bluuebunny/crossref_metadata_2025 huggingface dataset:
Analysis of function of the pair-rule genes hairy, even-skipped and fushi tarazu in mosaic Drosophiia embryos
Authors: Lawrence Peter A. , Johnston Paul
Published: April 2021

We report the first attempt of its kind to study genetic interactions using young Drosophila embryos that are mosaic for wildtype and mutant cells. Using nuclear transplantation we make mosaic embryos in which a patch of cells lacks a particular segmentation gene, A. With antibodies, we than look at the expression of another gene that is known to be downstream of gene A, with respect to the cells in the patch. We have examples of patches of hairy cells (where we monitor the effect on fushi tarazu (ftz) expression), even-skipped (monitoring ftz) and ftz (monitoring engrailed and Ultrabithorax).

Our main finding is that the dependence of engrailed expression on the ftz gene is strictly cell-autonomous. This result goes some way towards explaining the dependence of Ultrabithorax expression on ftz, a dependence we show to be locally cell-autonomous within parts of parasegments 6 and 8 but non autonomous within parasegment 7.
Molecular approaches to vertebrate limb morphogenesis
Authors: Smith Susan M. , Pang Kevin , Sundin Olof , Wedden Sarah E. , Thaller Christina , Eichele Gregor
Published: April 2021

It has long been proposed that concentration gradients of morphogens provide cues to specify cell fate in embryonic fields. Recent work jn a variety of vertebrate systems give bona fide evidence that retinoic acid, the biologically active form of vitamin A, is a candidate for such a morphogen. In the developing chick wing, for example, locally applied retinoic acid triggers striking changes in the pattern along the anteroposterior axis. Instead of giving rise to a wing with the normal 234 digit pattern, wing buds treated with retinoic acid develop a 432234 mirror-image symmetrical digit pattern.

For this review, we focus on three aspects of limb morphogenesis. (1) We summarize the experimental evidence supporting the notion that retinoic acid is a candidate morphogen. (2) Limb buds contain high levels of cellular retinoic-acid-binding protein (CRABP). Using order of magnitude calculations, we evaluate how the concentration of CRABP might affect the occupancy state of the retinoic acid receptor. (3) We discuss the spatio-temporal expression pattern of homeobox-con-taining genes in the developing limb and speculate about the possibility that retinoic acid influences the pattern of expression of homeobox genes.
Genetics of intercellular signalling in C. elegans
Authors: Austin Judith , Maine Eleanor M. , Kimble Judith
Published: April 2021

Cell-cell interactions play a significant role in controlling cell fate during development of the nematode Caenorhabditis elegans. It has been found that two genes, glp-1 and lin-12, are required for many of these decisions, glp-1 is required for induction of mitotic proliferation in the germline by the somatic distal tip cell and for induction of the anterior pharynx early in embryogenesis, lin-12 is required for the interactions between cells of equivalent developmental potential, which allow them to take on different fates. Comparison of these two genes on a molecular level indicates that they are similar in sequence and organization, suggesting that the mechanisms of these two different sets of cell-cell interactions are similar.
Role of the neuropeptide head activator for growth and development in hydra and mammals
Authors: Schaller H. Chica , Hoffmeister Sabine A. H. , Dübel Stefan
Published: April 2021

In hydra, HA is produced by nerve cells and released into the intercellular space bound to large-molecular-weight carrier(s). By additional interaction with extracellular matrix components and selfinactivation by dimérisation, a local action is ensured. HA acts as a mitogen on all dividing cell types in hydra forcing them to pass through G2, divide, and either start a new round of cell division or terminally differentiate. In addition, HA is required for head-specific determination and differentiation processes. To become a head-specific nerve cell, for example, an interstitial stem cell requires HA in early S-phase to become determined to the nerve cell pathway, in late G2 to progress through mitosis, and/or in Gj to differentiate to a head-, and not to a foot-, specific nerve cell. HA (with identical amino acid sequence) occurs in other animals including mammals. In mammals, it is produced by nerve or endocrine cells and it probably acts, as in hydra, on nerve-precursor cells. On the neural cell line NH15-CA2 and on the pituitary cell line AtT20, HA acts as mitogen by stimulating cells arrested in G2 to enter mitosis. The presence of HA early in neural development and in abnormal neural development, such as in brain and neuroendocrine tumors, are consistent with a function in growth control for HA in mammals.
Profile of a mammalian sperm receptor1,2
Authors: Wassarman Paul M.
Published: April 2021

Complementary molecules on the surface of eggs and sperm are responsible for species-specific interactions between gametes during fertilization in both plants and animals. In this essay, several aspects of current research on the mouse egg receptor for sperm, a zona pellucida glycoprotein called ZP3, are addressed. These include the structure, synthesis, and functions of the sperm receptor during oogenesis and fertilization in mice. Several conclusions are drawn from available information. These include (/) ZP3 is a member of a unique class of glycoproteins found exclusively in the extracellular coat (zona pellucida) of mammalian eggs. (If) ZP3 gene expression is an example of oocyte-specific and, therefore, sex-specific gene expression during mammalian development. (Ill) ZP3 is a structural glycoprotein Involved in assembly of the egg extracellular coat during mammalian oogenesis. (ZV) ZP3 is a sperm receptor involved in carbohydrate-mediated gamete recognition and adhesion during mammalian fertilization. (V) ZP3 is an inducer of sperm exocytosis (acrosome reaction) during mammalian fertilization. (Vf) ZP3 participates in the secondary block to polyspermy following fertilization in mammals. (VII) The extracellular coat of other mammalian eggs contains a glycoprotein that is functionally analogous to mouse ZP3. The unique nature, highly restricted expression, and multiple roles of ZP3 during mammalian development make this glycoprotein a particularly attractive subject for investigation at both the cellular and molecular levels.
Glue protein genes in Drosophila virilis: their organization, developmental control of transcription and specific mRNA degradation
Authors: Swida Ulrike , Lucka Lothar , Kress Horst
Published: April 2021

The gene Lgp-1, which is localized in the intermoult puff 16A of D. virilis polytene chromosomes, encodes the major larval glue protein lgp-1. The gene consists of two exons interrupted by a short intron. In the 5′ flanking region of Lgp-1, we find putative ecdysone receptor binding sites and two proximal conserved sequence motifs which are possibly involved in gene regulation. The amino acid sequence deduced from the DNA sequence reveals a relationship to the 68C glue protein family of D. melanogaster. The size of the Lgp-1 transcripts decreases in late third instar larvae concomitantly with their disappearance. This is caused by deadenylation followed by distinct nucleolytic attacks in the 3′untranslated region. Preliminary data suggest the presence of another glue protein gene in the 16A puff region which is related to the Lgp-1 gene.
A two-step model for the localization of maternal mRNA in Xenopus oocytes: Involvement of microtubules and microfilaments in the translocation and anchoring of Vg1 mRNA
Authors: Yisraeli Joel K. , Sokol Sergei , Melton D. A.
Published: April 2021

In an effort to understand how polarity is established in Xenopus oocytes, we have analyzed the process of localization of the maternal mRNA, vg1. In fully grown oocytes, vg1 mRNA is tightly localized at the vegetal cortex. Biochemical fractionation shows that the mRNA is preferentially associated with a detergent-insoluble subcellular fraction. The use of cytoskeletal inhibitors suggests that (1) microtubules are involved in the translocation of the message to the vegetal hemisphere and (2) microfilaments are important for the anchoring of the message at the cortex. Furthermore, immunohistochemistry reveals that a cytoplasmic microtubule array exists during translocation. These results suggest a role for the cytoskeleton in localizing information in the oocyte.
Genital ridges exert long-range effects on mouse primordial germ cell numbers and direction of migration in culture
Authors: Godin Isabelle , Wylie Christopher , Heasman Janet
Published: April 2021

The functional gametes of all vertebrates first arise in the early embryo as a migratory population of cells, the primordial germ cells (PGCs). These migrate to, and colonise, the genital ridges (GR) during the early organogenesis period, giving rise to the complete differentiating gonad. PGCs first become visible by alkaline phosphatase staining in the root of the developing allantois at 8.5 days post coltum (dpc). At 9.5 dpc they are found in the wall of the hind-gut and, during the following three days, they migrate along the hind-gut mesentery to the dorsal body wall, and then to the genital ridges. By 12.5 dpc, the great majority of PGCs have colonised the genital ridges. During this period the number of PGCs increases from less than 100 to approximately 4000.

In a previous paper (Donovan et al. 1986), we showed that 10.5 dpc PGCs can be explanted from the hind-gut mesentery, and will spread and migrate on feeder cell layers. We showed also that the intrinsic ability of PGCs to spread and migrate changes as they colonise the genital ridges. In this paper, we examine extrinsic factors that control PGC behaviour in vitro. Using PGCs taken from 8.5 dpc embryos, at the beginning of their migratory phase, we show that culture medium conditioned by 10.5 dpc genital ridges causes an increase in the number of PGCs in these cultures. We also show that PGCs migrate towards 10.5 dpc genital ridges in preference to other explanted organs.

These experiments show that genital ridges exert long-range effects on the migrating population of PGCs. They also suggest that tropic factors released from the genital ridges play a role in PGC guidance.
Requirements for integrins during Drosophila development
Authors: Zusman Susan , Patel-King Ramila S. , Ffrench-Constant Charles , Hynes Richard O.
Published: April 2021

The common subunit of the PS antigens of Drosophila is homologous with vertebrate integrins and is encoded by the lethal(1)myospheroid gene. We have generated flies mosaic for wild-type and mutant alleles of lethal-(1) myospheroid using adult gynandromorphs and radiation-induced somatic crossing over. The defects observed in the gynandromorphs demonstrate widespread requirements for PS integrins during development especially in ventrally derived structures, which also show strong expression of PS β integrin. Smaller lethal(1)myo-spheroid clones induced during larval development result in blister and vein defects in the wings and aberrant development of photoreceptor cells, demonstrating roles for PS integrins during development of both wings and eyes. PS integrins are required for the close apposition of the dorsal and ventral wing epithelia and for the proper arrangement of photoreceptor cells. However, many other adhesive and morphogenetic processes proceed normally in the absence of integrins containing the fl subunit encoded by lethal(1)myospheroid.
Position-specific activity of the Hox1.1 promoter in transgenic mice
Authors: Püschel Andreas W. , Balling Rudi , Gruss Peter
Published: April 2021

During development, positional values have to be assigned to groups of cells. The murine Hox genes are a class of genes that are predicted to be involved at some stage in this process. During embryogenesis they are expressed in distinct overlapping region- and stagespecific patterns and therefore must be regulated in response to positional information. In this study, we have analysed the activity of Hox1.1 promoter sequences in transgenic mice. The use of lacZ as a marker allows a detailed analysis of expression at the single cell level during early embryonic development. We show that 3.6 kbp of promoter and 1.7 kbp of 3’ sequences provide sufficient regulatory information to express a transgene in a spatial and temporal manner indistinguishable from the endogenous Hox1.1 gene during the period of development when Hox1.1 expression Is established. The activation occurs in a strict order in specific ectodermal and mesodermal domains. Within each of these domains the transgene is activated over a period of four hours apparently randomly in single cells. In a following second period, Hox1.1 and transgene expression patterns diverge. In this period, transgene expression persists in many mesodermally derived cells that do not express Hox1.1 indicating the absence of a negative regulatory element in the transgene. The anterior boundary of transgene expression is identical to that of Hox1.1. However, no posterior boundary of transgene expression is set, suggesting that a separate element absent from the transgene specifies this boundary.
TGF beta in murine morphogenetic processes: the early embryo and cardiogenesis
Authors: Akhurst Rosemary J. , Lehnert Sigrid A. , Faissner Andreas , Duffie Elizabeth
Published: April 2021

The tissue distribution of TGF beta-1 RNA was examined within whole mouse embryos from implantation to 10.5 days gestational age and, in the developing heart, up to 8 days postpartum. The earliest high level expression of TGF beta-1 RNA is at 7.0 days postcoUum ip.c.) in the cardiac mesoderm. At 8.0 days gestational age, cardiac TGF beta-1 RNA expression is limited to endocardia! cells. By 9.5 days p.c, this expression pattern becomes regionalised to those cells that overlie cardiac cushion tissue. High TGF beta-1 RNA levels continue to persist in endothelial cells of the heart valves until approximately one week postpartum.

The TGF beta-1 RNA distribution was compared with the extracellular distributions of polypeptides for TGF beta and Jl/tenascin. As previously reported, endothelial expression of TGF beta-1 RNA is correlated with mesenchymal expression of TGF beta polypeptide, suggesting a paracrine mode of action for this growth factor in cardiac development. Minor discrepancies in the distributions of TGF beta-1 RNA and the extracellular form of the TGF beta polypeptide suggest that translational or post-translational control of protein levels occurs and/or the possibility that the antibody used may also recognise other members of the TGF beta polypeptide family.

A correlation between endothelial TGF beta-1 expression and distribution of Jl/tenascin in the mesenchyme gives further support to the proposition that the biological effects of TGF beta-1 may, in part, be mediated by Jl/tenascin.
Regulation of fission yeast mating-type interconversion by chromosome imprinting
Authors: Klar Amar J. S.
Published: April 2021

Mating types of the fission yeast Schizosaccharomyces pombe interchange nonrandomly in a cell lineage so that only one cell among four granddaughters of a cell ever switches, and the sister of the newly switched cell switches efficiently in consecutive cell divisions, thereby producing chains of recurrent switching. The programme of cellular differentiation is mediated by inheritance of parental DNA chains at the mating type locus (matl) by progeny cells. This review summarizes recent results suggesting that two types of imprinting events at the matl locus are required to generate the specific pattern of switching in a cell lineage. One of those is a site- and strand-specific event that is required before the mat1 locus can be cleaved in vivo. The other is a double-stranded break at matl that is healed by gene conversion in the progeny cells resulting in switching the matl locus.
Gametic imprinting in maize in relation to the angiosperm life cycle
Authors: Kermicle Jerry L. , Alleman Mary
Published: April 2021

Differences in the activity of maternally and paternally derived genomes in maize endosperm have been observed at three levels of genetic manipulation. When the balance of entire chromosome sets departs from the standard ratio of two of maternal origin to one of paternal origin, development is impaired, often leading to seed failure. At the level of individual chromosomes, absence of a paternal representative for 8 of the 19 chromosome arms tested causes a marked reduction in kernel size. Replacement of the missing arms by ones of maternal origin does not complement this defect. At the gene level, some alleles of R confer solid coloration on the aleurone layer when transmitted maternally but patchy coloration (mottled) when transmitted via pollen. In contrast with the endosperm, no effect of parentage on R phenotype has been detected in embryonic and seedling tissues. Furthermore, gynogenetic and androgenetic haploid plants are viable in maize and are similar in appearance. The detection of parental effects in the endosperm, but not the embryo, points to the few cell divisions of the gametophytes as a critical stage in imprinting. Chromosomally based epigenetic variation originating at this stage would be reflected as imprinting effects. A separate fertilization establishes a line of genetic descent in the embryo that appears to be relatively free of imprinted genes.
Fusion between myoblasts and adult muscle fibers promotes remodeling of fibers into myotubes in vitro
Authors: Hinterberger Timothy J. , Barald Kate F.
Published: April 2021

Muscle satellite cells are residual embryonic myoblast precursors responsible for muscle growth and regeneration. In order to examine the role of satellite cells in the initial events of muscle regeneration, we placed individual mature rat muscle fibers in vitro along with their satellite cells. When the satellite cells were allowed to proliferate, they produced populations of myoblasts that fused together to form myotubes on the laminin substrate. These myoblasts and myotubes also fused with the adult fibers. When they did so, the fibers lost their adult morphology, and by 8 days in vitro, essentially all of them were remodeled into structures resembling embryonic myotubes. However, when proliferating satellite cells were eliminated by exposure to cytosine arabinoside (araC), the vast majority of fibers retained their adult shape. Addition of C2C12 cells (a myoblast line derived from adult mouse satellite cells) to araC-treated fiber cultures resulted in their fusion with the rat muscle fibers and restored the ability of the fibers to remodel, whereas addition of either a fibroblast cell line or a transformed, non-fusing variant of C2C12 cells, or addition of conditioned medium from C2C12 cells, failed to do so. These results imply that myoblast fusion is responsible for triggering adult fiber remodeling in vitro.
Expression of J1 / ten asci n in the crypt-villus unit of adult mouse small intestine: implications for its role in epithelial cell shedding
Authors: Probstmeier Rainer , Martini Rudolf , Schachner Melitta
Published: April 2021

The localization of the extracellular matrix recognition molecule J1/tenascin was investigated in the cryptvillus unit of the adult mouse ileum by immunoelectron microscopic techniques. In the villus region, J1/tenascin was detected strongly in the extracellular matrix (ECM) between fibroblasts of the lamina propria. It was generally absent in the ECM at the interface between sub-epithelial fibroblasts and intestinal epithelium, except for some restricted areas along the epithelial basal lamina of villi, but not of crypts. These restricted areas corresponded approximately to the basal part of one epithelial cell. In J1/tenascin-positive areas, epithelial cells contacted the basal lamina with numerous microvillus-like processes, whereas in J1/tenascin-negative areas the basal surface membranes of epithelial cells contacted their basal lamina in a smooth and continuous apposition. In order to characterize the functional role of J1/tenascin in the interaction between epithelial cells and ECM, the intestinal epithelial cell line HT-29 was tested for its ability to adhere to different ECM components. Cells adhered to substratum-immobilized fi bronectin, laminin and collagen types I to IV, but not to J1/tenascin. When laminin or collagen types I to IV were mixed with J1/tenascin, cell adhesion was as effective as without J1/tenascin. However, adhesion was completely abolished when cells were offered a mixture of fibronectin and J1/tenascin as substratum. The ability of J1/tenascin to reduce the adhesion of intestinal epithelial cells to their fibronectin-containing basal lamina suggests that J1/tenascin may be involved in the process of physiological cell shedding from the villus.
Guidance of commissural growth cones at the floor plate in embryonic rat spinal cord
Authors: Bovolenta Paola , Dodd Jane
Published: April 2021

The floor plate of the embryonic rat spinal cord has been proposed to act as an intermediate target that plays a role in the pattern of extension of commissural axons. To begin to examine the role of the floor plate in axon guidance at the midline, we have studied the precision of the commissural axon projection to and across the floor plate during development. To delineate the pathway, the fluorescent carbocyanine dye, Di-I, has been used as a probe. We show that commissural axons traverse the floor plate and turn rostrally at its contralateral border with remarkable precision. Axons were not observed to turn ipsilaterally and turned only upon reaching the contralateral edge of the floor plate. Virtually all com missural axons follow this route. The morphology of commissural growth cones was also examined. As they encountered the floor plate, commissural growth cones became larger and increased in complexity. The reorientation of axons in register with the floor plate boundary and the change in the morphological properties of commissural growth cones as they traverse the midline suggest that the floor plate may act as a guidepost with functions similar to cells that have been implicated in axon guidance in invertebrates.
Hydrogen peroxide levels in mouse oocytes and early cleavage stage embryos developed in vitro or in vivo
Authors: Nasr-Esfahani Mohammed H. , Aitken John R. , Johnson Martin H.
Published: April 2021

We describe a fluorimetric method for measuring the level of H2O2 in individual mouse oocytes and early embryos. Levels of H2O2 are low but detectable in unfertilized oocytes recovered freshly from the female reproductive tract. The levels in early cleaving embryos (1-cell to 8-cell stages) immediately after recovery from the female tract seem to be slightly higher the later the stage examined. However, when embryos are cultured in vitro from the 1-cell or early 2-cell stage, H2O2 levels rise when the embryos reach the mid-2-cell stage and remain elevated until they enter the early 4-cell stage. No equivalent elevation of H2O2 is seen during the transition from 1-cell to 2-cell or from 4-cell to 8-cell stages. Embryos that are able to develop successfully in vitro, as well as those that show a developmental block at the 2-cell stage on culture in vitro, both show this rise in H2O2 levels after in vitro culture. The relationship between the rise in H2O2 and the ‘2-cell block’ to development is discussed.
Lateral inhibition and the development of the sensory bristles of the adult peripheral nervous system of Drosophila
Authors: Simpson Pat
Published: April 2021

Cells in the neurectoderm of Drosophila face a choice between neural and epidermal fates. On the notum of the adult fly, neural cells differentiate sensory bristles in a precise pattern. Evidence has accumulated that the bristle pattern arises from the spatial distribution of small groups of cells, proneural clusters, from each of which a single bristle will result. One class of genes, which includes the genes of the achaete-scute complex, is responsible for the correct positioning of the proneural clusters. The cells of a proneural cluster constitute an equivalence group, each of them having the potential to become a neural cell. Only one cell, however, will adopt the primary, dominant, neural fate. This cell is selected by means of cellular interactions between the members of the group, since if the dominant cell is removed, one of the remaining, epidermal, cells will switch fates and become neural. The dominant cell therefore prevents the other cells of the group from becoming neural by a phenomenon known as lateral inhibition. They, then, adopt the secondary, epidermal, fate. A second class of genes, including the gene shaggy and the neurogenic genes mediate this process. There is some evidence that a proneural cluster is composed of a small number of cells, suggesting a contact-based mechanism of communication. The molecular nature of the protein products of the neurogenic genes is consistent with this idea.
Patterning of chick brain vesicles as revealed by peanut agglutinin and cholinesterases
Authors: Layer Paul G. , Alber Regina
Published: April 2021

Differentiation of individual rhombomeres of the chicken hindbrain directly follows the emergence of primary brain vesicles. Immediately after the constric tion of the prosencephalon at HH9, a series of vesicles of decreasing size is established almost simultaneously between HH9 and HH10, including mesencephalon, four preotic (R2–R5) and one postotic (R6/R7) rhombo meres. Thereby, the cranial neural tube is ventrally embedded in a mesodermal PNA-binding matrix that particularly accumulates underneath vesicular constric tion sites, as demonstrated for the segregation of the prosencephalon at HH9 and the cerebellar rhombomere R1 from R2 at HH13. The subsequent period of hind brain differentiation is analyzed by cholinesterase (AChE, BChE) and peanut lectin histochemistry, by the BrdU and the neurite-specific G4 antibodies. Preotically, differentiation of two pairs of rhombomeres (R4+R5, R2+R3) starts in R4, immediately followed by R2. The caudal rhombomeres of both pairs are delayed (R5, R3). Then the postotic rhombomere is subdivided, whereby R7 differentiates before R6. Thus, the development in the direct vicinity of the otic vesicle is delayed (R5, R6). R7 is the last rhombomere that is demarcated caudally.

Based on these findings, we postulate two processes that may regulate rhombomere formation in the chicken embryo: (a) an early rostrocaudal wave establishing the major brain vesicles, (b) a superimposed pairwise seg mentation emanating rostrally and caudally from the otic vesicle. The segregation of the cerebellar rhombo mere is a late step.

Abbreviations: AChE, acetylcholinesterase (E.C. 3.1.1.7);

BChE, butyrylcholinesterase (E.C. 3.1.1.8);

BrdU, bromodesoxyuridine;

D, diencephalon;

E, eye stalk;

G4, neurite-specific antigen;

M, mesencephalon;

NP, neuropore;

OV, otic vesicle;

PNA, peanut agglutinin;

P, prosencephalon;

Rl, rhombomere No. 1;

R, rhombencephalon; T, telencephalon.
Development of homogeneous fast and slow motor units in the neonatal mouse soleus muscle
Authors: Fladby Tormod , Jansen Jan K. S.
Published: April 2021

We studied the fiber type composition and contractile properties of mouse soleus motor units at 2 days, 5 days and 2 weeks of age. We used Lucifer Yellow injection to mark muscle fibers belonging to the same motor unit in the two youngest age groups, and the traditional method of glycogen depletion in the oldest. The age groups were chosen because 2 days is at the end of muscle fiber production; 5 days is at the start of synapse elimination in the muscle and 2 weeks is at the end. Muscle fibers were classified as fast (F) or slow (S) on the basis of their myosin heavy chain (MHC) content, as determined by different monoclonal antibodies.

Motor units are already dominated by either F- or S-fibers at 2 days, suggesting an early preferential innervation of the two types of fibers. A substantial part of the remaining refinement of the innervation takes place during the next 3 days, while the total number of terminals in the muscle remains constant. This is most easily explained by an exchange of aberrant for correct synapses during this period. A smaller part of the refinement of the innervation occurs during the sub sequent period of synapse elimination.
Autodifférenciation et induction de cartilage à partir de mésenchyme somitique de Poulet cultivéin vitro
Authors: Strudel par G.
Published: April 2021

Self-differentiation and induction of cartilage from chick somitic mesenchyme cultured in vitro

It has been shown (Strudel & Watterson) that in the absence of inductors (neural tube and notochord) the somitic mesenchyme of the chick embryo does not differentiate in situ into cartilage until after the 27-30 somite stage. In culture, in a medium based upon embryo extract (diluted or undiluted), cartilage only differentiates in mesenchyme taken from embryos of 27 somites. If, on the other hand, the mesenchyme is wrapped in vitelline membrane and cultured in a medium containing undiluted embryo extract it self-differentiates from the 17-somite stage. Differentiation into cartilage can be obtained in mesenchyme taken from younger embryos if extracts of inductors—neural tube and notochord—are added to the culture medium. In these circumstances chondrogenesis may occur in mesenchyme from an embryo of 20 somites when the embryo extract is diluted or from an embryo of 13 somites if it is not. If the somitic mesenchyme from 8 somite embryos is both wrapped in vitelline membrane and cultured in a medium based upon undiluted embryo extract then differentiation of cartilage occurs. These results show that the time-relations of chondrogenesis from chick somitic mesenchyme cultured in vitro varies with the composition of the medium and with the method of culture. These variations make it clear that the capacity of somitic mesenchyme to differentiate into cartilage may exist at very early stages. This early capacity is not revealed unless the somitic mesenchyme is cultured in a very favourable nutritive medium and in the presence of extracts of inducing tissues.
The Early Development of the Otic Vesicle in Staged Human Embryos
Authors: O’Rahilly Ronan
Published: April 2021

The early development of the otic vesicle has been investigated in thirty-six human embryos from stage 9 to stage 18 (3 to 6 post-ovulatory weeks). The otic zone was first distinguished at stage 9 and invagination had commenced in the more advanced specimens of stage 10. The otic pit deepened progressively during stage 11, and certain resemblances to the developing lens were noted. It is believed that contributions from the wall of the otic pit to the vestibulocochlear crest occurred, e.g. at stage 12, but they were not as conspicuous as the corresponding crest migrations from the optic primordium to the sheath of the optic vesicle. The otic pit became closed from the surface at stage 13 to form the otic vesicle, the dorsomedial portion of which became recognizable as the endolymphatic appendage. As in the case of the lens, a part of the basement membrane of the surface ectoderm became invaginated (stages 11 to 13) to form the basement membrane surrounding the otic vesicle. Although their precursors were detectable earlier, the cochlear and the semicircular ducts were clearly defined by the end of stage 18. During the formation of the semicircular ducts, portions of the epithelium lost their identity and their basement membrane disappeared, so that the epithelium merged with the underlying mesoderm, and obliteration of portions of the cavity of the otic vesicle ensued. It has been established previously that the various basement membranes associated with the embryonic eye persist into adulthood. Moreover, it is known that at least certain portions of the adult membranous labyrinth possess a basement membrane, derived presumably from that surrounding the otic vesicle in the embryo. The possible existence of a continuous and intact basement membrane throughout the adult membranous labyrinth, however, seems to require further investigation.
Localization of specific mRNAs in Xenopus embryos by whole-mount in situ hybridization
Authors: Hemmati-Brivanlou Ali , Frank Dale , Bolce Margaret E. , Brown Bob D. , Sive Hazel L. , Harland Richard M.
Published: April 2021

We have adapted a non-radioactive technique to detect localized mRNAs in whole-mount Xenopus embryos. Synthetic antisense RNA transcribed in the presence of digoxygenin-UTP is used as a probe and is detected via an anti-digoxygenin antibody. We show that localized mRNAs can be detected from late gastrula to tadpole stages and that high as well as low abundance RNAs can be detected. The method was tested on muscle actin and α-globin RNAs, whose localization has previously been characterized. In addition, we used the method to determine the distribution of XA-1 RNA, an anterior ectoderm-specific RNA, which we show is expressed in the periphery of the cement gland as well as in the region of the hatching gland. The sequence of an XA-1 cDNA predicts a protein rich in proline and histidine.
Gastric endocrine cells share a clonal origin with other gut cell lineages
Authors: Thompson E. M. , Fleming K. A. , Evans D. J. , Fundele R. , Surani M. A. , Wright N. A.
Published: April 2021

There has been considerable debate about the ontological origin of gut endocrine cells as being either from the neural crest (or primitive epiblast) or from the endodermal stem cell. We have attempted to define the ontological origin of endocrine cells by applying an experimental system that uses a marker to Identify one of the two phenotypes present in chimaeric mice as suggested by Ponder et al. (1985).

This study involved two separate experiments. The first made use of the unique staining properties of Dolichos bifloras agglutinin (DBA), a lectin that binds to the A-acetyl galactosamine sugar residues present on the surface of C57B1 mouse gut, but absent from RoRIII mouse gut, in C57Bl↔Roin mouse chimaeras at the ultrastructural level. A four-stage procedure for staining at the EM level was developed. Although mature villous endocrine cells stained for DBA, immature endocrine cells did not, either in the positive crypts of chimaeric mouse gut or in gut from C57B1 positive controls.

Thus a second marker was chosen. This experiment combined immunocytochemistry (to identify gastric antral gastrin cells chosen as a representative neuro-endocrine cell) with in situ DNA hybridization for the mouse male chromosome repeat sequence PY 353 (to identify XY cells) in XX→XY chimaeric mice. This study showed that the sex chromosomal pattern in the gastrin cells parallels that of other cells in the same gastric gland and therefore are clonal with them. This suggests that gut endocrine cells share a common stem cell with other epithelial cell lineages in the antrum and are endodermally derived.
Primary neurons that express the L2/HNK-1 carbohydrate during early development in the zebrafish
Authors: Metcalfe Walter K. , Myers Paul Z. , Trevarrowf Bill , Basst Michael B. , Kimmel Charles B.
Published: April 2021

In zebrafish, many nerve pathways in both the CNS and periphery are pioneered by a small and relatively simple set of ‘primary’ neurons that arise in the early embryo. We now have used monoclonal antibodies to show that, as they develop, primary neurons of several functional classes express on their surfaces the L2/HNK-1 tetrasac-charide that is associated with a variety of cell surface adhesion molecules. We have studied the early labeling patterns of these neurons, as well as some non-neural cells, and found that the time of onset and intensity of immunolabeling vary specifically according to cell type.

The first neuronal expression is by Rohon-Beard and trigeminal ganglion neurons, both of which are primary sensory neurons that mediate touch sensitivity. These cells express the epitope very strongly on their growth cones and axons, permitting study of their development unobscured by labeling in other cells. Both types initiate axogenesis at the same early time, and appear to be the first neurons in the embryo to do so. Their peripheral neurites display similar branching patterns and have similar distinctive growth cone morphologies. Their central axons grow at the same rate along the same longitudinal fiber pathway, but in opposite directions, and where they meet they appear to fasciculate with one another. The similarities suggest that Rohon-Beard and trigeminal ganglion neurons, despite their different positions, share a common program of early development.

Immunolabeling is also specifically present on a region of the brain surface where the newly arriving trigeminal sensory axons will enter the brain. Further, the trigeminal expression of the antigen persists in growth cones during the time that they contact an individually identified central target neuron, the Mauthner cell, which also expresses the epitope. These findings provide descriptive evidence for possible roles of L2/HNK-1 immunoreactive molecules in axonal growth and synaptogenesis.
Ca2+-activated ATPase of the mouse chorioallantoic placenta: developmental expression, characterization and cytohistochemical localization
Authors: Tuan Rocky S. , Bigioni Neil
Published: April 2021

A membrane-associated, Ca2+-activated, Mg2+-dependent ATPase activity has been identified in the mouse chorioallantoic placenta. The enzyme activity is expressed and increases as a function of gestation. Bio-chemical characterization shows that the enzyme is highly specific for Ca2+ and nucleotide triphosphates, with a Km of 0.97 mM [Ca2+] and a Vmax of 1.05 nmol Pt released mg-1 placental protein min-1. The mouse placental Ca2+-ATPase activity has a pl of approximately 6.8, and corresponds to two apparent Mr values of 118 and ISOxlO3, based on Ferguson analysis of non-denaturing electrophoretograms. Enzyme activity is inhibited by phenothiazin (suggesting a calmodulin dependence), vanadate, erythrosin B and quercetin, but not by ouabain or levamisole. Enzyme cytohistochemistry revealed that the Ca2+-ATPase is localized to’polyploid trophoblastic cells of the mouse inner placenta. These results suggest that the enzyme may be a functional component of transplacental calcium transport during mouse embryonic development.
Primordial germ cells in the mouse embryo during gastrulation
Authors: Ginsburg Malka , Snow Michael H. L. , McLaren Anne
Published: April 2021

With the aid of a whole-mount technique, we have detected a small cluster of alkaline phosphatase (ALP)-positive cells in whole mounts of mid-primitive-streakstage embryos, days post coitum (dpc). Within the cluster, about 8 cells contain a small cytoplasmic spot, intensely stained for ALP activity and possibly associated with an active Golgi complex. The cluster lies just posterior to the definitive primitive streak in the extraembryonic mesoderm, separated from the embryo by the amniotic fold. Towards the end of gastrulation, the number of cells containing the ALP-positive spot rises to between 50 and 80. Thereafter the number of cells in the extraembryonic cluster declines, and similar cells start to be seen in the mesoderm of the primitive streak and then in the endoderm. At 8 dpc, about 125 ALP-stained cells are found, mainly in the hindgut endoderm and also at the base of the allantois, their appearance and location at this stage agreeing closely with previous reports on primordial germ cells (PGCs). Embryos from which the cluster area has been removed at the 7-day stage are devoid of PGCs after culture for 48 h, whereas the excised tissue is rich in PGCs. We argue that the cells in the cluster are indeed primordial germ cells, at a stage significantly earlier than any reported previously. This would indicate that the PGC lineage in the mouse is set aside at least as early as 7 dpc, possibly as one of the first ‘mesodermal’ cell types to emerge, and that its differentiation, as expressed by ALP activity, is gradual.
Two alkaline phosphatase genes are expressed during early development in the mouse embryo
Authors: Hahnel Ann C. , Rappolee Daniel A. , Millan Jose Luis , Manes Thomas , Ziomek Carol A. , Theodosiou Nicoleta G. , Werb Zena , Pedersen Roger A. , Schultz Gilbert A.
Published: April 2021

Alkaline phosphatase (AP) activity is stage specific in mouse embryos and may be associated with compaction and separation of trophectoderm from inner cell mass in preimplantation development. We previously sequenced a cDNA and two mouse AP genes that could contribute to the AP activity in embryos. Oligonucleotide primers were constructed from the three sequences and used in the reverse transcription-polymerase chain reaction technique to establish that two of the three AP isozymes are transcribed during preimplantation development. The predominant transcript (E-AP) is from a gene highly homologous to the human tissue-specific APs, but different from the mouse intestinal AP. Tissue non-specific (TN) AP also is transcribed, but there is approximately 10 times less TN-AP than E-AP transcript. The TN-AP isozyme is the predominant transcript of 7 to 14 day embryos and primordial germ cells. A switch in predominance from E-AP to TN-AP must occur during early postimplantation development. This study establishes a framework for experiments to determine the functions of the two isozymes during preimplantation development.
Ontogenetic transition of wound healing pattern in rat skin occurring at the fetal stage
Authors: Ihara Setsunosuke , Motobayashi Yumiko , Nagao Eriko , Kistler Andreas
Published: April 2021

Full-thickness excisional wounds were made in the dorsal skin of rat fetuses at day 16 and day 18 of gestation. A small patch of skin surrounding the open wound was cut out, mounted on a plastic ring and incubated in an organ culture system. In the presence of serum, the open wound in the day-16 fetal skin closed within three days of culture. During the wound-closure process, no new structures were formed in the wound space, and no conspicuous changes were noted in the histological architecture of the surrounding skin during culture, indicating that the wound closure may result from a centripetal movement of the surrounding skin only. In contrast, the size of the open wound in the day- 18 fetal skin remained almost unchanged for one week, but a thin acellular network spread over the wound space within one day of culture. The predominant component of the network was cross-linked fibrin, as disclosed by scanning electron microscopy and sodium dodecylsulfate-polyacrylamide gel electrophoresis followed by immunoblotting. The network served as a scaffold for the ingrowth of fibroblast-like cells. These stage-dependent differences in fetal wound healing were consistent with an in vivo study showing that the day-16 wound was covered with the surrounding skin itself, whereas the day-18 wound was covered with newly formed epidermis and invaded by inflammatory cells. The present investigation strongly indicates the prenatal occurrence of a fetal-to-adult transition in the woundhealing pattern of rat skin.
Expression of a surface epitope on cells that link branches in the tracheal network of Manduca sexta
Authors: Nardi James B.
Published: April 2021

A monoclonal antibody (MAb 2F5) to a cell surface epitope labels a small subpopulation of tracheal epithelial cells in each thoracic and abdominal segment of Manduca. These cells (nodes) represent the sites within the tracheal network at which invaginating tracheal tubes join during embryonic establishment of the tracheal network. Tracheal nodes are also the sites at which tracheal cuticle fractures during each molt. Since tracheal cuticle is shed through each spiracle, a tracheal node lies between each pair of contralateral spiracles within a segment (commissural node) and between each pair of adjacent, ipsilateral spiracles (lateral longitudinal node). MAb 2F5 first labels presumptive nodal cells of tracheal epithelium immediately prior to the linking of epithelial tubes from successive and opposite spiracles. One cell at the tip of each invaginating tracheal branch labels with MAb 2F5. The highly localized expression of the cell surface epitope recognized by MAb 2F5 may be instrumental in the orderly coupling of tracheal branches during embryonic development. On the basis of immunolabeling of Western blots and tissues, MAb 2F5 is believed to recognize Manduca fasciclin II, a member of a class of molecules involved in cell adhesion/recognition.
Concurrent reduction in the sulfation of heparan sulfate and basement membrane assembly in a cell model system
Authors: Brauer Philip R. , Keller Katharyn M. , Keller John M.
Published: April 2021

Basement membranes (BMs) are specialized extracellular matrices that have important roles in cell attachment, migration, growth and differentiation. The murine teratocarcinoma cell line, M1536-B3, has been shown to produce a model BM composed of laminin, entactin and heparan sulfate proteoglycans but lacking collagen. Therefore, M1536-B3 cells are an excellent model system in which to study the role of non-collagenous components in BM assembly. We have used these cells to test for a requirement of mature heparan sulfate (HS) chains in BM assembly. Growth of Ml536-B3 cells in the presence of chlorate, an inhibitor of activated sulfate synthesis, resulted in a dose-dependent decrease in the sulfation of glycosaminoglycans and reduction in the charge density of the isolated HS. The undersulfated HS from chlorate-treated cells had a decreased binding capacity for laminin when compared with control HS. Concurrent with these changes in sulfation, chlorate treatment of M1536-B3 cells resulted in the failure of BM assembly, which was restored upon removal of the chlorate from the growth medium. These results were not due to major alterations in cell attachment, spreading, growth, protein synthesis, or to an inability of the cells to synthesize and secrete laminin. These data suggest that the sulfation of HS and its subsequent ability to interact with other BM components play major roles in the assembly and structure of BMs.
Inhibition of proteoglycan synthesis eliminates left-right asymmetry in Xenopus laevis cardiac looping
Authors: Yost H. Joseph
Published: April 2021

The heart of any vertebrate is formed from an apparently symmetric cardiac tube that loops consistently in the same direction along the left-right axis of the embryo. In the amphibian Xenopus laevis, inhibition of proteoglycan synthesis by p-nitrophenyl-β-D-xylopyranoside during a narrow period of development from late gastrula to early neurula specifically eliminated the looping of the cardiac tube. Most of the proteoglycans synthesized during this period were heparan sulfate proteoglycans. Treatment with p-nitrophenyl-α-D-xylopyranoside, an analogue that does not inhibit proteoglycan synthesis, did not interfere with cardiac looping. The critical period for proteoglycan synthesis was coincident with the migration of cardiac primordia to the ventral midline. The inhibition of cardiac looping was further explored in expiants of cardiac primordia and anterioventral ectoderm. In recombinate embryos in which half the embryo, and thus one of the two heart primordia, was treated with p-nitrophenyl-β-D-xylopyranoside, and the other half was untreated, cardiac looping occurred normally. It is proposed that the left-right axis in Xenopus, as reflected in cardiac looping, is established early in development, and that proteoglycan synthesis is involved in the transduction of left-right axial information to the cardiac primordia during migration.