bluuebunny/crossref_metadata_2025 huggingface dataset:
Detrimental effects of two active X chromosomes on early mouse development
Authors: Takagi Nobuo , Abe Kenji
Published: April 2021

Matings between female mice carrying Searle’s translocation, T(X;16)16H, and normal males give rise to chromosomally unbalanced zygotes with two complete sets of autosomes, one normal X chromosome and one X16 translocation chromosome (XnX16 embryos). Since X chromosome inactivation does not occur in these embryos, probably due to the lack of the inactivation center on X16, XnX16 embryos are functionally disomie for the proximal 63 % of the X chromosome and trisomic for the distal segment of chromosome 16. Developmental abnormalities found in XnX16 embryos include: (1) growth retardation detected as early as stage 9, (2) continual loss of embryonic ectoderm cells either by death or by expulsion into the proamniotic cavity, (3) underdevelopment of the ectoplacental cone throughout the course of development, (4) very limited, if any, mesoderm formation, (5) failure In early organogenesis including the embryo, amnion, chorion and yolk sac. Death occurred at 10 days p.c. Since the combination of XO and trisomy 16 does not severely affect early mouse development, it is likely that regulatory mechanisms essential for early embryogenesis do not function correctly in XnX16 embryos due to activity of the extra X chromosome segment of X16.
The mechanical basis of cell rearrangement I. Epithelial morphogenesis during Fundulus epiboly
Authors: Weliky Michael , Oster George
Published: April 2021

Many morphogenetic processes are accomplished by coordinated cell rearrangements. These rearrangements are accompanied by substantial shifts in the neighbor relationships between cells. Here we propose a model for studying morphogenesis in epithelial sheets by directed cell neighbor change. Our model describes cell rearrangements by accounting for the balance of forces between neighboring cells within an epithelium. Cell rearrangement and cell shape changes occur when these forces are not in mechanical equilibrium. We will show that cell rearrangement within the epidermal enveloping layer (EVL) of the teleost fish Fundulus during epiboly can be explained solely in terms of the balance of forces generated among constituent epithelial cells.

Within a cell, we account for circumferential elastic forces and the force generated by hydrostatic and osmotic pressure. The model treats epithelial cells as two-dimensional polygons where the mechanical forces are applied to the polygonal nodes. A cell node protrudes or contracts when the nodal forces are not in mechanical equilibrium. In an epithelial sheet, adjacent cells share common boundary nodes; in this way, mechanical force is transmitted from cell to cell, mimicking junctional coupling. These junctional nodes can slide, and nodes may appear or disappear, so that the number of polygonal sides is variable.

Computer graphics allows us to compare numerical simulations of the model with time-lapse cinemicroscopy of cell rearrangements in the living embryo, and data obtained from fixed and silver stained embryos. By manipulating the mechanical properties of the model cells we can study the conditions necessary to reproduce normal cell behavior during Fundulus epiboly. We find that simple stress relaxation is sufficient to account for cell rearrangements among interior cells of the EVL when they are isotropically contractile. Experimental observations show that the number of EVL marginal cells continuously decreases throughout epiboly. In order for the simulation to reproduce this behavior, cells at the EVL boundary must generate protrusive forces rather than contractile tension forces. Therefore, the simulation results suggest that the mechanical properties of EVL marginal cells at their leading edge must be quite different from EVL interior cells.
Multiple functions for NGF receptor in developing, aging and injured rat teeth are suggested by epithelial, mesenchymal and neural immunoreactivity
Authors: Byers M. R. , Schatteman G. C. , Bothwell M.
Published: April 2021

We have used immunocytochemistry to analyse expression of nerve growth factor receptor (NGFR) in developing, aging and injured molar teeth of rats. The patterns of NGFR immunoreactivity (IR) in developing epithelia and mesenchyme matched the location of NGFR mRNA assayed by in situ hybridization with a complementary S35-labeled RNA probe. The following categories of NGFR expression were found. (1) There was NGFR-IR in the dental lamina epithelium and in adjacent mesenchyme during early stages of third molar formation. (2) NGFR-IR nerve fibers were posterior and close to the bud epithelium. (3) During crown morphogenesis NGFR expression was prominent in internal enamel epithelium and preodontoblasts; it faded as preameloblasts elongated and as odontoblasts began to make predentin matrix; and it was weak or absent from outer enamel epithelium, the cervical loop, and differentiated ameloblasts and odontoblasts. (4) When NGFR-IR nerve fibers entered the molars late in the bell stage, they innervated the most mature peripheral pulp and dentin in an asymmetric pattern which correlated more with asymmetric enamel synthesis than with mesenchymal NGFR-IR distribution. (5) The mesenchymal pulp cells continued to have intense NGFR expression in adult teeth, especially near coronal tubular dentin. (6) The pulpal NGFR-IR decreased in very old rats or subjacent to reparative dentin (naturally occurring or experimentally induced). (7) During root formation, the preodontoblasts had NGFR-IR but most root mesenchymal cells and Hertwig’s epithelial root sheath did not. This work suggests that there are important epithelial and mesenchymal targets of NGF regulation during molar morphogenesis that differ for crown and root development and that do not correlate with neural development. The continuing expression of NGFR-IR by pulpal mesenchymal cells in adult rats was most intense near coronal odontoblasts making tubular dentin; and it was lost during aging, or subjacent to sites of dentin injury that caused a phenotypic change in the odontoblast layer.
Myelinization of the Optic Nerve and its Dependence on Visual Function— A Quantitative Investigation in Mice
Authors: Gyllensten Lars , Malmfors Torbjörn
Published: April 2021

Quantitative studies have been made of the number, myelinization and diameter of the fibres of the optic nerve during post-natal development in black mice, using phase-contrast and electron microscopy. The influence of complete darkness was investigated. In mice reared in the light no myelinated fibres are present up to 5 days of age, whereas a few hundred are demonstrable at 10 days. Between 15 and 20 days of age, myelinization is intense, the mean number of such fibres per nerve being about 22,000 at 20 days. After this time there is only a slight increase, i.e., to about 25,000 at 30 days. Up to 30 days, only a few per cent of the fibres are more than 1 μ. in diameter (measured in fixed and embedded specimens). In adult mice, three maxima in fibre diameter are observed; the largest group comprises those with a diameter less than 1 μ. (0·6–0·8 μ.), and the two smaller ones fibres more than 1 μ. in diameter (1·1–1·3 and about 2·0 μ.). The fibres are found to be more closely packed in the central part of the optic nerve than in its peripheral parts, but the coarser fibres are more numerous peripherally. No appreciable number of unmyelinated fibres is visible in mature animals. In mice reared in complete darkness for 20 and 30 days, delay in myelinization of the optic nerve fibres causes a reduction in the number of those myelinized by some 12–13 per cent. The coarser fibres are affected to a greater extent than the thin ones; thus at 30 days the number of those more than 1 p in diameter is reduced by almost 60 per cent. In mice reared in darkness, a retardation is observed in growth of the nuclei of the retinal ganglion cells, as well as in their shape maturation (decrease in eccentricity).
Observations of Dependent Histogenesis in Salamander Limb Development
Authors: Finnegan Cyril V.
Published: April 2021

The prospective limb portion of the limb disc of Ambystoma punctatum and Taricha torosa tail-bud stages was transplanted homoplastically and heteroplastically to the ventral hypomere region of similar-aged host animals. The transplants were observed for growth (elongation) and for histogenesis of limb elements and in all cases less limb development was seen, both macroscopically and histologically, than was to be expected from the developmental age of the donors. In a parallel series of experiments a small amount of epibrachial somite tissue was transplanted with the limb cell mass to the ventral hypomere region. The limb histogenesis and growth were enhanced to a considerable degree. It was seen that chondrogenesis (of a girdle) occurred in the somite tissue and the limb cell mass seemed to demonstrate histogenesis (proximally) and elongation (distally) in relation to that development. The limb cell mass was transplanted to various superficial sites on the nonlimb trunk somites and it was observed that more advanced growth and histogenesis were obtained when the transplant was associated with the ventral somite region. Pronephric tissue was developed in all cases where limb cell mass was transplanted to the ventral hypomere region of Taricha torosa hosts. When the amount of dorsal tissue was increased, by adding dorsal ectoderm and/or epibrachial somite to the limb cell mass, no pronephric tissue developed in the graft area. No nephric differentiation was seen with similar transplants to Ambystoma punctatum hosts. The results are considered as indicative of a somite-dependent histogenesis in the development of the proximal portion of the transplanted limb.
The Development of the Inner Ear in Mice Homozygous for Shaker-with-Syndactylism
Authors: Deol M. S.
Published: April 2021

A reinvestigation of the effects of the gene sy on the inner ear showed that the first abnormalities appear several days earlier than had hitherto been believed, and that they concern the periotic labyrinth and not the otic labyrinth. An excessive amount of mesenchymal tissue is laid down round certain, if not all, parts of the otic labyrinth in 13-day embryos, and as the periotic labyrinth is derived from this tissue it is abnormal. The abnormalities of the otic labyrinth appear at least 4 days later, and consist in a dedifferentiation of the epithelium and its penetration by the periotic mesenchymal tissue. As the otic labyrinth is surrounded by the periotic labyrinth, it is believed that the abnormalities of the former are consequent upon those of the latter, although the underlying mechanism is not understood. A possible connection has been suggested between the three major effects of the gene sy, namely, abnormalities of the inner ear, syndactylism, and defects of ossification involving the whole skeleton.
Constitutive Enzymes of the Chick Embryo. II. Cysteine Desulphydrase
Authors: Solomon J. B.
Published: April 2021

Optimum substrate and co-factor concentrations and pH are given for the assay of cysteine desulphydrase in chick tissues. Endogenous desulphydrase activity (no added co-factor) was very low in the yolk from 4 — 12 days and in the whole chick embryo from 3 — 8 days of incubation, there was little in the chorio-allantoic membrane or in the brain and heart of older embryos. Specific desulphydrase activity in the area opaca of blastoderm increased rapidly with development of the yolk sac and was maximal at 7 — 11 days of incubation. Activity had fallen to half the value by 16 days of incubation. There was twice as much activity in White Leghorn yolk sac homogenates than those of Brown Leghorns. There was no deficiency of co-factor in such homogenates. The desulphydrase of Brown Leghorn livers increased from 7 — 16 days of incubation (per mg. protein) but remained constant during the embryonic development of White Leghorn livers. Livers showed a deficiency of co-factor. Just before hatching the liver desulphydrase of both breeds begins to decrease and in White Leghorns had decreased to adult level by 1 week after hatching; whereas in Brown Leghorns the rate of decrease was slower. Liver desulphydrase was increased when adult cockerels were fed on a hard-boiled egg diet. Attempts at enzyme induction by injection of cysteine were generally unsuccessful. When cysteine (16 mg.) was injected at 7 and 10 days of incubation liver desulphydrase had risen by 60 and 100 per cent, respectively 1 day later. Multiple injection of cysteine into young chickens produced only a 40 per cent, increase in liver desulphydrase. The paucity of evidence in favour of substrate-induced enzyme inductions in embryos is discussed.
Expression of TGF-β isoforms during first trimester human embryogenesis
Authors: Gatherer Derek , Duke Peter Ten , Baird David T. , Akhurst Rosemary J.
Published: April 2021

We have studied the expression of the genes encoding transforming growth factors (TGFs) β1, β2 and β3 in human embryos ranging from 32 to 57 days post-coitum, using in situ hybridization. The spatial and temporal pattern of expression of each gene is distinct, though each occasionally overlaps. TGF-β1 is expressed in haematopoietic, endothelial and osteogenic tissues. TGF-β1 and TGF-β3 are expressed in a wide variety of mesenchymal tissues including areas of chondrogenic activity. TGF-β2 is also found in several epithelia and in the ventral nervous system. The differential transcript distributions are broadly similar to those seen in mouse embryos suggesting that there is conservation of TGF-β3 gene regulatory sequences and developmental function across this species boundary.
Dose-dependent regulation of pair-rule stripes by gap proteins and the initiation of segment polarity
Authors: Warrior Rahul , Levine Michael
Published: April 2021

A key step in Drosophila segmentation is the establishment of periodic patterns of pair-rule gene expression in response to gap gene products. From an examination of the distribution of gap and pair-rule proteins in various mutants, we conclude that the on/off periodicity of pairrule stripes depends on both the exact concentrations and combinations of gap proteins expressed in different embryonic cells.

It has been suggested that the distribution of gap gene products depends on cross-regulatory interactions among these genes. Here we provide evidence that autoregulation also plays an important role in this process since there is a reduction in the levels of Kruppel (Kr) RNA and protein in a Kr null mutant.

Once initiated by the gap genes each pair-rule stripe is bell shaped and has ill-defined margins. By the end of the fourteenth nuclear division cycle, the stripes of the pairrule gene even-skipped (eve) sharpen and polarize, a process that is essential for the precisely localized expression of segment polarity genes. This sharpening process appears to depend on a threshold response of the eve promoter to the combinatorial action of eve and a second pair-rule gene hairy. The eve and hairy expression patterns overlap but are out of register and the cells of maximal overlap form the anterior margin of the polarized eve stripe. We propose that the relative placement of the eve and hairy stripes may be an important factor in the initiation of segment polarity.
Re-establishment of the interphase microtubule array in vacuolated plant cells, studied by confocal microscopy and 3-D imaging
Authors: Flanders David J. , Rawlins David J. , Shaw Peter J. , Lloyd Clive W.
Published: April 2021

There are two conflicting ideas about the site of reassembly of cortical microtubules following cytokinesis. Some observations indicate that microtubules (MTs) radiate from the surface of the postcytokinetic nuclear envelope, before becoming organized at the cortex. On the other hand, results of regrowth experiments, following MT depolymerization by drugs, suggest that the array may assemble directly upon the cortex.

In this study, we have taken advantage of the significant separation between nucleus and cortex, in large, vacuolated epidermal cells, to determine which of these two potential sites supports the earliest stages of regrowth in the undrugged, native state.

MTs in stem epidermis of Datura stramonium L. were stained by indirect immunofluorescence. This was performed on hand-cut sections of tissue in which the cells were not separated by enzymes or distorted by airdrying. Epidermal cells with these sheets were optically sectioned by confocal laser scanning microscopy and three-dimensional images reconstructed, rotated and viewed stereoscopically using computer methods.

During metaphase, no MTs can be detected at the cortex but MTs begin to re-colonize the cell surface during early cytokinesis. Thick cables of MTs radiate from the nucleus parallel to the cell plate as well as in other directions, along transvacuolar strands, out to the cortex. Microtubules grow out over the cortex where the thick bundles make contact, as well as from the edges of the fully developed phragmoplast. These early cortical MTs do not form regular transverse arrays: they either appear to be random or to grow in branching V-shaped patterns. The cortical array is therefore not organized immediately but at a later stage.

It is concluded that MT bundles, radiating from the nucleus, are involved in the earliest stages of cortical array formation.
Polysialic acid and N-CAM localisation in embryonic rat kidney: mesenchymal and epithelial elements show different patterns of expression
Authors: Lackie Peter M. , Zuber Christian , Roth Jürgen
Published: April 2021

The expression of polysialic acid (PSA) and the neural cell adhesion molecule (N-CAM) during the embryonic development of rat kidney was investigated using immunocytochemistry and immunoblotting. A monoclonal antibody (mAb 735), which recognised only long chain PSA, and polyclonal antibodies specific for N-CAM were employed.

At the earliest stages of metanephros formation, PSA and N-CAM immunostaining was found in both embryonic anlagen, namely the uretic bud and the metanephrogenic mesenchyme. Reactivity in uretic bud derivatives declined during embryonic development and was generally absent in the collecting duct system by embryonic day 18 (E18). Uninduced metanephrogenic mesenchyme was immunostained throughout development while induced mesenchymal cells showed greatly increased PSA and N-CAM immunoreactivity during their transformation into epithelium. This staining declined rapidly as nephrons differentiated. These processes were preceded by sorting of PSA and N-CAM to the basolateral plasma membrane. Similar N-CAM and PSA patterns were observed in mesonephros development. In adult kidney parenchyma both PSA and N-CAM were undetectable.

Immunoblotting of samples of embryonic kidney with mAb 735 revealed a broad band ranging from 140 to greater than 200×103Mr. N-CAM antibodies revealed reactivity in a band of 140×103Mr after removal of PSA by endoneuraminidase treatment.

Expression of N-CAM and PSA in both embryonic anlagen indicates that neither molecular species acts primarily as an inductive signal. These molecules were localised in areas where changes in cell adhesion during organogenesis might be important and thus may be involved in the grouping of developing cells.
An extra maternally derived X chromosome is deleterious to early mouse development
Authors: Shao Changshun , Takagi Nobuo
Published: April 2021

An extra copy of the X chromosome, unlike autosomes, exerts only minor effects on development in mammals including man and mice, because all X chromosomes except one are genetically inactivated. Contrary to this contention, we found that an additional maternally derived X (XM) chromosome, but probably not a paternally derived one (Xp), consistently contributes to early death of 41,XXY and 41,XXX embryos in mice. Because of imprinted resistance to inactivation, two doses of XM remain active in the trophectoderm, and seem to be responsible for the failure in the development of the ectoplacental cone and extraembryonic ectoderm, and hence, from early embryonic death. Discordant observations in man indicating viability of XMXMXP and XMXMY individuals suggest that imprinting on the human X chromosome is either weak, unstable or erased before the initiation of X-inactivation in progenitors of extraembryonic membranes.
Evidence for allelism of the recessive insertional mutation add and the dominant mouse mutation extra-toes (Xt)
Authors: Pohl Thomas M. , Mattei Marie-Geneviève , Rüther Ulrich
Published: April 2021

A recessive mutant caused by insertional mutagenesis in transgenic mice has been detected in which the anterior part of the forelimb is disorganized. The morphology of the thumb is always altered and sometimes the adjacent finger has an extra phalanx. This phenotype suggests that a body plan gene is affected. We have named the mutation add (anterior digit-pattern deformity). Using the cloned DNA from the flanking region of the integrated transgene, add has been mapped close to the centromere of chromosome 13. This position links add to a genetically mapped locus called extra-toes (Xt). The phenotype of the double-mutant addjXt as well as the molecular analysis suggest that add and Xt are allelic.
Identification of vimentin and novel vimentin-related proteins in Xenopus oocytes and early embryos
Authors: Torpey N. P. , Heasman J. , Wylie C. C.
Published: April 2021

We have made antibodies against fusion proteins of Xenopus vimentin. We show for the first time the distribution of vimentin in larval stages, where it is found in cells of mesenchymal origin, and in radial glial cells. In sections of Xenopus oocytes and early embryos, immunocytochemistry reveals the presence of an extensive cytoplasmic network, distributed in an animal-vegetal gradient. Germ plasm stains particularly strongly. The form of the IF proteins in this network is unusual. In immunoblot experiments the anti-vimentin antibodies detect a number of distinct proteins. We have identified those that are the products of the two known vimentin genes, by injection of synthetic mRNA transcribed from cloned vimentin cDNAs into oocytes, followed by two-dimensional Western blotting. This has demonstrated unambiguously that one Xenopus vimentin, Vim1, is present in oocytes and early embryos. However, two other immunoreactive proteins detected in Triton extracts of oocytes and early embryos are not the products of Vim1, since depletion of vimentin mRNA by antisense oligonucleltide injection has no effect on the synthesis of these proteins. These results suggest that novel IF-like proteins are expressed in Xenopus oocytes and early embryos.
Decoding positional information: regulation of the pair-rule gene hairy
Authors: Howard Kenneth R. , Struhl Gary
Published: April 2021

In the series of local gene activations that occur during early Drosophila development, the striped expression patterns of the pair-rule genes provide the first indication of segmental periodicity. The experiments that we report here address the question of how these patterns arise, by studying the regulation of one of these genes, hairy. We show that each of the seven stripes of hairy expression is controlled by a distinct subset of cis-acting regulatory elements, some mediating transcriptional activation and others transcriptional repression. In general, elements necessary and sufficient for triggering a particular stripe response are clustered on the DNA and appear to overlap or be interspersed with elements involved in at least one other stripe response. Our results extend previous findings suggesting that periodic hairy expression arises by a decoding process in which each stripe is triggered by particular combinations or concentrations of regulatory factors. These regulatory factors are likely to include the products of the gap class of segmentation genes that are required for activating or positioning particular subsets of hairy stripes and are expressed with overlapping distributions during early embryogenesis.
Drosophila neurotactin, a surface glycoprotein with homology to serine esterases, is dynamically expressed during embryogenesis
Authors: Hortsch Michael , Patel Nipam H. , Bieber Allan J. , Traquina Zaida R. , Goodman Corey S.
Published: April 2021

Drosophila neurotactin is a transmembrane glycoprotein with an apparent molecular mass of 135 x 103 Neurotac tin is regionally expressed at the cellular blastoderm stage; later in embryogenesis the expression of the protein becomes restricted to cells of the peripheral and central nervous system. Immunocytochemical localiz ation shows neurotactin protein at points of cell-cell contact. Using the anti-neurotactin monoclonal antibody BP-106, a neurotactin cDNA was isolated that encodes a 846 residue polypeptide. The chromosomal location of the neurotactin gene is 73C. The extracellular domain at the carboxyterminal end of the neurotactin protein shows a strong structural and sequence homology to serine esterases without retaining the amino acids forming the active center. Neurotactin therefore belongs to a growing group of proteins including Drosophila glutactin and thyroglobulins that are known to share this serine esterase protein domain motif without retaining the active center of the enzyme.
Carbonic anhydrase III, an early mesodermal marker, is expressed in embryonic mouse skeletal muscle and notochord
Authors: Lyons Gary E. , Buckingham Margaret E. , Tweedie Susan , Edwards Yvonne H.
Published: April 2021

Carbonic anhydrase HI (CAHI) is an abundant soluble protein in adult mammalian slow twitch skeletal muscle fibers. It is thought to be an early marker for myogenesis based upon its high level of expression in myoblasts in vitro prior to fusion. Using in situ hybridization, we have studied the in vivo distribution of CAIH gene transcripts in mouse embryos and fetuses from 7.25 days to 17.5 days post coitum (p.c.). CAIH mRNAs are first detected in the myotomes of somites between 9.5 and 10.5 days p.c. (20–30 somites). At 15.5 daysp.c., CAIH begins to be restricted to developing slow muscle fibers. By two weeks post partum (p.p.), CAIH mRNAs are detected mainly in slow muscle fibers.

CAHI transcripts are detected at an earlier stage (7.25 days p.c.) in the developing notochord. CAIH is expressed at a much higher level in the notochord than it is in developing skeletal muscle. As the notochord forms the nucleus pulposus in fetal mice, CAIH mRNA levels remain very high. Expression of CAIH in the notochord is of interest in the context of skeletal myogenesis because the notochord is thought to play an important role in somite formation. In addition to the notochord, CAIH transcripts are detected prenatally in several other non-muscle tissues: in cells of the choroid plexus, endocardial cushion and ureter, and in adipocytes.
Endodermal origin of yolk-sac-derived teratomas
Authors: Sobis H. , Verstuyf A. , Vandeputte M.
Published: April 2021

Mutant mice deficient in glucose-6-phosphate dehydrogenase were used to induce teratomas. This enzyme is linked to the X chromosome, which can be inactivated in female embryo. The differences in the enzyme activity between yolk sac mesoderm and embryo versus yolk sac endoderm can be detected in female concepti by using appropriate crosses of wild-type and G6PD-deficient mice. Histochemical study showed that the dual cell population was observed in heterozygous embryos and in the embryomas derived from them. The teratomas derived from the corresponding yolk sac, however, were G6PD-positive from wild-type and G6PD-negative from homozygous enzyme-deficient mothers. We conclude that yolk-sac-derived teratomas are of endodermal origin because of the fact that the paternal X chromosome is inactivated in the yolk sac endoderm, whereas in the yolk sac mesoderm, as in the embryo, the inactivation is at random.
Expression of a novel cadherin (EP-cadherin) in unfertilized eggs and early Xenopus embryos
Authors: Ginsberg Dorit , Desimone Douglas , Geiger Benjamin
Published: April 2021

Two distinct cadherin cDNA clones of Xenopus laevis were isolated from a stage 17 embryo cDNA library. Analysis of the complete deduced amino acid sequences indicated that one of these molecules is closely homologous to chicken and mouse N-cadherin, while the other displays comparable homology to both E- and P-cadherins and was thus denoted EP-cadherin. This molecule has an apparent relative molecular mass of 125 ×103 (compared to approx. 138 ×103 or approx. 140 x103 of E-cadherin and N-cadherins, respectively). Northern and Western blot analyses indicated that N-cadherin is first expressed at the neurula stage while EP-cadherin is the only cadherin detected in unfertilized eggs and cleavage stage embryos. Immunolabeling of Xenopus eggs with antibodies prepared against a fusion protein, containing a segment of EP-cadherin, indicated that the protein is highly enriched at the periphery of the animal hemisphere. EP-cadherin was also found in A6 epithelial cells derived from Xenopus kidneys, and was apparently localized in the intercellular adherens junctions.
Involvement of Bone Morphogenetic Protein-4 (BMP-4) and Vgr-1 in morphogenesis and neurogenesis in the mouse
Authors: Jones C. Michael , Lyons Karen M. , Hogan Brigid L. M.
Published: April 2021

Bone Morphogenetic Protein-4 (BMP-4) and Vgr-1 are members of the TGF-/J gene family most closely related to the Drosophila Decapentaplegic and Xenopus Vg-1 genes. Members of this gene family have been implicated in diverse processes during embryogenesis including epithelial-mesenchymal interactions. Here, we use in situ hybridization to localize BMP-4 and Vgr-1 transcripts during murine development. BMP-4 mRNA is found in a variety of tissues. In the 8.5 days p.c. embryo, transcripts are localized to the mesoderm posterior to the last somite. Later gestation embryos show expression in developing limbs, the embryonic heart, the facial processes and condensed mesenchymal cells associated with early whisker follicle formation. In the developing central nervous system (CNS), BMP-4 expression is restricted to the floor of the diencephalon associated with pituitary development. In contrast, Vgr-1 transcripts are found along the anteroposterior axis of the CNS, in cells immediately adjacent to the floor plate and in the roof plate extending to the forebrain. Together, the data support the hypothesis that polypeptide growth factors of the TGF-β superfamily play key roles in the initial stages of neurogenesis and organogenesis during murine development.
Primitive erythropoiesis of mouse teratocarcinoma stem cells PCC3/A/1 in serum-free medium
Authors: Miwa Yoko , Atsumi Tadao , Imai Nobuo , Ikawa Yoji
Published: April 2021

Mouse teratocarcinoma stem cells PCC3/A/1 differentiated into various types of cells, such as red cells, when they were grown in serum-free medium containing transferrin and bovine serum albumin on a KCF cell feeder layer. These red cells were stained well with 2,7-diaminofluorene (DAF), and therefore were erythroid cells. They were nucleated and contained embryonic globin chains, immunologically identified with anti-embryonic hemoglobin antisera after acid urea Triton X-100 polyacrylamide gel electrophoresis (UT-PAGE). The addition of erythropoietin to the culture medium enhanced the production of both embryonic and adult globin chains. The addition of interleukin-3 also enhanced the production of embryonic globin chains, but not the production of adult globin chains. These results indicated that primitive erythropoiesis of PCC3/A/1 teratocarcinoma cells did not require exogenous addition of any hematopoietic factor such as erythropoietin or interleukin-3. This culture system will be a new model system for investigating the factors regulating the primitive erythropoiesis in yolk sac blood islands.
Spatial and temporal changes in the distribution of proteoglycans during avian neural crest development
Authors: Perris Roberto , Krotoski Danuta , Lallier Thomas , Domingo Carmen , Sorrell J. Michael , Bronner-Fraser Marianne
Published: April 2021

In this study, we describe the distribution of various classes of proteoglycans and their potential matrix ligand, hyaluronan, during neural crest development in the trunk region of the chicken embryo. Different types of chondroitin and keratan sulfate proteoglycans were recognized using a panel of monoclonal antibodies produced against specific epitopes on their glycosamino-glycan chains. A heparan sulfate proteoglycan was identified by an antibody against its core protein. The distribution of hyaluronan was mapped using a biotinyl-ated fragment that corresponds to the hyaluronan-binding region of cartilage proteoglycans. Four major patterns of proteoglycan immunoreactivity were ob-served. (1) Chondroitin-6-sulfate-rich proteoglycans and certain keratan sulfate proteoglycans were absent from regions containing migrating neural crest cells, but were present in interstitial matrices and basement membranes along prospective migratory pathways such as the ventral portion of the sclerotome. Although initially distributed uniformly along the rostrocaudal extent of the sclerotome, these proteoglycans became rearranged to the caudal portion of the sclerotome with progressive migration of neural crest cells through the rostral sclerotome and their aggregation into peripheral ganglia. (2) A subset of chondroitin/keratan sulfate proteoglycans bearing primarily unsulfated chondroitin chains was observed exclusively in regions where neural crest cells were absent or delayed from entering, such as the perinotochordal and subepidermal spaces. (3) A subset of chondroitin/keratan sulfate proteoglycans was restricted to the perinotochordal region and, following gangliogenesis, was arranged in a metameric pattern corresponding to the sites where presumptive vertebral arches form. (4) Certain keratan sulfate proteoglycans and a heparan sulfate proteoglycan were observed in basement membranes and in an interstitial matrix uniformly distributed along the rostrocaudal extent of the sclerotome. After gangliogenesis, the neural crest-derived dorsal root and sympathetic ganglia contained both these proteoglycan types, but were essentially free of other chondroitin/keratan-proteoglycan subsets. Hyaluronan generally colocalized with the first set of proteoglycans, but also was concentrated around mi-grating neural crest cells and was reduced in neural crest-derived ganglia. These observations demonstrate that proteoglycans have diverse and dynamic distri-butions during times of neural crest development and chondrogenesis of the presumptive vertebrae. In gen-eral, chondroitin/keratan sulfate proteoglycans are abundant in regions where neural crest cells are absent, and their segmental distribution inversely correlates with that of neural crest-derived ganglia.
Expression of a novel Müller glia specific antigen during development and after optic nerve lesion
Authors: Schlosshauer Burkhard , Grauer Dorothee , Dütting Dieter , Vanselow Jens
Published: April 2021

To generate monoclonal antibodies, immunogen fractions were purified from embryonic chick retinae by temperature-induced detergent-phase separation employing Triton X-114. Under reducing conditions, the monoclonal antibody (mAb) 2M6 identifies a protein doublet at 40 and 46×103Mr, which appears to form disulfide-coupled multimers. The 2M6 antigen is regulated developmentally during retinal histogenesis and its expression correlates with Müller glial cell differentiation. Isolated glial endfeet and retinal glial cells in vitro were found to be 2M6-positive, identified with the aid of the general glia marker mAb R5. mAb 2M6 does not bind to any other glial cell type in the CNS as judged from immunohistochemical data. Cell-type specificity was further substantiated by employing retinal explant and single cell cultures on laminin in conjunction with two novel neuron-specific monoclonal antibodies. MAb 2M6 does not bind either to neurites or to neuronal cell bodies. Incubation of retinal cells in vitro with bromodeoxyuridine (BrdU) and subsequent immunodouble labeling with mAb 2M6 and anti-BrdU reveal that mitotic Müller cells can also express the 2M6 antigen.

To investigate whether Müller cell differentiation depends on interactions with earlier differentiating ganglion cells, transections of early embryonic optic nerves in vivo were performed. This operation eliminates ganglion cells. Müller cell development and 2M6 antigen expression were not affected, suggesting a ganglion-cell-independent differentiation process. If, however, the optic nerve of juvenile chicken was crushed to induce a transient degeneration/regeneration process in the retina, a significant increase of 2M6 immunoreactivity became evident. These data are in line with the hypothesis that Müller glial cells, in contrast to other distinct glial cell types, might facilitate neural regeneration.
Gap-junctional permeability in early and cleavage-arrested ascidian embryos
Authors: Dale Brian , Santella Luigi A , Tosti Elisabetta
Published: April 2021

Using the whole-cell voltage clamp technique, we have studied junctional conductance (Gj), and Lucifer Yellow (LY) coupling in 2-cell and 32-cell ascidian embryos. Gj ranges from 17.5 to 35.3 nS in the 2-cell embryo where there is no passage of LY, and from 3.5 to 12.2 nS in the later embryo where LY dye spread is extensive. In both cases, Gj is independent of the transjunctional potential (Vj). Manually apposed 2-cell or 32-cell embryos established a junctional conductance of up to 10 nS within 30 min of contact. Furthermore, since we did not observe any significant number of cytoplasmic bridges at the EM and Gj is sensitive to octanol, it is probable that blastomeres in the 2-cell and 32-cell embryos are in communication by gap junctions. In order to compare Gj in the two stages and to circumvent problems of cell size, movement and spatial location, we used cytochalasin B to arrest cleavage. Gj in cleavage-arrested 2-cell embryos ranged from 25.0 to 38.0 nS and remained constant over a period of 2.5 h. LY injected into a blastomere of these arrested embryos did not spread to the neighbour cell until they attained the developmental age of a 32- to 64-cell control embryo. Our experiments indicate a change in selectivity of gap junctions at the 32cell stage that is not reflected by a macroscopic change in ionic permeability.
Integrin expression during human epidermal development in vivo and in vitro
Authors: Hertle Mark D. , Adams Josephine C. , Watt Fiona M.
Published: April 2021

In order to investigate the role of extracellular matrix receptors of the integrin family in establishing the spatial organization of epidermal keratinocytes, we used immunofluorescence microscopy to examine the expression of a range of integrin subunits during development of human palm and sole skin. All of the integrins expressed during development were also present in mature epidermis and were largely confined to the basal layer of keratinocytes in a pericellular distribution. The α3 and β1 subunits were expressed prior to initiation of stratification and did not change in abundance or distribution during subsequent development, α4 and β3 were not detected at any time in the epidermis. Every other subunit examined showed spatial or temporal changes in expression. Staining for α3 was strong before stratification and until mid-development, but was greatly decreased in neonatal epidermis. α2 was first detected in small patches of basal cells prior to stratification, and thereafter was found in the entire basal layer, with greater staining in developing sweat glands, α5 was not expressed until mid-development, and then primarily in developing sweat glands, with faint expression in neonatal epidermis, αv was detected following stratification, in developing sweat glands, and occasionally in neonatal epidermis, α6 and β4 were peribasally expressed before stratification, but there-after became concentrated at the basal cell surface in contact with the basement membrane, co-localizing with hemidesmosomes as determined by staining with bullous pemphigoid antiserum. We also examined the distri-bution of three known ligands for keratinocyte integrins: laminin and collagen type IV were present in the basement membrane zone at all stages of development, whereas fibronectin was only evident there until about 13 weeks estimated gestational age. Finally, we found that the changes in integrin expression that occur on initiation of stratification in vivo could be reproduced in organ cultures of developing skin; such cultures there-fore provide a useful experimental model for further studies of the role of integrins in epidermal stratification.
Common precursors for neural and mesectodermal derivatives in the cephalic neural crest
Authors: Baroffio Anne , Dupin Elisabeth , Douarin Nicole M. Le
Published: April 2021

The cephalic neural crest (NC) of vertebrate embryos yields a variety of cell types belonging to the neuronal, glial, melanocytic and mesectodermal lineages. Using clonal cultures of quail migrating cephalic NC cells, we demonstrated that neurons and glial cells of the peripheral nervous system can originate from the same progenitors as cartilage, one of the mesectodermal derivatives of the NC. Moreover, we obtained evidence that the migrating cephalic NC contains a few highly multipotent precursors that are common to neurons, glia, cartilage and pigment cells and which we interpreté as representative of a stem cell population. In contrast, other NC cells, although provided with identical culture conditions, give rise to clones composed of only one or some of these cell types. These cells thus appear restricted in their developmental potentialities compared to multipotent cells. It is therefore proposed that, in vivo, the active proliferation of pluripotent NC cells during the migration process generates distinct subpopulations of cells that become progressively committed to different developmental fates.
Gene regulatory factors of the sea urchin embryo: I. Purification by affinity chromatography and cloning of P3A2, a novel DNA-binding protein
Authors: Calzone Frank J. , Höog Christer , Teplow David B. , Cutting Ann E. , Zeller Robert W. , Britten Roy J. , Davidson Eric H.
Published: April 2021

The P3A2 regulatory protein interacts with specific sites in the control region of the Cyllla actin gene. Previous studies showed that this interaction is required to confine expression of a Cyllla. CAT fusion to the aboral ectoderm, the embryonic territory in which Cyllla is normally utilized. P3A2 also binds specifically to similar target sites located in the regulatory region of the SM50 gene, which is expressed only in skeletogenic mesenchyme lineages. The P3A2 factor was purified by affinity chromatography from nuclear extracts of 24 h sea urchin embryos, and partial peptide sequences were used to isolate a cDNA clone encoding the complete protein. There are no significant similarities between P3A2 and any other protein in existing sequence data bases. P3A2 thus includes a novel type of DNA-binding domain. To examine the differential utilization of P3A2 in Cyllla and SM50 genes, we measured the specific affinity of this protein for the various target sites in the regulatory DNAs of each gene, and identified the core target site sequences. The stability of P3A2 complexes formed with SM50 target sites is 50-100 times greater than that of the complexes formed with Cyllla target sites, though the factor binds to very similar core sequence elements. P3A2 is one of at least twelve different proteins whose interaction with CyIIIa regulatory DNA is required for correct developmental expression. The results reported demonstrate that it might be possible to purify most of these regulatory proteins, or any other specific DNA-binding proteins of the sea urchin embryo, by using the simple procedures described for P3A2.
Changes in transcellular ionic currents associated with cytokinesis and polar lobe formation in embryos of Bithynia tentaculata (Mollusca)
Authors: Zivkovic Danica , Dohmen René
Published: April 2021

Using the vibrating probe technique, we have measured transcellular ionic currents ranging from 0.05 to 3.00 μA cm-2 around polar-lobe-forming embryos of Bithynia tentaculata.

During maturation and the first two mitotic division cycles, we have detected recurrent changes in the current pattern, which correlated temporally and spatially with cytokinesis and polar lobe formation. The maximum inward current at the animal pole coincides with the onset of cleavages at that pole, and the maximum inward current at the vegetal pole slightly precedes the formation of the two meiotic polar lobes and the first mitotic polar lobe at that pole. This correlation was less clear with regard to the second mitotic polar lobe. The resorption of the polar lobe correlates with a reversal of the current from inward to outward at the vegetal pole. After the resorption of the first mitotic polar lobe, the transmembrane ion flow in the membrane area, which formerly covered the lobe, is different from the surrounding membrane.
The program for cellular differentiation in Volvox carteri as revealed by molecular analysis of development in a gonidialess/somatic regenerator mutant
Authors: Tam L.W. , Kirk D.L.
Published: April 2021

Development of a ‘gonidialess’/‘somatic regenerator’ double mutant of Volvox carteri was analyzed with a number of cell-type-specific cDNA probes that had been identified in a previous study. Whereas in wild-type strains somatic cells and gonidia (asexual reproductive cells) constitute two distinct cell lineages, in this mutant all cells first differentiate as somatic cells and then redifferentiate as gonidia. During the initial period of somatic differentiation, we found that both gonidial and ‘early’ somatic transcripts were accumulated in the mutant, consistent with the idea that it is the regA gene product (which is defective in this mutant) that normally acts to suppress gonidial gene expression in somatic cells. Later in development, levels of early somatic transcripts fell abruptly, levels of the late somatic transcripts remained extremely low, and levels of gonidial transcripts rose as the cells redifferentiated. Thus it appears that in the mutant cells the gonidial program of development takes over and somatic differentiation is aborted before the stage at which late somatic genes are normally activated. These results provide molecular genetic support for a model which postulates that three types of genes (including the two that are defective in the strain studied here) are crucial for converting the sequential program of differentiation seen in more primitive volvocalean algae to the dichotomous program of germ-soma differentiation that occurs in wild-type V. carteri.
Rodent CNS neuroblasts exhibit both perpendicular and parallel contact guidance on the aligned parallel neurite bundle
Authors: Nagata Isao , Nakatsuji Norio
Published: April 2021

Mouse cerebellar granule cells showed two types of migration behavior in microexplant cultures. They first migrated along their neurites, showing the typical contact guidance, and then oriented themselves at right angles to the parallel neurites, thus exhibiting the ‘perpendicular contact guidance’ (Nakatsuji, N. and Nagata, I. 1989 Development, 106, 441–447).

To study whether other neurons have the capacity to show similar ‘perpendicular contact guidance’, we cultured dissociated neuroblasts from various parts of CNS or PNS on parallel neurite bundles. The PNS neuroblasts always extended their processes parallel to the neurite bundle. In contrast, almost all kinds of CNS neuroblasts tested oriented their processes both perpendicular and parallel to the neurite bundles that were all free of glia. Time-lapse video recording revealed that neuroblasts migrated in both directions. Thus, CNS neuroblasts possess the capacity to migrate and extend their processes at right angles to the substratum of heterotypic neurite bundles, which may play an important role in histogenesis of the CNS during development.