNK-1 epitope after they have segregated from the neural keel. The truncal neural crest cells begin to express the epitope while they still reside in the dorsal region of the neural keel; this has not been observed in other vertebrates. The cephalic and anterior truncal neural crest cells migrate under the ectoderm; the cephalic cells then enter into the gill arches and the anterior truncal cells into the mesentery of the digestive tract where they cease migration. These cephalic and anterior trunk pathways are similar to those described in Xenopus and chick. The neural crest cells of the trunk, after segregation, accumulate in the dorsal wedges between the somites, however, unlike in chick and rat, they do not migrate in the anterior halves of the somites but predominantly between the neural tube and the somites, the major pathway observed in carp and amphibians; some cells migrate over the somites. The HNK-1 staining of whole-mount embryos revealed a structure resembling the Rohon-Beard and extramedullary cells, the primary sensory system in amphibians. Such a system has not been described in fish.
Expression of extracellular matrix-degrading metalloproteinases and metalloproteinase inhibitors is developmentally regulated during endoderm differentiation of embryonal carcinoma cells
Authors: Adler Richard R. , Brenner Carol A. , Werb Zena
Published: April 2021

The differentiation of F9 and PSA-1 embryonal carcinoma cells to embryoid bodies composed of a mixture of parietal and visceral endoderm was accompanied by changes in their secretion of metalloproteinases. Differentiation was induced by retinoic acid and dibutyryl cyclic AMP (for F9 cells) or by removing cells from a substrate of feeder cells to alter cell-cell interaction and adhesion (for. PSA-1 cells). The embryoid bodies attached to gelatin-coated dishes, and the parietal endoderm cells spread out over the matrix. The differentiated cells secreted specific gelatin- and casein-degrading proteinases, including enzymes that comigrated with proenzyme forms of collagenase and stromelysin. Total proteinase activity as well as specific collagenase activity increased with the time of differentiation. All of the gelatin- and casein-degrading proteinases detectable by substrate gel zymography were inhibited by inhibitors of metalloproteinases but not by inhibitors of serine or cysteine proteinases, indicating that they were metalloproteinases. Both cell lines showed increased collagenolytic activity, which was activated by treatment with plasmin. In addition, both cell lines showed increased secretion of specific metalloproteinase inhibitors, including tissue inhibitor of metalloproteinases, with differentiation. Analysis of mRNA from undifferentiated and differentiated F9 cells by RNA blot analysis or reverse transcription coupled with the polymerase chain reaction showed that increased expression of genes for collagenase, stromelysin and tissue inhibitor of metalloproteinases is associated with differentiation of these cells. These results suggest that the expression of extracellular matrix-degrading metalloproteinases and their inhibitors is developmentally regulated during the differentiation and spreading of the parietal endoderm.
Receptors for epidermal growth factor and insulin-like growth factor-I on preimplantation trophoderm of the pig
Authors: Corps A. N. , Brigstock D. R. , Littlewood C. J. , Brown K. D.
Published: April 2021

12sI-labelled epidermal growth factor (125I-EGF) and 125I-labelled insulin-like growth factor-I (12SI-IGF-I) bound to trophodenn cells from pig blastocysts obtained on days 15 –19 of pregnancy. Specific binding was detected on freshly isolated cell suspensions and on cells cultured for several days. The binding of 12SI-EGF was inhibited by increasing concentrations of EGF, but not by various other growth factors and hormones. Chemical cross-linking of 12SI-EGF to its receptors using disuccinimidyl suberate (DSS) revealed a radiolabelled band of relative molecular mass 160 000, similar to that identified as the EGF receptor in other cell types. The binding of l2iI-IGF-I was inhibited by both IGF-I and insulin, indicating that the receptors were either type I IGF receptors or insulin receptors. Cross-linking of f25I-IGF-I to serum-free supernatants from trophoderm cultures showed that the cells secreted an IGF-binding protein, giving a complex of relative molecular mass about 45 000. The presence of receptors for EGF and IGF/insulin suggests that these factors could be involved in regulating the growth and development of the early blastocyst.