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Nucleotides in embryos in the stages of morula, gastrula, and neurula.

Levels of nucleotides and sugar nucleotides in embryos of Bufo arenarum at the stages of morula, gastrula, and neurula have been measured. The total amounts of purine nucleoside diphosphates decreased from morula to gastrula, but increased sharply from gastrula to neurula. The levels of ADP followed this pattern, but those of GDP did not change significantly through the three stages. Purine nucleoside triphosphate levels, which had increased immediately after fertilization, remained almost constant through morula, gastrula, and neurula. As with the purine nucleoside diphosphates, the adenine nucleotide decreased from morula to gastrula, and increased from gastrula to neurula. In contrast, the level of GTP showed a sharp maximum at gastrula. The total pyrimidine nucleoside triphosphate did not change significantly from morula through neurula. As in previous stages of development, only uridine sugar nucleotides were detected. A sharp increase of the galactosyl ester of nucleotides was found at gastrula.

Adenosine Diphosphate

Extracellular matrix synthesis in blastula and gastrula stages of normal and hybrid frog embryos. I. Toluidine blue and lanthanum staining.

Four developmental stages of Rana pipiens, R. pipiens female X R. catesbeiana female, and R. pipiens female X R. esculenta male embryos were fixed in standard electron microscopic fixatives with or without 1% lanthanum nitrate (LN) added. Embryos fixed without LN were embedded and sectioned at 0-5 micrometer and stained with toluidine blue. Embryos fixed with LN were embedded and sectioned at 70-80 nm and examined in the electron microscope. Both techiques show a dramatic increase in the amount of stainable material in the extracellular matrix during development. Blastula stage embryos have a small amount of material in the extracellular matrix. Early gastrulae have more stainable material than blastulae and this increased amount appears to be due to an accumulation of material in the marginal zone of the embryo, especially near the dorsal lip of the blastopore. Late gastrulae have large amounts of stainable material in the extracellular spaces in almost all parts of the embryo. Interspecific arrested hybrid embryos do not show the same dramatic accumulation of stainable material in their extracellular matrices.

Animals

Intercellular junctions, intramembranous particles and cytoskeletal elements of deep cells of the Fundulus gastrula.

The fine structure of motile deep cells of the gastrula stage of Fundulus heteroclitus was studied with transmission electron microscopy, using both thin sectioning and freeze-cleave techniques. Gastrula deep cells form extensive non-junctiona appositions with each other, in which the apposed plasma membranes are parallel and separated by a distance of 26-28 nm. They also form gap junctions. Tight junctions, desmosomes, and extensive interdigitations of apposed plasma membranes were not observed. The plasma membranes of deep cells contain numerous unclustered intramembranous particles. Cytoplasmic microtubules were found, but they appear to be small in number, sparsely distributed, and mainly randomly oriented. Microfilaments are also present and are localized largely in the cortical cytoplasm and in thin cell extensions. The significance of these findings for the contact and locomotory behavior of deep cells is discussed.

Animals

Non-polyadenylated 22 s ribonucleoprotein particle is insensitive to translational inhibitor RNA of cryptobiotic gastrulae of Artemia salina.

A free cytoplasmic 22 S ribonucleoprotein particle exhibiting a major template activity in rabbit reticulocyte system has been identified in the cryptobiotic gastrulae of Artemia salina. This particle contains non-polyadenylated 9 S messenger RNA which codes primarily for a non-histone basic protein with an apparent molecular weight of 26 000 daltons. We have previously demonstrated the presence of a translational inhibitor RNA which is apparently responsible for transforming polyadenylated messenger (Slegers et al., FEBS Letters 80, 390-394, 1977). This inhibitor RNA was found to be completely ineffective on the template activity of non-polyadenylated 22 S messenger ribonucleoprotein, confirming the specificity of this regulatory RNA for polyadenylate sequences.

Animals

Extracellular matrix synthesis in blastula and gastrula stages of normal and hybrid frog embryos. IV. Biochemical and autoradiographic observations on fucose-, glucose-, and mannose-labelled materials.

Normal Rana pipiens gastrulae show more incorporation of isotopically labelled fucose, glucose, and mannose into TCA-insoluble materials than blastulae. Interspecific hybrid embryos which undergo developmental arrest at the onset of gastrulation often synthesize reduced amounts of fucose-, glucose- and mannose-labelled materials. These materials are high-molecular weight and are degraded by Pronase into fragments which are included in Sepharose CL-4B but excluded from Sephadex G-50. Labelled materials contain label predominantly in fucose and galactose with little label in glucose or mannose. Labelled materials migrate slowly on cellulose acetate, bind to DEAE-cellulose and elute at low ionic strength and are precipitated by cetyl pyridinium chloride (CPC) without the addition of carrier compounds. Pulse-chase labelling experiments and light- and electron-microscopic autoradiography were used to examine the site of synthesis, mode of transport, and sites of deposition of fucose-, glucose- and mannose-labelled materials in different developmental stages of normal developing Rana pipiens embryos and interspecific hybrid embryos formed by fertilizing the eggs of R. pipiens with the sperm of R. catesbeiana. In both normal and hybrid embryos, after a 15-30-min pulse, grains are closely associated with juxtanuclear and cytoplasmic collections of membrane-bound vesicles which resemble the Golgi apparatus. In normal embryos following a 15-30-min pulse and a 60-min chase, grains are largely cleared from the cytoplasmic vesicles and deposited in the extracellular spaces or along cell surfaces. In contrast, arrested hybrid embryos given a 15-30-min pulse and a 60-min chase show a marked accumulation of grains over cytoplasmic structures such as the Golgi apparatus and vesicular elements in the cell cortex. Certain interesting features of regional variation in synthetic activity in developing normal embryos are also described.

Animals

Embryo surface morphology during post-gastrula development of the frog, Bombina orientalis, as revealed by scanning electron microscopy.

To further study the morphology of the developing amphibian embryo, and to provide a basis for the characterization of abnormal embryogenesis, surface features of the Bombina orientalis embryo during post-gastrula development were examined with the scanning electron microscope. Ciliated cells similar to those reported for Rama pipiens and Ambystoma mexicanum were observed in the epidermis. Epidermal secretory cells displaying large numbers of exocytotis apertures become prominent at the early tail bud stage and are prevalent through hatching. The surface features of the adhesive organs and gill filaments are also described. Removal of the embryo from the fertilization envelope resulted in the accumulation of a prominent mucoid matrix over the surface cells. These findings should be considered when experiments are designed which involve the effects of applied molecules on amphibian embryo epidermal cells.

Animals

Archenteron cells are responsible for the increase in ribosomal RNA synthesis in sea urchin gastrulae.

Paracentrotus lividus embryos were continuously labeled with P32 from hatching blastula to pluteus. The archenteron cells were then separated from the rest of the embryo and the radioactivity accumulated into the ribosomal RNA of the two cell groups measured. The results clearly indicate that the bulk of ribosomal RNA is mainly if not entirely, synthesized in this time interval by the archenteron cells.

Animals

Maternal redd1 mRNA decline triggers mTORC1 activation during the blastula-gastrula transition in zebrafish embryos.

During early metazoan development, maternal mRNAs and proteins stored in the egg sustain initial cellular functions. After the blastula stage, developmental control shifts to zygotic gene expression, and maternal transcripts are progressively degraded. Although mTORC1 is a central regulator of global mRNA translation and cell growth, its role in controlling maternal mRNA translation prior to gastrulation remains poorly understood. In zebrafish embryos, the mTORC1 inhibitor redd1 is abundantly expressed after fertilization but decreases following the maternal-to-zygotic transition (MZT), inversely correlating with mTORC1 activity. Overexpression of redd1 suppresses mTORC1, impairs gastrulation, and reduces translation of 5'TOP mRNAs and key regulatory genes, underscoring the necessity of relieving mTORC1 inhibition after the blastula stage. To investigate redd1 translation under conditions of low mTORC1 activity, we injected reporter mRNAs containing its 5' and 3' UTRs. The 3'UTR promoted polyadenylation and enhanced translation, while both UTRs enabled efficient reporter expression despite mTORC1 suppression, indicating that redd1 mRNA is translated independently of canonical mTORC1 pathways. Similarly, maternal mRNAs such as nanog, myca, pou5f3, and ccnb1, as well as the early zygotic transcript dharma, are translated through mTORC1-independent mechanisms. Together, these findings reveal a transient phase of mTORC1 suppression in early zebrafish embryos and demonstrate that select maternal and zygotic mRNAs bypass this regulation to ensure proper developmental progression.

Animals

Characterization of protein components of poly(A)-containing messenger ribonucleoproteins from cryptobiotic gastrulae of Artemia salina.

Conditions for the isolation of intact poly(A)+mRNP from cryptobiotic gastruale of A. salina are described. In the presence of Mg2+ ions nucleolytic cleavage occurs in vitro in the vicinity of the 3'-poly(A) segment of mRNP during the isolation procedure. The resulting two parts of poly(A)+mRNP complex are separated by thermal elution from oligo(dT)-cellulsoe affinity column. Analysis by SDS-gel electrophoresis of protein components associated with intact poly(A)+mRNP has revealed the existence of 20--30 S RNP complex containing five major proteins with Mr 68,000, 53,000, 50,000, 45,000 and 38,000, respectively, but completely lacking the poly(A)-specific Mr 76,000 protein.

Animals

Somitogenesis in amphibia. II. Origins in early embryogenesis of two factors involved in somite specification.

A somite pre-pattern is established shortly before visible segmentation. The pre-pattern results from the interaction of two components: a wave of cell behavioural change that passes along the axis, and, an underlying co-ordination of the cells that is the basis for their association into large somite-sized groupings. The evidence is derived from studies of the zones of abnormal segmentation that follow temperature shocks delivered between the neurula and tail-bud stages (Pearson & Elsdale, 1979). Temperature shock given earlier at the mid-gastrula stage is however ineffective in inducing abnormalities in somitogenesis. Shocks given before the mid-gastrula stage reveal a prior period of sensitivity stretching back into the blastula. Thus early and late sensitive periods can be defined separated by a short refactory period. Quite different patterns in the distribution of somite abnormalities characterize the results of shock during the two sensitive periods, suggesting different aetiologies. It is concluded that the wave of rapid cell change is set up early in embryogenesis during the blastula stage, and each cell of the prospective paraxial mesoderm carries a determination to change after a specific length of time, i.e. a countdown is set in each cell. As a result of the movements of gastrulation, the prospective paraxial mesoderm cells become laid out along the axis of the neurula in the order (antero-posterior sequence) in which they will change. The achievement of the correct redistribution of the cells depends crucially on the conservation of the sequence in the blastula by the maintenance of topological integrity throughout gastrulation. It is suggested that early shock disturbs gastrulation movements, causing some mixing up of the cells resulting in incoherence of the wavefront. Whereas early shocks are thus assumed to affect the wave, the evidence suggests that late shock undergoes co-ordination. It is concluded therefore that co-ordination is established later, after the refractory period, around the late gastrula stage.

Animals

Distinct single-copy sequence sets in sea urchin nuclear RNAs.

The purpose of this study was to determine whether nuclear RNAs (nRNA) of sea urchin embryos and adult tissues contain identical or partially distinct sets of single-copy sequence transcripts. A DNA tracer was prepared consisting mainly of sequences absent from gastrula nRNA; 3.6% of this tracer reacted with adult intestine nRNA but not with gastrula nRNA. The existence of a differentially transcribed DNA fraction was verified by its partial purification and rehybridization to intestine and gastrula nRNAs. About one-third of the genomic single-copy sequence is represented in both nRNAs, or about 2 x 10(8) nucleotides. The differentially transcribed portion of the single-copy genome identified in this work includes about 3.5 x 10(7) nucleotides.

Animals

[Activity of carbohydrate metabolism enzymes in loach embryos under the influence of hormones].

The activities of hexokinase, glucokinase, phosphofructokinase, glucose-6-phosphate dehydrogenase, glucose-6-phosphatase, and fructose-1,6-diphosphatase were determined in loach embryos developed in solutions of insulin, hydrocortisone, estrone and thyroxin at different stages of embryogenesis. Glucokinase and fructose-1,6-diphosphatase activties are shown not to change markedly under the influence of the above-mentioned hormones. During some periods of early development the hexokinase activity is inhibited by insulin, estrone and thyroxin. The glucose-6-phosphate dehydrogenase activity is suppressed by each of the used hormones at all the stages of early embryogenesis while the glocose-6-phosphatase activity decreased only under the influence of insulin at the cleavage, blastula and gastrula stages. Insulin increased the activity of phosphofructokinase at the cleavage, blastula and early gastrula stages and hydrocortisone, estrone and thyroxine during certain periods of these stages. From middle gastrula two last hormones decreased the phosphofructokinase activity in the loach embryos.

Animals