bluuebunny/crossref_metadata_2025 huggingface dataset:
Differential expression of Hox 3.1 protein in subregions of the embryonic and adult spinal cord
Authors: Awgulewitsch Alexander , Jacobs Donna
Published: April 2021

Synthetic oligopeptides derived from the predicted Hox 3.1 protein coding sequence were used for the production of antibodies (anti-aa2) that specifically recognize Hox 3.1 protein in tissue sections. These antibodies were applied in immunohistochemical studies to monitor the expression of Hox 3.1 protein within the central nervous system (CNS) of embryonic and adult mice. We demonstrate congruency between the distinct Hox 3.1 RNA and protein expression patterns in the developing spinal cord by direct comparison of in situ hybridization and immunohistochemical staining in frozen sagittal sections from embryos of 12.5 days of gestation. A distinct pattern of spatially restricted expression of Hox 3.1 protein within the spinal cord was first detected at around 10.5 days of embryonic development. Within certain anteroposterior limits the geometries of this expression pattern change drastically during subsequent embryonic stages, concomitant with important cytoarchitectural changes in the developing spinal cord. Analyses on subcellular levels indicate predominant accumulation of Hox 3.1 protein within nuclei of neuronal cells. In addition to the nuclear localization in subsets of embryonic cells, persistent accumulation of Hox 3.1 protein was shown in nuclei of fully differentiated and mature neuronal cells of the adult CNS.
Genome imprinting and development in the mouse
Authors: Surani M. Azim , Kothary Rashmi , Allen Nicholas D. , Singh Prim B. , Fundele Reinald , Ferguson-Smith Anne C. , Barton Sheila C.
Published: April 2021

Development in mammals is influenced by genome imprinting which results in differences in the expression of some homologous maternal and paternal alleles. This process, initiated in the germline, can continue following fertilization with interactions between oocyte cytoplasmic factors and the parental genomes involving modifier genes. Further epigenetic modifications may follow to render the ‘imprints’ heritable through subsequent cell divisions during development. Imprinting of genes can be critical for their dosage affecting embryonic growth, cell proliferation and differentiation.

The cumulative effects of all the imprinted genes are observed in androgenones (AG) and parthenogenones (PG), which reveal complementary phenotypes with respect to embryonic and extraembryonic tissues. The presence of PG cells in chimeras causes growth retardation, while that of AG cells enhanced growth. AG cells apparently have a higher cell proliferation rate and, unlike PG cells, are less prone to selective elimination. However, the PG germ cells are exempt from cell selection. In chimeras, PG cells are more likely to be found in ectodermal derivatives such as epidermis and brain in contrast to AG cells which make pronounced contributions to many mesodermal derivatives such as muscle, kidney, dermis and skeleton. The presence of androgenetic cells in chimeras also results in the disproportionate elongation of the anterior-posterior axis and sometimes in the abnormal development of skeletal elements along the axis. Genetic studies highlight the influence of subsets of imprinted genes, and identify those that are critical for development.
Imprinting by DNA methylation: from transgenes to endogenous gene sequences
Authors: Reik Wolf , Howlett Sarah K. , Surani M. Azim
Published: April 2021

A number of transgenes in the mouse show variation in methylation and expression phenotypes dependent on parental transmission. It appears that there exist at least two types of transgene imprinting; one is retained on an essentially homozygous background, while the other requires heterozygosity at some modifying loci in the genome and is observed as differences in phenotype in reciprocal crosses. For this type of imprinting to occur, the parental origin of the modifier locus itself is important, and parental asymmetry may involve specific interactions between egg cytoplasm and the chromosomes. Based on the identification of ‘methylation polymorphism’ in the mouse genome, we also show that endogenous gene sequences can undergo imprinting by DNA methylation.
Expression of the IGF-ll/mannose-6-phosphate receptor mRNA and protein in the developing rat
Authors: Senior Paul V. , Byrne Simon , Brammar William J. , Beck Felix
Published: April 2021

The insulin-like growth factor II (IGF-II) receptor is identical to the mannose-6-phosphate receptor (M-6-P), but its role as a somatomedin transducer is uncertain. IGF-II/M-6-P receptor expression was studied by in situ hybridization (ISH) in the developing rat. Expression occurs in extra-embryonic membranes at the time of IGF-n mRNA induction and later at paracrine/ autocrine sites of IGF-II action (skeletal muscle and perichondrium) in the embryo. Highest levels of receptor mRNA occur in heart and major vessels. Postnatally transcription is strongly down-regulated. This suggests a role for the IGF-II/M-6-P receptor in IGF-II action or turnover during development distinct from its role in lysosomal transport.
Effects of ectopic expression of caudal during Drosophila development
Authors: Mlodzik Marek , Gibson Greg , Gehring Walter J.
Published: April 2021

The effects of heat-shock-induced ectopic expression of the homeobox gene caudal (cad) at all stages of Drosophila development have been examined. Presence of cad protein (CAD) at the anterior end of cellular blastoderm embryos was found to disrupt head development and segmentation, due to alteration of the expression of segmentation genes such as fushi taraza and engrailed, as well as repression of head-determining genes such as Deformed. These results support the conclusion that, while CAD is probably required to activate transcription of fushi tarazu in the posterior half of the embryo, it should not be expressed in the anterior half prior to gastrulation, and thus suggest a role for the CAD gradient. Ectopic expression of CAD at later stages of development has no obvious effects on embryogenesis or imaginai disc development, suggesting that the homeotic genes of the Antennapedia and Bithorax Complexes are almost completely epistatic to caudal.
Structure and expression of the mouse Oct2a and Oct2b, two differentially spliced products of the same gene
Authors: Hatzopoulos Antonis K. , Stoykova Anastasia S. , R. Erselius Jörn , Goulding Martyn , Neuman Toomas , Gruss Peter
Published: April 2021

A large family of tissue-specific nuclear proteins interact with the octamer motif ATTTGCAT, a transcriptional regulatory element found in the promoter and enhancer sequences of many genes. As a step towards elucidating the mechanism of this regulation, cDNA clones of the mouse Oct2 protein were isolated. One, called here Oct2b, encodes a larger variant of the previously described Oct2a proteins. The Oct2b cDNA has an insertion of 74 bp close to the 3 ′ end which creates an open reading frame distinct from Oct2a. As a result, the Oct2b protein has a carboxy end which is similar to that of the ubiquitous octamer-binding protein Oct1. Analysis of the Oct2 gene shows that Oct2a and Oct2b are differentially spliced products of the same gene. The insertion in the Oct2b cDNA results from the inclusion of an additional exon in the mRNA which would otherwise reside in an intron sequence of the Oct2a transcript.

RNA analysis demonstrates that both Oct2a and 2b mRNAs are most abundant in B-cells but they are also expressed in a variety of tissues including brain, intestine, testis, kidney, as well as in embryos. Interestingly, the ratio of Oct2a and 2b varies among tissues. In situ hybridization studies during mouse embryogenesis show that the Oct2 gene is widely expressed in the developing nervous system. In contrast, expression in the adult brain is confined to very specific areas which include the suprachiasmatic and medial mammillary nuclei, hippocampus, olfactory tract and the olfactory bulb. Oct2 proteins are present in both neuronal and oligodendroglial cells, although they are more abundant in glial cells.
Stimulatory effects of basic fibroblast growth factor on DNA synthesis in the human embryonic cornea
Authors: Hyldahl Louise , Schofield Paul N. , Engström Wilhelm
Published: April 2021

First trimester human embryonic eye globes were micro dissected so that a passage was opened between the outer environment and the anterior chamber, which rendered free access of tissue culture medium to the endothelial cell monolayer. The dissected eye globes were main tained in organ culture for 24 h in the continuous presence of tritiated thymidine. Sections were cut through the whole eye globes and were subject to autoradiographic analysis in order to estimate the mitogenic response of human corneal endothelial cells to externally supplied growth factors and hormones.

It was found that the corneal endothelial cells could be stimulated to initiate DNA synthesis by exposure to basic fibroblast growth factor (bFGF). The thymidine label ling index nearly doubled after bFGF addition.

Northern blot analysis revealed the presence of bFGF transcripts in the embryonic eye. In contrast we were unable to trace any bFGF transcripts in other first trimester human embryonic organs. In an attempt to determine the topographical distribution of bFGF mRNA within the eye, we found that transcript levels were higher in the posterior regions of the eye globe. Immunostaining with the appropriate antibody showed conclusively that bFGF protein was present in both the anterior and posterior human eye. These results suggest that local production of bFGF may stimulate cell pro liferation in vivo.
The mechanics of notochord elongation, straightening and stiffening in the embryo of Xenopus laevis
Authors: Adams Dany Spencer , Keller Ray , Koehl M. A. R.
Published: April 2021

We have examined the biomechanical development of the notochord of Xenopus early tail-bud embryos by: (1) quantifying morphological and mechanical changes in the embryo during stages 20 – 28, and (2) conducting manipulative experiments to elucidate mechanical roles of various components of the notochord. The notochord, which is composed of a stack of flat cells surrounded by a connective tissue sheath, elongates dramatically and begins straightening between stages 21 and 25. At this time the fiber density in the notochord sheath goes up, the osmotic activity of the notochord cells increases, vacuoles within these cells swell, the internal pressure of the notochord increases 2- to 3-fold, and the flexural stiffness of the notochord rises by an order of magnitude. We suggest that the tendency of the notochord cells to osmotically swell is resisted by the sheath, thereby permitting the internal pressure to rise. This pressure increase results in the greater stiffness that permits the notochord to elongate and straighten without being buckled by the surrounding tissues.
The closely related Drosophila sry β and sry δ zinc finger proteins show differential embryonic expression and distinct patterns of binding sites on polytene chromosomes
Authors: Payre François , Noselli Stéphane , Lefrère Valérie , Vincent Alain
Published: April 2021

Serendipity (sry) beta (β) and delta (δ) are two finger protein genes resulting from a duplication event. Comparison of their respective protein products shows interspersed blocks of conserved and divergent amino-acid sequences. The most extensively conserved region corresponds to the predicted DNA-binding domain which includes 6 contiguous fingers; no significant sequence conservation is found upstream and downstream of the protein-coding region. We have analysed the evolutionary divergence of the sry β and δ proteins on two separate levels, their embryonic pattern of expression and their DNA-binding properties in vitro and in vivo. By using specific antibodies and transformant lines containing β -galactosidase fusion genes, we show that the sry β and sry δ proteins are maternally inherited and present in embryonic nuclei at the onset of zygotic transcription, suggesting that they are transcription factors involved in this process. Zygotic synthesis of the sry β protein starts during nuclear division cycles 12– 13, prior to cellularisation of the blastoderm, while the zygotic sry δ protein is not detectable before germ band extension (stage 10 embryos). Contrary to sry δ, the zygotic sry β protein constitutes only a minor fraction of the total embryonic protein. The sry δ and δ proteins made in E. coli bind to DNA, with partly overlapping specificities. Their in vivo patterns of binding to DNA, visualised by immunostaining polytene chromosomes, differ both in the number and position of their binding sites. Thus changes in expression pattern and DNA-binding specificity have contributed to the evolution of the sry β and δ genes.
Pattern of the insulin-like growth factor II gene expression during early mouse embryogenesis
Authors: Lee Jacqueline E. , Pintar John , Efstratiadis Argiris
Published: April 2021

The mouse insulin-like growth factor II (IGF-II) gene encodes a polypeptide that plays a role in embryonic growth. We have examined the temporal and spatial pattern of expression of this gene in sections of the mouse conceptus between embryonic days 4.0 and 8.5 by in situ hybridization. Abundant IGF-II transcripts were detected in all the trophectodermal derivatives, after implantation. Labeling was then observed in primitive endoderm, but was transient and disappeared after formation of the yolk sac. Expression was next detected in extraembryonic mesoderm at the early primitive streak stage. Labeling in the embryo proper appeared first at the late primitive streak/neural plate stage in lateral mesoderm and in anterior-proximal cells located between the visceral endoderm and the most cranial region of the embryonic ectoderm. The position of the latter cells suggests that their descendants are likely to participate in the formation of the heart and the epithelium of the ventral and lateral walls of the foregut, where intense labeling was observed at the neural fold stage. Hybridization was also detected in cranial mesenchyme, including neural crest cells. The intensity of hybridization signal increased progressively in paraxial (presomitic and somitic) mesoderm, while declining in the ectoplacental cone. The neuroectoderm and surface ectoderm did not exhibit hybridization at any stage. Immunohistochemical analysis indicated co-localization of IGF-II transcripts, translated pre-pro-IGF-II, and the cognate IGF-II/mannose-6-phosphate receptor. These correlations are consistent with the hypothesis that IGF-II has an autocrine function.
Ectoderm nuclei from sea urchin embryos contain a Spec–DNA binding protein similar to the vertebrate transcription factor USF
Authors: Tomlinson Craig R. , Kozlowski Mark T. , Klein William H.
Published: April 2021

The Spec gene family of Strongylocentrotus purpuratus is expressed exclusively in aboral ectoderm cells during embryogenesis. To investigate the regulation of Spec gene activity, the region around the Spec1 transcriptional initiation site was analyzed for sites of protein–DNA interaction. One high-affinity site bound a factor termed SpFl within the Spec1 5′ untranslated leader region at position +39 to +60. The core sequence recognized by SpFl, CACGTG, is the same as that of the upstream stimulatory factor (USF), a widely occurring vertebrate transcription factor containing a myc-HLH motif. A comparison of USF- and SpFl-binding activities suggested that SpFl was a sea urchin version of USF. SpFl activity was detectable only in ectoderm cells of the embryo, implying that it has a role as a cell type-specific transcription factor. SpFl-binding sites were also found upstream of the Spec2a and Spec2c genes in the same conserved sequence block as Spec1. Extracts from Lyte-chinus pictus embryos showed an SpFl-like activity, suggesting that SpFl is conserved in sea urchins. Surprisingly, changes in the Spec1, Spec2a, or Specie genes that removed or modified the SpFl-binding site had no effect on expression when reporter gene fusions containing these mutations were injected into sea urchin eggs and analyzed for expression during embryogenesis. We propose that, while SpFl may not be essential for expression of the exogenously introduced reporter genes, it may be required for proper regulation of the endogenous Spec genes.
Induction of a specialized muscle basal lamina at chimaeric synapses in culture
Authors: Swenarchuk Lauren E. , Champaneria Shashikant , Anderson M. John
Published: April 2021

To identify mechanisms that regulate the formation of the neuromuscular junction, we examined the cellular origin of a heparan sulfate proteoglycan (HSPG) that becomes highly concentrated within the synaptic cleft during the initial deposition of the junctional basal lamina. Using cultured nerve and muscle cells from anuran and urodele embryos, we prepared species-chimaeric synapses that displayed spontaneous cholinergic potentials, and eventually developed organized accumulations of acetylcholine receptors and HSPG at the sites of nerve-muscle contact. To determine the cellular origin of synaptic HSPG molecules, these chimaeric junctions were stained with both species-specific and cross-reactive monoclonal antibodies, labeled with contrasting fluorochromes. Our results demonstrate that synaptic HSPG is derived almost exclusively from muscle. Since it has already been shown that muscle cells can assemble virtually all of the known constituents of the junctional basal lamina into organized surface accumulations, without any input from nerve cells, we consider the possibility that the specialized synaptic basal lamina may be generated primarily by the myofibre, in response to another ‘inductive’ positional signal at the site of nerve-muscle contact.
Rotation of the tectal primordium reveals plasticity of target recognition in retinotectal projection
Authors: Ichijo Hiroyuki , Fujita Setsuya , Matsuno Toru , Nakamura Harukazu
Published: April 2021

Retinotectal projection is precisely organized in a retinotopic manner. In normal projection, temporal retinal axons project to the rostral part of the tectum, and nasal axons to the caudal part of the tectum. The two-dimensional relationship between the retina and the tectum offers a useful experimental system for analysis of neuronal target recognition. We carried out rotation of the tectal primordium in birds at an early stage of development, around the 10-somite stage, to achieve a better understanding of the characteristics of target recognition, especially the rostrocaudal specificity of the tectum. Our results showed that temporal retinal axons projected to the rostral part of the rotated tectum, which was originally caudal, and that nasal axons projected to the caudal part of the rotated tectum, which was originally rostral. Therefore, the tectum that had been rotated at the 10-somite stage received normal topographic projection from the retinal ganglion cells. Rostrocaudal specificity of the tectum for target recognition is not determined by the 10-somite stage and is acquired through interactions between the tectal primordium and its surrounding structures.
Acrosin activation follows its surface exposure and precedes membrane fusion in human sperm acrosome reaction
Authors: Tesarik Jan , Drahorad Josef , Testart Jacques , Mendoza Carmen
Published: April 2021

Evidence has accumulated suggesting multiple roles of acrosin in fertilization, including its participation in early steps of gamete recognition and binding. However, the implication of acrosin in many of these processes is not compatible with its presumptive sequestration within the sperm acrosome until a late phase of the acrosome reaction. In an earlier study (J. Tesarik, J. Drahorad, J. Peknicova, 1988, Fertil. Steril. 50, 133-141), we reported the binding of an anti-acrosin monoclonal antibody (M0-AKR.1) to the plasma membrane overlying the acrosome of human spermatozoa starting the acrosome reaction. In this study, we characterized further this antibody with regard to its reactivity with different forms of acrosin and found that it recognizes specifically an active form of this enzyme and does not react with its proenzyme form. MO-AKR.l was thus used as a probe for in situ analysis of acrosin activation during the acrosome reaction. When suspensions of living spermatozoa were incubated with MO-AKR.l and with another monoclonal antibody (T6) directed to an intra-acrosomal cytoskeletal protein, significantly more spermatozoa reacted with the former antibody than with the latter; this indicated that some of the spermatozoa showing acrosin immunoreactivity carried activated acrosin on the cell surface, while their acrosome was still impermeable to intra-acrosomal-directed probes. The size of this particular sperm subpopulation was increased by the action of follicular fluid (a natural acrosome reaction inducer), but not ionophore A23187 (an artificial acrosome reaction inducer); it corresponded to the proportion of spermatozoa showing acrosin immunoreactivity on the plasma membrane but neither intra-acrosomal staining nor perceptible membrane perturbations when examined by immunoelectron microscopy. When spermatozoa were pre-incubated with protease inhibitors before the addition of acrosome reaction-inducing agents, the percentage of cells binding MO-AKR.l was markedly reduced. These data suggest that limited acrosin activation on the sperm plasma membrane is an early event in the physiological acrosome reaction.
The embryonic development of larval muscles in Drosophila
Authors: Bate Michael
Published: April 2021

Each of the abdominal hemisegments A2–A7 in the Drosophila larva has a stereotyped pattern of 30 muscles. The pattern is complete by 13 h after egg laying, but the development of individual muscles has begun with the definition of precursors at least by the onset of germ band shortening, some 5.5 h earlier. The earliest signs of muscle differentiation are cell fusions, which occur in the ventralmost mesoderm overlying the CNS and at stereotyped positions in the rest of the mesoderm as the germ band shortens. At the end of shortening, the pattern of muscle precursors produced by these fusions is complete. Precursors filled with dye reveal extensive fine processes probably involved in itially in cell fusion and, subsequently, in navigation over the epidermis to form attachment points. The muscle pattern is formed before innervation and without cell death. Thus, neither of these processes is involved in determining the distribution of precursors. Evidence is presented for the view that the development of the larval muscle pattern in Drosophila depends on a prior segregation of founder cells at appropriate locations in the mesoderm with which other cells fuse to form the precursors.
Zygotic activity of the nullo locus is required to stabilize the actin-myosin network during cellularization in Drosophila
Authors: Simpson Lesilee , Wieschaus Eric
Published: April 2021

Cellularization of the Drosophila embryo requires the establishment of a hexagonal network of actin and myosin filaments that are interconnected around the nuclei in the cortex of the syncytial blastoderm. This cytoskeletal network provides the framework and possibly the contractile force for the membrane invaginations that synchronously subdivide the syncytial embryo into individual cells. Zygotic expression of the nullo locus is essential for the preservation of an intact actin-myosin network. Embryos deleted for the nullo locus have a disrupted network, resulting in the formation of many multinucleate cells. We show that nullo is not required for the initial formation of the actin-myosin network, but is necessary for the maintenanee of its hexagonal shape during cellularization. The phenotype of embryonic mosaics is nonautonomous, indicating that nullo does not have to be expressed in every nucleus for proper cellularization. Examination of nullo mutant clones in adults reveals that nullo activity is not required for cell division in imaginai discs. Furthermore, germline clone experiments suggest that maternal expression of the nullo locus is not essential for either germline proliferation or the cellularization of progeny. We propose a model in which nullo functions specifically at cellularization to stabilize the actinmyosin network during contraction.
A cytoplasmic determinant for dorsal axis formation in an early embryo of Xenopus laevis
Authors: Yuge M. , Kobayakawa Y. , Fujisue M. , Yamana K.
Published: April 2021

In Xenopus laevis, dorsal cells that arise at the future dorsal side of an early cleaving embryo have already acquired the ability to cause axis formation. Since the distribution of cytoplasmic components is markedly heterogeneous in an egg and embryo, it has been supposed that the dorsal cells are endowed with the activity to form axial structures by inheriting a unique cytoplasmic component or components localized in the dorsal region of an egg or embryo. However, there has been no direct evidence for this.

To examine the activity of the cytoplasm of dorsal cells, we injected cytoplasm (dorsal cytoplasm) from dorsal vegetal cells of a Xenopus 16-cell embryo into ventral vegetal cells of a simultaneous recipient. The cytoplasm caused secondary axis formation in 42 % of recipients. Histological examination revealed that well-developed secondary axes included notochord, as well as a neural tube and somites. However, injection of cytoplasm of ventral vegetal cells never caused secondary axis and most recipients became normal tailbud embryos. Furthermore, about two-thirds of ventral isolated halves injected with dorsal cytoplasm formed axial structures.

These results show that dorsal, but not ventral, cytoplasm contains the component or components responsible for axis formation. This can be the first step towards identifying the molecular basis of dorsal axis formation.
Dynamic changes in the distribution of cytoplasmic myosin during Drosophila embryogenesis
Authors: Young Paul E. , Pesacreta Thomas C. , Kiehart Daniel P.
Published: April 2021

Dramatic changes in the localization of conventional non-muscle myosin characterize early embryogenesis in Drosophila melanogaster. During cellularization, myosin is concentrated around the furrow canals that form the leading margin of the plasma membrane as it plunges inward to package each somatic nucleus into a columnar epithelial cell. During gastrulation, there is specific antimyosin staining at the apical ends of those cells that change shape in regions of invagination.

Both of these localizations appear to result from a redistribution of a cortical store of maternal myosin. In the preblastoderm embryo, myosin is localized to the egg cortex, sub-cortical arrays of inclusions, and, diffusely, the yolk-free periplasm. At the syncytial blastoderm stage, myosin is found within cytoskeietal caps associated with the somatic nuclei at the embryonic surface. Following the final syncytial division, these myosin caps give rise to the myosin rings observed during cellularization.

These distributions are observed with both whole immune serum and affinity-purified antibodies directed against Drosophila non-muscle myosin heavy chain. They are not detected in embryos stained with antiDrosophila muscle myosin antiserum or with preimmune serum. Although immunolocalization can only suggest possible function, these myosin localizations and the coincident changes in cell morphology are consistent with a key role for non-muscle myosin in powering cellularization and gastrulation during embryogenesis.
Identification of a cytoskeletal protein localized in the myoplasm of ascidian eggs: localization is modified during anural development
Authors: Swalla Billie J. , Badgett Marty R. , Jeffery William R.
Published: April 2021

The myoplasm of ascidian eggs is a localized cytoskeletal domain that is segregated to presumptive larval tail muscle cells during embryonic development. We have identified a cytoskeletal protein recognized by a vertebrate neurofilament monoclonal antibody (NN18) which is concentrated in the myoplasm in eggs and embryos of a variety of ascidian species. The NN18 antigen is localized in the periphery of unfertilized eggs, segregates with the myoplasm after fertilization, and enters the larval tail muscle cells during embryonic development. Western blots of one-dimensional and twodimensional gels showed that the major component recognized by NN18 antibody is a 55x103Mr protein (p58), which exists in at least three different isoforms. The enrichment of p58 in the Triton X-100-insoluble fraction of eggs and its reticular staining pattern in eggs and embryos suggests that it is a cytoskeletal protein. In subsequent experiments, p58 was used as a marker to determine whether changes in the myoplasm occur in eggs of anural ascidian species, i.e. those exhibiting a life cycle lacking tadpole larvae with differentiated muscle cells. Although p58 was localized in the myoplasm in eggs of four urodele ascidian species that develop into swimming tadpole larvae, this protein was distributed uniformly in eggs of three anural ascidian species. The eggs of two of these anural species contained the actin lamina, another component of the myoplasm, whereas the third anural species lacked the actin lamina. There was no defectible localization of p58 after fertilization or segregation into muscle lineage cells during cleavage of anural eggs. NN18 antigen was uniformly distributed in pre-vitellogenic oocytes and then localized in the perinuclear zone during vitellogenesis of urodele and anural ascidians. Subsequently, NN18 antigen was concentrated in the peripheral cytoplasm of post-vitellogenic oocytes and mature eggs of urodele, but not anural, ascidians. It is concluded that the myoplasm of ascidian eggs contains an intermediate filament-like cytoskeletal network which is missing in anural species that have modified or eliminated the tadpole larva.
A newt type II keratin restricted to normal and regenerating limbs and tails is responsive to retinoic acid
Authors: Ferretti Patrizia , Brockes Jeremy P. , Brown Robin
Published: April 2021

In order to understand the molecular mechanisms underlying the regenerative ability of the urodele limb, it is important to identify regeneration-associated proteins and to study their regulation. We have recently shown that the anti-cytokeratin monoclonal antibody LP1K reacts strongly with newt blastemal cells, while its reactivity is restricted in normal limbs. By screening a cDNA expression library from the newt blastema with LP1K, we have identified cDNA clones coding for a type II keratin (NvKH) expressed both in the mesenchyme and the specialized wound epithelium of the blastema. While the rod domain of the protein is highly conserved, the homology between NvKII and mammalian type II keratins drops markedly at the N- and C-terminal regions. The expression of this keratin was analysed by Northern blotting and RNAase protection analysis of various newt tissues, and appears to be organ specific, since it is restricted to normal and regenerating limbs and tails. In particular, we have investigated the expression of this keratin mRNA in normal and regenerating limbs. The transcript is barely detectable in the proximal portion of the normal limb, but its level is high in the distal one. After amputation, NvKH mRNA is expressed both in proximal and distal blastemas, although at higher levels distally, indicating that this keratin is regeneration associated. The NvKH transcript is detectable both in mesenchyme and in the wound epithelium of the regenerate, while no transcript is detectable in normal epidermis. The level of NvKH mRNA is markedly down-regulated both in normal and regenerating limbs following intraperitoneal injection with retinoic acid, a putative endogenous morphogen in limb regeneration.
Changes in neural and lens competence in Xenopus ectoderm: evidence for an autonomous developmental timer
Authors: Servetnick Marc , Grainger Robert M.
Published: April 2021

The ability of a tissue to respond to induction, termed its competence, is often critical in determining both the timing of inductive interactions and the extent of induced tissue. We have examined the lens-forming competence of Xenopus embryonic ectoderm by transplanting it into the presumptive lens region of open neural plate stage embryos. We find that early gastrula ectoderm has little lens-forming competence, but instead forms neural tissue, despite its location outside the neural plate; we believe that the transplants are being neuralized by a signal originating in the host neural plate. This neural competence is not localized to a particular region within the ectoderm since both dorsal and ventral portions of early gastrula ectoderm show the same response. As ectoderm is taken from gastrulae of increasing age, its neural competence is gradually lost, while lens competence appears and then rapidly disappears during later gastrula stages. To determine whether these developmental changes in competence result from tissue interactions during gastrulation, or are due to autonomous changes within the ectoderm itself, ectoderm was removed from early gastrulae and cultured for various periods of time before transplantation. The loss of neural competence, and the gain and loss of lens competence, all occur in ectoderm cultured in vitro with approximately the same time course as seen in ectoderm in vivo. Thus, at least from the beginning of gastrulation onwards, changes in competence occur autonomously within ectoderm. We propose that there is a developmental timing mechanism in embryonic ectoderm that specifies a sequence of competences solely on the basis of the age of the ectoderm.
A chicken caudal homologue, CHox-cad, is expressed in the epiblast with posterior localization and in the early endodermal lineage
Authors: Frumkin Ayala , Rangini Zehava , Ben-Yehuda Adi , Gruenbaum Yosef , Fainsod Abraham
Published: April 2021

CHox-cad is a chicken homeobox gene whose homeo domain is homologous to the Drosophila caudal and the murine Cdx1 genes. Based on sequence analysis of a 2.5 kb CHox-cad cDNA clone, we deduced that the primary translation product consists of 248 amino acids. Comparison between the cDNA and genomic clones revealed the presence of an intron within the CHox-cad homeodomain between amino acids 44 and 45. The onset of CHox-cad transcription correlates temporarily with the beginning of gastrulation. During primitive streak stages CHox-cad exhibits a caudally localized pattern of expression restricted to the epiblast and the primitive streak. At these stages, CHox-cad transcripts can also be detected in the definitive endoderm cells. Later in embryogenesis CHox-cad is expressed in the epithelial lining of the embryonic gut and yolk sac. After four days of chicken development, no CHox-cad transcripts could be detected. The early CHox-cad posterior expression in the germ layer undergoing gastrulation and its continuous expression in the early endodermal lineage raise the possibility of CHox-cad involvement in the establishment of the definitive endoderm.
Spontaneous immortalisation of Schwann cells in culture: short-term cultured Schwann cells secrete growth inhibitory activity
Authors: Eccleston P. Ann , Mirsky Rhona , Jessen Kristjan R.
Published: April 2021

In the developing peripheral nerve, Schwann cells proliferate rapidly and then become quiescent, an essential step in control of Schwann cell differentiation. Cell proliferation is controlled by growth factors that can exert positive or inhibitory influences on DNA synthesis. It has been well established that neonatal Schwann cells divide very slowly in culture when separated from neurons but here we show that when culture was continued for several months some cells began to proliferate rapidly and non-clonal lines of immortalised Schwann cells were established which could be passaged for over two years. These cells had a similar molecular phenotype to short-term cultured Schwann cells, except that they expressed intracellular and cell surface fibronectin. The difference in proliferation rates between short- and long-term cultured Schwann cells appeared to be due in part to the secretion by short-term cultured Schwann cells of growth inhibitory activity since DNA synthesis of long-term, immortalised Schwann cells was inhibited by conditioned medium from short-term cultures. This conditioned medium also inhibited DNA synthesis in short-term Schwann cells stimulated to divide by glial growth factor or elevation of intracellular cAMP. The growth inhibitory activity was not detected in the medium of long-term immortalised Schwann cells, epineurial fibro-blasts, a Schwannoma (33B), astrocytes or a fibroblast-like cell-line (3T3) and it did not inhibit serum-induced DNA synthesis in epineurial fibroblasts, 33B cells or 3T3 cells. The activity was apparently distinct from trans-forming growth factor-β, activin, IL6, epidermal growth factor, atrial natriuretic peptide and gamma-interferon and was heat and acid stable, resistant to collagenase and destroyed by trypsin treatment. We raise the possibility that loss of an inhibitory autocrine loop may contribute to the rapid proliferation of long-term cultured Schwann cells and that an autocrine growth inhibitor may have a role in the cessation of Schwann cell division that precedes differentation in peripheral nerve development.
Gene regulatory factors of the sea urchin embryo: II. Two dissimilar proteins, P3A1 and P3A2, bind to the same target sites that are required for early territorial gene expression
Authors: Hoog Christer , Calzone Frank J. , Cutting Ann E. , Britten Roy J. , Davidson Eric H.
Published: April 2021

Previous work demonstrated that a negative regulatory interaction mediated by factor(s) termed ‘P3A’ is required for correct territory-specific gene expression in the sea urchin embryo. A probe derived from a P3A target site in the skeletogenic SM50 gene of Strongylocentrotus purpuratus was used to isolate a cDNA clone coding for a factor that binds specifically to this site. This factor, called P3A1, contains two sequence elements that belong to the Zn finger class of DNA-binding motifs, and in these regions is most closely similar to the Drosophila hunchback factor. The P3A1 factor also binds to a similar target sequence in a second gene, Cyllla, expressed in embryonic aboral ectoderm. Another sea urchin embryo protein factor, P3A2, has been isolated by affinity chromatography and cloned, as described in Calzone et al. Development 112, 335–350 (1991). P3A2 footprints the same target sites in the SM50 and CyIIIa genes as does P3A1, but lacks the Zn finger sequence motifs and in amino acid sequence is almost entirely dissimilar to P3A1. A deletion analysis of P3A2 delimited the DNA-binđing region, revealing that five specific amino acids in the first P3A1 finger region and four in the second P3A1 finger region are also present in equivalent positions in P3A2. The P3A1 and P3A2 factors could function as regulatory antagonists, having evolved similar target specificities from dissimilar DNA-binding domains.
Molecular composition and development of the plant glycocalyx associated with the peribacteroid membrane of pea root nodules
Authors: Perotto Silvia , Vandenbosch Kathryn A. , Butcher Geoffrey W. , Brewin Nicholas J.
Published: April 2021

In root nodules of pea (Pisum sativum), endosymbiotic Rhizobium bacteroids are individually enclosed by a plant-derived membrane, the peribacteroid membrane, creating an organelle-like structure termed the symbiosome. In order to investigate the structure and function of the peribacteroid membrane in plant-microbe surface interactions, monoclonal antibodies were used to identify the major antigenic components of this membrane and to investigate their cellular and tissue distribution. Immunofluorescence studies with isolated intact and ruptured symbiosomes indicated that the relevant antigens were confined to the luminal (inner) face of the peribacteroid membrane. Biochemical analysis indicated that the antibodies recognised carbohydrate epitopes associated with Golgi-derived glycoproteins and glycolipids. These glycoconjugate molecules are collectively termed the glycocalyx. Three different classes of glycocalyx antigens, reacting with different groups of monoclonal antibodies, could be distinguished. Immunolocalisation studies of longitudinal sections of pea nodules revealed enhanced expression of glycocalyx antigens in infected nodule tissue, with the three classes of antigen showing different degrees of expression in different developmental zones of the tissue. One of the classes of antigen was absent from the apical meristematic region of pea nodules but was re-expressed during cell invasion by Rhizobium and the development of peribacteroid membranes. In Phaseolus nodules, as opposed to those of Pisum, this group of antigens was entirely lacking from the central infected tissue.
Xenopus temporal retinal neurites collapse on contact with glial cells from caudal tectum in vitro
Authors: Johnston Alexander R. , Gooday Douglas J.
Published: April 2021

Nasal and temporal retinal neurites were confronted in culture with glial cells from the rostral and caudal parts of the optic tectum and with glial cells from the diencephalon. Twenty of each of the six classes of encounter between individual growth cones and isolated glial cells were analysed by time-lapse videorecording. The results show that growth cones from the temporal retina collapse when they contact glial cells from the caudal tectum, but do not collapse when they contact glia from other areas. Growth cones of nasal retinal fibres do not collapse on contact with any of the glial types examined. This suggests that the inhibitory phenomena described by others are in part due to the cell surface characteristics of glial cells, and that there are differences between glia from the front and back of the optic tectum.
Chicken homeobox gene Msx-1: structure, expression in limb buds and effect of retinoic acid
Authors: Yokouchi Yuji , Ohsugi Kojune , Sasaki Hiroshi , Kuroiwa Atsushi
Published: April 2021

A chicken gene carrying a homeobox highly homologous to the Drosophila muscle segment homeobox (msh) gene was isolated and designated as Msx-1. Conceptual translation from the longest ORF gave a protein of 259 amino acids lacking the conserved hexapeptide. Northern analysis detected a single 2.6 kb transcript. As early as day 2 of incubation, the transcript was detected but was not found in adult tissue. In situ hybridization analysis revealed that Msx-1 expression is closely related to a particular mesenchymal cell lineage during limb bud formation. In early stage embryos, Msx-1 was expressed in the somatopleure. When primordial mesenchyme cells for limb bud were generated from the Wolffian ridge of the somatopleure, Msx-1 expression began to diminish in the posterior half of the limb bud then in the presumptive cartilage-forming mesenchyme.

In developing limb buds, remarkable expression was seen in the apical ectodermal ridge (AER), which is responsible for the sustained outgrowth and development of the limb. The Msx-1 transcripts were found in the limb mesenchymal cells in the region covering the necrotic zone and ectodermal cells overlying such mesenchymal cells. Both ectodermal and mesenchymal expression in limb bud were rapidly suppressed by local treatment of retinoic acid which can generate mirrorimage duplication of digits. This indicates that retinoic acid alters the marginal presumptive non-cartilage forming mesenchyme cell lineage through suppression of Msx-1 expression
Pattern formation in the nematode epidermis: determination of the arrangement of peripheral sense organs in the C. elegans male tail
Authors: Baird Scott E. , Fitch David H. A. , Kassem Iman A. A. , Emmons Scott W.
Published: April 2021

The developmental process that determines the arrangement of ray sensilla in the Caenorhabditis elegans male tail has been studied. It is shown that the adult arrangement of rays is determined by the placement of ray cells at specific sites in the epidermis of the last larval (L4) stage. Placement of ray cells at specific epidermal sites results from the generation of neurons and support cells in the epidermis near to their final positions, and the subsequent refinement of these positions by an active mechanism involving specific cellular associations. Positions of ray cells and adjacent epidermal cells have been studied during ray development by means of indirect immunofluorescence staining with an antibody to a cell junctional antigen. Mutations are described in six genes that alter the adult arrangement of the rays, frequently resulting in fusion of rays. Changes in the adult pattern of rays in mutants appear to result from prior changes in the epidermal positions of ray cells, and for two mutants it is suggested that this may be due to the inappropriate clustering of processes from neurons and support cells of adjacent rays. Development of the wild-type arrangement of rays appears to require the specification of molecular differences between the rays that affect the specificity of their cellular associations.
Effect of egg composition on the developmental capacity of androgenetic mouse embryos
Authors: Latham Keith E. , Solter Davor
Published: April 2021

Analysis of the developmental capacities of androgenetic and gynogenetic mouse embryos (bearing two paternal or two maternal pronuclei, respectively) revealed a defect in blastocyst formation of androgenetic, but not gynogenetic, embryos that was a function of the maternal genotype. Androgenetic embryos constructed using fertilized eggs from C57BL/6 or (B6D2)F1 mice developed to the blastocyst stage at frequencies similar to those previously reported, whereas androgenetic embryos constructed with fertilized eggs from DBA/2 mice developed poorly, the majority failing to progress beyond the 16-cell stage and unable to form a blastocoel-like cavity, regardless of whether the male pronuclei were of C57BL6 or DBA/2 origin. This impaired development was observed even in androgenetic embryos constructed by transplanting two male pronuclei from fertilized DBA/2 eggs to enucleated C57BL/6 eggs, indicating that the defect cannot be explained as the lack of some essential component in the DBA/2 cytoplasm that might otherwise compensate for androgeny.

Rather, the DBA/2 egg cytoplasm apparently modifies the incoming male pronuclei differently than does C57BL/6 egg cytoplasm. Several specific alterations in the protein synthesis pattern of DBA/2 androgenones were observed that reflect a defect in the regulatory mechanisms that normally modulate the synthesis of these proteins between the 8-cell and blastocyst stages. These results are consistent with a model in which cytoplasmic factors present in the egg direct a strain dependent modification of paternal genome function in response to epigenetic modifications (genomic imprinting) established during gametogenesis and indicate that preimplantation development can be affected by these modifications at both the morphological and biochemical levels.
The determination of sense organs in Drosophila: effect of the neurogenic mutations in the embryo
Authors: Goriely Anne , Dumont Nathalie , Dambly-Chaudière Christine , Ghysen Alain
Published: April 2021

We have examined the early pattern of sensory mother cells in embryos mutant for six different neurogenic loci. Our results show that the neurogenic loci are required to restrict the number of competent cells that will become sensory mother cells, but are not involved in controlling the localization or the position-dependent specification of competent cells. We conclude that these loci are involved in setting up a system of mutual inhibition, which transforms graded differences within the proneural clusters into an all- or-none difference between one cell, which becomes the sense organ progenitor cell, and the other cells, which remain epidermal.
Genetic approaches to inflorescence and leaf development in maize
Authors: Veit Bruce , Greene Ben , Lowe Brenda , Mathern Julie , Sinha Neelima , Vollbrecht Erik , Walko Richard , Hake Sarah
Published: April 2021

The application of genetic methods to the analysis of morphogenesis in maize is described. Several classes of floral mutants are differentiated through developmental studies and tests of epistasis. The results of mosaic and dosage analysis of Kn1, a dominant mutation affecting leaf development, are related to molecular studies of the gene.