bluuebunny/crossref_metadata_2025 huggingface dataset:
The fate of medial edge epithelial cells during palatal fusion in vitro: an analysis by Dil labelling and confocal microscopy
Authors: Carette Martin J. M. , Ferguson Mark W. J.
Published: April 2021

Fusion of bilateral shelves, to form the definitive mammalian secondary palate, is critically dependent on removal of the medial edge cells that constitute the midline epithelial seam. Conflicting views suggest that programmed apoptotic death or epithelial-mesenchymal transformation of these cells is predominantly involved. Due in part to the potentially ambiguous interpretation of static images and the notable absence of fate mapping studies, the process by which this is achieved has, however, remained mechanistically equivocal.

Using an in vitro mouse model, we have selectively labelled palatal epithelia with Dil and examined the fate of medial edge epithelial (MEE) cells during palatal fusion by localisation using a combination of conventional histology and confocal laser scanning microscopy (CLSM). In dynamic studies using CLSM, we have made repetitive observations of the same palatal cultures in time-course investigations.

Our results concurred with the established morphological criteria of seam degeneration; however, they provided no evidence of MEE cell death or transformation. Instead we report that MEE cells migrate nasally and orally out of the seam and are recruited into, and constitute, epithelial triangles on both the oral and nasal aspects of the palate. Subsequently these cells become incorporated into the oral and nasal epithelia on the surface of the palate. We hypothesize an alternative method of seam degeneration in vivo which largely conserves the MEE population by recruiting it into the nasal and oral epithelia.
Characterisation of dystrophin during development of human skeletal muscle
Authors: Clerk Angela , Strong Peter N. , Sewry Caroline A.
Published: April 2021

Dystrophin, the 427 × 103Mr product of the Duchenne muscular dystrophy (DMD) gene, was studied in human foetal skeletal muscle from 9 to 26 weeks of gestation. Dystrophin could be detected from at least 9 weeks of gestation at the sarcolemmal membrane of most myotubes, though there was differential staining with antibodies raised to various regions of the protein. Dystrophin immunostaining increased and became more uniform with age and by 26 weeks of gestation there was intense sarcolemmal staining of all myotubes. On a Western blot, a doublet of smaller relative molecular mass than that seen in adult tissue was detected in all foetuses studied. There was a gradual increase in abundance of the upper band from 9 to 26 weeks, and the lower band, although present in low amounts in young foetuses, increased significantly between 20 and 26 weeks of gestation. These data indicate that there are several specific isoforms of dystrophin present in developing skeletal muscle, though the role of these is unknown.
Hair follicle differentiation: expression, structure and evolutionary conservation of the hair type II keratin intermediate filament gene family
Authors: Powell Barry , Crocker Lesley , Rogers George
Published: April 2021

During hair follicle development several cell streams are programmed to differentiate from the cell population of the follicle bulb. In the hair cells, a number of keratin gene families are transcriptionally activated. We describe the characterization of the type II keratin intermediate filament (IF) gene family which is expressed early in follicle differentiation. In sheep wool, four type II IF proteins are expressed. One gene has been completely sequenced and the expression of three of the genes examined in detail. The sequenced gene encodes a 55 × 103Mr protein of the type II keratin IF protein family, designated KU-9 in the new nomenclature we have adopted and described in the Introduction. The gene has a similar exon/intron structure to the epidermal type II keratin IF genes. In situ hybridization experiments show that the genes are expressed in the hair cortical cells but not in the cells of the outer root sheath, inner root sheath or medulla. During hair keratinocyte differentiation the type II IF genes are sequentially activated and coexpressed in the same cells. Expression is first detected in cells in the middle of the follicle bulb located near the dermal papilla and, subsequently, two of the genes are transcriptionally activated in the differentiating keratinocytes as they migrate upwards, in the upper part of the bulb. A fourth type II IF gene is activated later. The genes with the same expression pattern are also closely related in sequence and a number of conserved elements are present in the promoters of those genes, including a novel element which is also found in the promoter of a coexpressed type I IF gene and three other hair keratin genes.
U-cadherin in Xenopus oogenesis and oocyte maturation
Authors: Müller A. H. J. , Angres B. , Hausen P.
Published: April 2021

U-cadherin is a member of the cadherin family in Xenopus that participates in interblastomere adhesion in the early embryo from the first cleavage onwards. Though a maternal pool of U-cadherin is available in the egg, it is not present on the egg membrane (Angres et al., 1991. Development 111, 829-844). To assess the origin of this unexpected distribution in the egg, the accumulation and localization of the cadherin during oogenesis and oocyte maturation were investigated.

We report here that U-cadherin is present in Xenopus oocytes throughout oogenesis. It is localized at the oocyte-follicle cell contacts suggesting that it functions in the adhesion of the two cell types.

When oocytes mature and the contacts to the follicle cells break, U-cadherin disappears from the oocyte surface. Evidence for a translocation of U-cadherin from the membrane to the inside of the oocyte was obtained when the fate of membrane-bound U-cadherin, which was labelled on the surface of oocytes prior to maturation, was followed through maturation.

The total U-cadherin content of the oocyte increases during maturation. Metabolic labelling experiments indicate that at maturation the translation of U-cadherin is elevated well above the level that one would expect from the general increase in protein synthesis occurring during maturation. This enhanced synthesis is presumably the main source of the maternal pool of U-cadherin in the egg.
Localized enhancement and repression of the activity of the TGF-P family member, decapentaplegic, is necessary for dorsal-ventral pattern formation in the Drosophila embryo
Authors: Ferguson Edwin L. , Anderson Kathryn V.
Published: April 2021

Seven zygotic.ally active genes are required for normal patterning of the dorsal 40% of the Drosophila embryo. Among these genes, decapentaplegic (dpp) has the strongest mutant phenotype: in the absence of dpp, all cells in the dorsal and dorsolateral regions of the embryo adopt fates characteristic of more ventrally derived cells (Irish and Gelbart (1987)Genes Dev. 1, 868-879). Here we describe the phenotypes caused by alleles of another of this set of genes, tolloid, and show that tolloid is required for dorsal, but not dorsolateral, pattern. Extragenlc suppressors of tolloid mutations were isolated that proved to be mutations that elevate dpp activity. We studied the relationship between tolloid and dpp by analyzing the phenotypes of tolloid embryos with elevated numbers of the dpp gene and found that doubling the dpp+ gene dosage completely suppressed weak tolloid mutants and partially suppressed the phenotypes of tolloid null mutants. We conclude that the function of tolloid is to increase dpp activity. We also examined the effect of doubling dpp+ gene dosage on the phenotypes caused by other mutations affecting dorsal development. Like tolloid, the phenotypes of mutant embryos lacking shrew gene function were suppressed by elevated dpp, indicating that shrew also acts upstream of dpp to increase dpp activity. In contrast, increasing the number of copies of the dpp gene enhanced the short gastrulation (sog) mutant phenotype, causing ventrola teral cells to adopt dorsal fates. This indicates that sog gene product normally blocks dpp activity ventrally. We propose that the tolloid, shrew and sog genes are required to generate a gradient of dpp activity, which directly specifies the pattern of the dorsal 40% of the embryo.
Cholinergic neuronal differentiation factors: evidence for the presence of both CNTF-like and non-CNTF-like factors in developing rat footpad
Authors: Rohrer H.
Published: April 2021

Catecholaminergic sympathetic neurons are able to change their transmitter phenotype during development and to acquire cholinergic properties. Cholinergic sympathetic differentiation is only observed in fibers innervating specific targets like the sweat glands in the rat footpad. A function for ciliary neurotrophic factor (CNTF) in this process has been implied as it is able to induce cholinergic properties (ChAT,VIP) in cultured chick and rat neurons. We show here that a CNTF-like, VIP-inducing activity is present in rat footpads and that it increases 6-fold during the period of cholinergic sympathetic differentiation. Immunohistochemical analysis of P21 rat footpads demonstrated CNTF-like immunoreactivity in Schwann cells but not in sweat glands, the target tissue of cholinergic sympathetic neurons. The expression of this factor in footpads seems to be dependent on the presence of intact nerve axons, as nerve transection results in a loss of CNTF-like cholinergic activity and immunoreactivity. Immunoprecipitation experiments with rat footpad extracts provided evidence for the presence of ChAT-inducing factors other than CNTF, which may independently or together with CNTF be involved in the determination of sympathetic neuron phenotype.
Transforming growth factor-alpha (TGF-α) and insulin gene expression in human fetal pancreas
Authors: Miettine Pätvi J. , Heikinheimo Kristiina
Published: April 2021

Transforming growth factor-alpha (TGF-α) mRNA is expressed in several pancreatic cancer cell lines, but its expression during normal fetal pancreas development has not been studied. We investigated the expression of TGF-α, its receptor (EGF-R) and insulin mRNA and their corresponding peptides in human fetal pancreata (15–20 gestation weeks). Polymerase chain reaction (PCR) and RNAase protection analysis revealed that TGF-α and insulin mRNAs were detectable in pancreas during the developmental span studied. In northern blot analysis a single band of 4.8 kilobases (kb) corresponding to the TGF-α transcript and a 0.6 kb for the insulin mRNA were detected in the pancreas. Using in situ hybridization, TGF-α mRNA expression was seen in a low copy number in both the exo- and endocrine pancreas. By immunohistochemistry TGF-α-immuno-reactive cells were detected in the ducts, acini and islets showing that the mRNA was translated into protein. By contrast, insulin transcripts were detected in a high copy number, restricted to the islets of Langerhans. However, monoclonal insulin antibody detected less insulin containing cells than could be expected from the mRNA pattern suggesting that fetal β-cells rapidly secrete insulin instead of storing it in the secretory granules. Alternatively, the translation of insulin mRNA could be inefficient. By double labeling the pancreas sections with polyclonal TGF-α antiserum and monoclonal insulin antibody the TGF-α- and insulin-like immunoreactivity was localized to β-cells. Furthermore, mRNA for the TGF-α receptor, EGF-R, together with EGF-R-immu-noreactive cells were also present in pancreas. The results suggest that TGF-α may participate auto- and/or paracrinically in the development of human fetal pancreas.
Function of an Ultrabithorax minigene in imaginal cells
Authors: Castelli-Gair James , Müller Jürg , Bienz Mariann
Published: April 2021

An Ultrabithorax (Ubx) minigene constructed from three key Ubx control regions is capable of supporting development of Ubx null mutants throughout larval life and beyond to pharate flies, thereby rescuing the larval lethality due to the homeotic mutation. The cuticle of these flies shows that the minigene provides at least partial Ubx function in each of the four compartments whose morphogenetic pathways are determined by Ubx. We analyse β-galactosidase patterns in imaginal discs conferred by each individual Ubx control region. From the comparison of these patterns with Ubx expression in Cbx mutants, we infer that long-range repressor elements in the chromosomal Ubx gene play an important role in the generation of Ubx expression patterns in imaginal discs. Expression and function of our Ubx minigenes indicate that Ubx control regions are capable of functioning properly out of context and detached from their normal chromosomal location within the homeotic gene complex.
Diffusible rod-promoting signals in the developing rat retina
Authors: Watanabe Takashi , Raff Martin C.
Published: April 2021

We previously developed a reaggregate cell culture system in which embryonic rat retinal neuroepithelial cells proliferate and give rise to opsin-expressing rod photoreceptor cells (rods) on the same schedule in vitro as they do in vivo. We showed that the proportion of neuroepithelial cells in the embryonic day 15 (E15) retina that differentiated into opsin+ rods after 5 –6 days in such cultures increased by ∼40-fold when the E15 cells were cultured in the presence of an excess of postnatal day 1 (Pl) neural retinal cells. In the present study, we have further analyzed this rod-promoting activity of neonatal neural retinal cells. We show that the activity is mediated by a diffusible signal(s) that seems to act over a relatively short distance. Whereas neonatal (P1-P3) neural retina has rod-promoting activity, E15 and adult neural retina, neonatal thymus, cerebrum and cerebellum do not. Finally, we show that neonatal neural retina promotes rod but not amacrine cell development.
Regeneration of dopaminergic neurons in goldfish retina
Authors: Braisted Janet E. , Raymond Pamela A.
Published: April 2021

The conditions necessary to trigger regeneration of dopaminergic neurons were investigated in the goldfish retina. Intraocular injection of 6-hydroxydopamine (6-OHDA) was used to destroy dopaminergic neurons, and neuronal regeneration was monitored by injections of the thymidine analog bromodeoxyuridine (BUdR). Regenerated dopaminergic neurons, (identified by doublelabeling with anti-tyrosine hydroxylase and anti-BUdR antibodies) were found within 3 weeks after 2 injections of 0.6 mg/ml 6-OHDA (estimated intraocular concentration), but not after injection of lower doses. All retinas with regenerated dopaminergic neurons also contained other types of regenerated neurons, including cones and ganglion cells, consistent with nuclear counts which revealed non-selective cell loss (34 –36%) in both the outer and inner nuclear layers after exposure to the high dose, but not lower doses of 6-OHDA. Regenerated neurons were produced by clusters of dividing neuroepithelial cells probably derived from rod precursors in the outer nuclear layer. These results demonstrate that dopaminergic neurons will not regenerate after they are selectively ablated but only as part of a developmental process that involves generation of multiple cell types.
Evidence of a role for endogenous electrical fields in chick embryo development
Authors: Hotary Kevin B. , Robinson Kenneth R.
Published: April 2021

We have tested directly the hypothesis that the endogenous electrical field in the chick embryo plays a causal role in development. Conductive implants, which shunt currents out of the embryo and thus alter the internal held, were placed under the dorsal skin at the mid-trunk level of stage 11 – 15 embryos. Currents leaving the posterior intestinal portal (p.i.p.) of these embryos were reduced by an average of 30%. Control embryos receiving non-conductive implants showed no change in p.i.p. currents. In the group receiving current shunts, 92% of the embryos exhibited some developmental abnormality. Only 11% of the control group displayed defects. The most common defect in the experimental group (81%) was in tail development. Tail defects ranged from complete absence to the formation of a normal length, but morphologically abnormal tail. Internally, tail structures (neural tube, notochord and somites) were frequently absent or aberrantly formed. In 33% of the experimental embryos, the notochord continued lengthening in the absence of any other tail development. This led to the formation of ourenteric outgrowths from the hindgut. Defects in limb bud and head development were also found in experimentally treated embryos, but at a much lower frequency than tail defects.

The abnormalities observed in experimental embryos were very similar to those produced naturally in rumpless mutant chicks. A vibrating probe analysis of these mutants (from both dominant and recessive strains) showed that currents leaving the p.i.p. were significantly lower in phenotypically abnormal mutants than in wild-type and phenotypically normal mutant embryos from both strains. There was no apparent correlation between the average transepithelial potential (TEP) of these mutants and the development of tail abnormalities. The possible role of endogenous electrical fields in chick tail development is discussed.
Expression and functional involvement of N-cadherin in embryonic limb chondrogenesis
Authors: Oberlender Steven A. , Tuan Rocky S.
Published: April 2021

Cell adhesion molecules have been shown to be important mediators of morphogenesis and pattern formation. In this study, we have shown that N-cadherin is expressed in a specific spatiotemporal manner in the developing limb bud during chondrogenesis in vivo and in cultured limb mesenchyme in vitro. The time period of maximal expression of N-cadherin corresponds to the period of active cellular condensation, an event believed to be a necessary prerequisite for chondrogenic differentiation. To directly assess the functional involvement of N-cadherin in cellular condensation, we have examined the effects of perturbing N-cadherin activity on both cell aggregation and chondrogenesis using NCD-2, a rat monoclonal antibody directed against the binding region of N-cadherin. Non-immune rat IgG was used as a control. Our results show that functional N-cadherin is necessary for chondrogenesis to proceed both in vivo and in vitro. Limb mesenchymal cells exhibited characteristic Ca2+-dependent cell aggregation in suspension, which was inhibited in the presence of exogenous NCD-2. In micromass cultures of limb mesenchymal cells, NCD-2 inhibited overt chondrogenesis in a dose-dependent manner. Furthermore, NCD-2 inhibition of chondrogenesis in micromass cultures was time-dependent, suggesting that N-cadherin is crucially involved during the latter half of the first 24 hours of culture, a time period most likely corresponding to active cellular condensation. NCD-2 also significantly influenced limb development when injected into embryonic limb buds in vivo. In addition to significant inhibition of chondrogenesis and developmental delays, gross developmental deformities and perturbation of overall pattern formation were also observed. Taken together, these results demonstrate that N-cadherin is functionally required in mediating the cell-cell interactions among mesenchymal cells important for chondrogenesis in micromass culture in vitro and in the intact limb bud in vivo.
engrailed-mediated repression of Ultrabithorax is necessary for the parasegment 6 identity in Drosophila
Authors: Mann Richard S.
Published: April 2021

The homeotic genes of Drosophila are expressed in overlapping domains along the anterior-to-posterior axis and specify the distinct morphological patterns of each parasegment. Within single parasegments, the levels of homeotic gene expression are often modulated, in part because of cross-regulation by other homeotic gene products. However, the functional significance of different levels of homeotic gene expression is unclear. Here modulations in Ultrabithorax (Ubx) expression within parasegment 6 are examined. Specifically, Ubx is shown to be down-regulated in the posterior compartment of this parasegment by engrailed (en). The significance of Ubx repression by en was demonstrated by characterizing the expression of the Ubx target gene, Distal-less (Dll). In the posterior compartment of parasegment 6, Dll is normally expressed in a small cluster of cells. If Ubx is expressed uniformly via a heatshock promoter, Dll is inappropriately repressed in these posterior compartment cells. In the anterior compartment of parasegment 6, Dll is normally repressed by high levels of Ubx. However, if en is expressed uniformly via a heatshock promoter, Ubx is repressed and Dll is derepressed. Because Dll is required for the development of larval sensory structures, these results demonstrate that en-mediated repression of Ubx in the posterior compartment is necessary for the morphology of parasegment 6. Thus, different levels of homeotic gene expression can be important for their segmental patterning functions.
GATA factor activity is required for the trophoblast-specific transcriptional regulation of the mouse placental lactogen I gene
Authors: Ng Yuk-Kiu , George Kathleen M. , Engel James Douglas , Linzer Daniel I. H.
Published: April 2021

The molecular determinants governing tissue-specific gene expression in the placenta are at present only poorly defined, particularly with respect to the regulation of specific hormone genes whose products are vital to embryonic development and the maintenance of a nurturing maternal environment. In continuing our analysis of the trophoblast-specific expression of the mouse placental lactogen I gene, we now demonstrate that the transcription factors GATA-2 and GATA-3 regulate the activity of this gene promoter. These factors are expressed in placental trophoblast cells, with peak levels of the GATA-2, GATA-3 and placental lactogen I mRNAs each accumulating at midgestation. Analysis of a region of the placental lactogen I gene promoter, previously shown to be sufficient for directing trophoblast-specific transcription, revealed the presence of three consensus binding sites for GATA-2 or GATA-3. Both GATA-2 and GATA-3 bind to these sites in vitro and mutation of these sites results in a significant decrease in promoter activity as assayed by transient transfection into the choriocarcinoma-derived cell line Rcho-1, which expresses endogenous GATA-2 and GATA-3. Furthermore, overexpression of GATA factors in Rcho-1 cells stimulates transcription from a co-transfected placental lactogen I gene promoter. Most significantly, expression of GATA-2 or GATA-3 was found to induce transcription from this promoter in transfected non-trophoblast (fibroblast) cells. These data indicate that GATA factors are both limiting and required transcriptional regulatory molecules in placental trophoblasts, and that the tissue specificity of the placental lactogen I gene is determined, at least in part, by GATA-2 and/or GATA-3.
Differential expression of Broad-Complex transcription factors may forecast tissue-specific developmental fates during Drosophila metamorphosis
Authors: Emery Ivette F. , Bedian Vahe , Guild Gregory M.
Published: April 2021

The steroid hormone ecdysone initiates metamorphosis in Drosophila melanogaster by activating a cascade of gene activity that includes primary response transcriptional regulators and secondary response structural genes. The Broad-Complex (BR-C) primary response gene is composed of several distinct genetic functions and encodes a family of related transcription factor isoforms. Our objective was to determine whether BR-C isoforms were components of the primary ecdysone response in all tissues and whether tissue-specific isoform expression is associated with tissue-specific metamorphic outcomes. We used specific antibody reagents that recognize and distinguish among the Z1, Z2 and Z3 BR-C protein isoforms to study protein expression patterns during the initial stages of metamorphosis. Western blot analyses demonstrated that BR-C isoforms are induced at the onset of metamorphosis, each with unique kinetics of induction and repression. Whole-mount immunostaining showed that the BR-C proteins accumulate in the nuclei of all larval and imaginal tissues indicating that the BR-C is induced as a primary response in many tissues. Several tissues express different levels and combinations of the BR-C isoforms suggesting that the BR-C is important in determining the tissue-specific outcome of many parallel ecdysone response cascades. For example, prepupal salivary glands (destined for histolysis during metamorphosis) express Z1 isoforms while imaginal discs (destined for cell differentiation and morphogenesis) shift from the synthesis of Z2 isoforms to the synthesis of Z1 isoforms. The prepupal central nervous system (destined for tissue remodeling) expresses all isoforms, with Z3 pre-dominating. Salivary gland chromosome immunostaining indicated that BR-C proteins interact directly with numerous loci in the polytene genome. Finally, western blot analyses showed that distinct BR-C genetic functions can be correlated with single and specific BR-C protein isoforms.
Roles of heterotrimeric and monomeric G proteins in sperm-induced activation of mouse eggs
Authors: Moore Grace D. , Ayabe Takuya , Visconti Pablo E. , Schultz Richard M. , Kopf Gregory S.
Published: April 2021

Results of several lines of experimentation suggest that sperm-induced egg activation has several features in common with G protein-coupled receptor signal transduction mechanisms. We report that microinjection of GDPβS into metaphase II-arrested mouse eggs blocks sperm-induced egg activation. Since GDPβS inactivates both het-erotrimeric and monomeric classes of G proteins, the involvement of members of each of these families in sperm-induced egg activation was evaluated. Neither pertussis toxin treatment of eggs nor microinjection of eggs with inhibitory antibodies toward Gαq blocked sperm-induced egg activation. Nevertheless, microinjection of phosducin, a protein that binds tightly to free G protein βγ subunits, specifically inhibited second polar body emission, the fertilization evoked decrease of H1 kinase activity and pronucleus formation. Microinjection of phosducin, however, did not inhibit the fertilization-induced modifications of the zona pellucida and microinjection of βγt did not result in egg activation in the absence of sperm. Inactivation of the monomeric Rho family of G proteins with C3 transferase from Clostridium botulinum inhibited emission of the second polar body and cleavage to the 2-cell stage, but did not affect the modifications of the zona pellucida or pronu-cleus formation. Microinjection of Rasval12, which is a constitutively active form of Ras, did not result in egg activation in the absence of sperm. Moreover, microinjection of either an anti-Ras neutralizing antibody (Y13-259) or a dominant negative form of Ras (RasT) did not affect events of sperm-induced egg activation. In contrast, microinjection of RasT inhibited embryo cleavage to the 2-cell stage. These results suggest that both heterotrimeric and monomeric G proteins are involved in various aspects of sperm-induced egg activation.
Combinatorial specification of blastomere identity by glp-1-dependent cellular interactions in the nematode Caenorhabditis elegans
Authors: Moskowitz Ivan P. G. , Gendreau Steven B. , Rothman Joel H.
Published: April 2021

Most somatic cells in the nematode Caenorhabditis elegans arise from AB, the anterior blastomere of the 2-cell embryo. While the daughters of AB, ABa and ABp, are equivalent in potential at birth, they adopt different fates as a result of their unique positions. One such difference is that the distribution of epidermal precursors arising from ABp is reversed along the anterior-posterior axis relative to those arising from ABa. We have found that a strong mutation in the glp-1 gene eliminates this ABa/ABp difference. Furthermore, extensive cell lineage analyses showed that ABp adopts an ABa-like fate in this mutant. This suggests that glp-1 acts in a cellular interaction that makes ABp distinct from ABa. One ABp-specific cell type was previously shown to be induced by an interaction with a neighboring cell, P2. By removing P2 from early embryos, we have found that the widespread differences between ABa and ABp arise from induction of the entire ABp fate by P2. Lineage analyses of genetically and physically manipulated embryos further suggest that the identities of the AB greatgranddaughters (AB8 cells) are controlled by three regulatory inputs that act in various combinations. These inputs are: (1) induction of the ABp-specific fate by P2, (2) a previously described induction of particular AB8 cells by a cell called MS, and (3) a process that controls whether an AB8 cell is an epidermal precursor in the absence of either induction. When an AB8 cell is caused to receive a new combination of these regulatory inputs, its lineage pattern is transformed to resemble the lineage of the wild-type AB8 cell normally receiving that combination of inputs. These lineage patterns are faithfully reproduced irrespective of position in the embryo, suggesting that each combination of regulatory inputs directs a unique lineage program that is intrinsic to each AB8 cell.
Wingless can bring about a mesoderm-to-ectoderm induction in Drosophila embryos
Authors: Lawrence Peter A. , Johnston Paul , Vincent Jean-Paul
Published: April 2021

By means of nuclear transplantations, we make mosaics in which largely wingless− embryos contain patches of wingless+ cells. In these genetic mosaics, using a standard assay for wingless function (the maintenance of engrailed expression), we uncover an induction across germ layers: Wingless made in the mesoderm can sustain engrailed expression in the ectoderm. This result makes clear that Wingless is expressed in the mesoderm until at least one hour after gastrulation and may function in this germ layer in the wild type.
Cardiovascular development in the zebrafish: II. Endocardial progenitors are sequestered within the heart field
Authors: Lee Robert K. K. , Stainier Didier Y. R. , Weinstein Brant M. , Fishman Mark C.
Published: April 2021

We have examined the zebrafish embryo to ascertain the location of endocardial and myocardial progenitors prior to gastrulation, in an attempt to define the earliest stages of cardiac patterning. Currently there is uncertainty as to the spatial and lineage relationship of the progenitors for these two phenotypically distinct cell types that form the two concentric layers of the primitive heart tube. By single-cell injection and tracking, we distinguish a region in the early and midblastula which has the properties of a heart field, in that it defines a zone of cardiac progenitors within which there is a spatial gradient of propensity to generate heart cells, and which regulates, in the sense of adapting to the transplantation of pluripotential cells. This zone extends from the future ventral axis dorsally along the margin, with cardiogenic propensity tapering off laterally and dorsally. Myocardial progenitors are spread through-out this region, but endocardial precursors are restricted to the ventral marginal region. The cardiovascular progeny of the ventral cells include, in addition to endocardium and myocardium, cells in the endothelium and blood.
Micromere fate maps in leech embryos: lineage-specific differences in rates of cell proliferation
Authors: Smith Constance M. , Weisblat David A.
Published: April 2021

Stereotyped early cleavages in glossiphoniid leech embryos yield 25 micromeres, along with 3 macromeres and 10 teloblasts. The micromeres generate prostomial tissues and also give rise to most of the squamous epithelium of a provisional integument that spreads epibolically from the animal pole, covering the rest of the embryo during germinal plate formation.

We systematically injected individual micromeres with fluorescent cell lineage tracers at the time of their birth and quantitatively mapped the contributions of all these cells to the late stage 7 embryo, a time in development that is early in the epibolic expansion. At this time, micromere derivatives comprise two types of cells: squamous epithelial (superficial) cells that cover the germinal bands and the region of the animal cap between the germinal bands; and underlying (deep) cells that are confined to the distal ends of the germinal bands and in the area between their distal ends. We find that individual micromeres contribute clones of deep and/or superficial progeny that are stereotyped with respect to both numbers and types of cells in the clone and the domains that they occupy. The N teloblasts also contribute cells to the squamous epithelium.

We find significant differences in the rate of cell proliferation between different micromere clones. These differences appear to reflect lineage-specific traits, since there is little or no regulation of cell number after ablation of individual micromeres.
Sequential fates in a single cell are established by the neurogenic cascade in the Malpighian tubules of Drosophila
Authors: Hoch Michael , Broadie Kendal , Jäckle Herbert , Skaer Helen
Published: April 2021

In each Malpighian tubule of Drosophila, one cell is singled out, the tip cell, whose function during embryogenesis is to promote cell division in its neighbours. We follow the seg-regation of this cell, explore the genetic interactions that underlie its specification and demonstrate that tip cell allocation closely resembles neurogenesis. The tip cell arises by division of a tip mother cell, which is selected from a cluster of equivalent cells in each tubule primordium. Each cluster is marked out by the expression of proneural genes and the selection of a single cell from each group involves lateral inhibition, mediated by the neurogenic genes. We confirm the mitogenic role of the tip cell during embryogenesis by mutational analysis and show that it subsequently adopts a second fate, differentiating neural characteristics. We demonstrate that both stages in the differentiation of this cell are established by the same sequence of genetic inter-actions, which have not previously been shown to occur outside the neurogenic ectoderm.
Mesenchymal regulation of epithelial gene expression in developing avian stomach: 5′-flanking region of pepsinogen gene can mediate mesenchymal influence on its expression
Authors: Fukuda Kimiko , Ishii Yasuo , Saiga Hidetoshi , Shiokawa Koichiro , Yasugi Sadao
Published: April 2021

The expression of a gene encoding an embryonic chick pepsinogen was investigated in developing avian gut. Expression is restricted to the epithelial layer of the embryonic proventriculus (glandular stomach). We can therefore regard this gene as a marker gene for proventricular epithelial differentiation. There is some consider-able evidence in favour of epithelilal-mesenchymal inter-actions being important during the development of the gastrointestinal system; for example, pepsinogen expression is induced in proventricular and gizzard (muscular stomach) epithelia by the proventricular mes-enchyme but is suppressed by the gizzard mesenchyme. In the present paper, we studied how the mesenchymes influence this gene expression pattern. For this we produced constructs containing various portions of the 5′-flanking region of the embryonic chick pepsinogen gene, driving reporter sequences (β-galactocidase or luciferase), and these constructs were transfected into dissociated epithelial cells either from the proventriculus or gizzard. We then recombined these cells with mesenchymal cells and cultured them as cell aggregates. In this way, we were able to dissect the timing and other requirements of the epithelial-mesenchymal interactions for expression of embryonic chick pepsinogen gene. We also report that 1.1 kb of 5′-flanking sequence is sufficient to drive correct expression of embryonic chick pepsinogen gene, although further enhancement was seen if the constructs contained 3.2 kb of upstream sequence.
Control of CNS midline transcription by asymmetric E-box-like elements: similarity to xenobiotic responsive regulation
Authors: Wharton Keith A. , Franks Robert G. , Kasai Yumi , Crews Stephen T.
Published: April 2021

Central nervous system midline cells constitute a discrete group of Drosophila embryonic cells with numerous functional and developmental roles. Corresponding to their separate identity, the midline cells display patterns of gene expression distinct from the lateral central nervous system. A conserved 5 base pair sequence (ACGTG) was identified in central nervous system midline transcriptional enhancers of three genes. Germ-line transformation experiments indicate that this motif forms the core of an element required for central nervous system midline transcription.

The central nervous system midline element is related to the mammalian xenobiotic response element, which regulates transcription of genes that metabolize aromatic hydrocarbons. These data suggest a model whereby related basic-helix-loop-helix-PAS proteins interact with asymmetric E-box-like target sequences to control these disparate processes.
The novel homeodomain gene buttonless specifies differentiation and axonal guidance functions of Drosophila dorsal median cells
Authors: Chiang Chin , Patel Nipam H. , Young Keith E. , Beachy Philip A.
Published: April 2021

We have identified a novel homeodomain gene, buttonless (btn), that is specifically expressed in 20 cells of a single type during Drosophila embryonic development. These cells, the dorsal median (DM) cells, are arranged as a single pair within each segment along the dorsal midline of the CNS. Distinctive features of the DM cells include a large cell body and a long thick process extending laterally to the muscle attachment site. In the absence of btn gene function the initial commitment to the DM cell fate is made but differentiation fails to occur and the DM cells are lost. The btn mutation thus specifically eliminates the DM cells, and this genetic ablation in turn reveals a requirement for DM cells as cellular cues for axonal guidance during transverse nerve outgrowth and bifurcation of the median nerve.
Aprotinin, a Kunitz-type protease inhibitor, stimulates skeletal muscle differentiation
Authors: Wells James M. , Strickland Sidney
Published: April 2021

Aprotinin, a Kunitz-type inhibitor of serine proteases, stimulates myotube formation by mouse G8-1 and C2C12 skeletal muscle myoblasts. This stimulation of morphological differentiation is accompanied by accumulation of myogenin transcripts and production of muscle-specific proteins. In contrast, active TGFβ prevents differentiation of G8-1 and C2C12 myoblasts. When active TGFβ and aprotinin are both added to myoblast cultures, differeniation is inhibited, suggesting the active growth factor acts downstream of the protease inhibitor. TGFβ is found in serum as a latent, dimeric propolypeptide that is cleaved by limited proteolysis to release the biologically active carboxy-terminal dimer. To address the possibility that aprotinin may effect myogenesis by inhibiting proteolytic activation of latent TGFβ, levels of the endogenous growth factor were measured in differentiating myoblast cultures. Latent TGFβ is rapidly depleted from control cultures within 24 hours of plating, but remains relatively stable in aprotinin-treated cultures. Consistent with this, aprotinin-treated cultures have reduced levels of active TGFβ. These data indicate that Kunitz-domain containing protease inhibitors may help orchestrate the onset of myogenesis, possibly by regulating the activity of TGFβ-like molecules.
Identification of a candidate primary sex determination locus, fox-1, on the X chromosome of Caenorhabditis elegans
Authors: Hodgkin Jonathan , Zellan Jonathan D. , Albertson Donna G.
Published: April 2021

Sex in Caenorhabditis elegans (XX hermaphrodite, XO male) is determined by the X:A ratio, which is the ratio of X chromosome number to autosomal set number. Recent genetic results with X chromosome duplications have suggested that there may be only a small number of major numerator sites on the X chromosome that contribute to this ratio. Mapping of duplication endpoints delimited a region of less than 300 kb, likely to contain one such element. Cosmid clones from this region were tested for numerator activity by constructing transgenic lines carrying extra copies of each tested cosmid. Most cosmid arrays have no effect on the viability of either XX or XO animals. One cosmid array was found to be viable in XX animals, but lethal and feminizing in XO animals, consistent with it containing a major numerator element. Further experiments defined a region of 12-30 kb with apparent numerator activity, which is designated fox-1, ‘Feminizing locus On X’. A cDNA clone hybridizing across part of this region encodes a predicted RNA-binding protein.
Expression of peptide YY in all four islet cell types in the developing mouse pancreas suggests a common peptide YY-producing progenitor
Authors: Upchurch Brent H. , Aponte Gregory W. , Leiter Andrew B.
Published: April 2021

The islets of Langerhans contain four distinct endocrine cell types producing the hormones glucagon, insulin, somatostatin and pancreatic polypeptide. These cell lineages are thought to arise from a common, multipotential progenitor cell whose identity has not been well established. The pancreatic and intestinal hormone, peptide YY, has been previously identified in glucagon-producing cells in islets; however, transgenic mice expressing Simian Virus 40 large T antigen under the control of the peptide YY gene expressed the oncoprotein in β, δ and pancreatic polypeptide cells, and occasionally developed insulinomas, suggesting relationships between peptide YY-producing cells and several islet cell lineages. The four established pancreatic islet cell types were examined for coexpression of peptide YY in islets of normal and transgenic mice throughout development. Peptide YY immunoreactivity was identified in the earliest endocrine cells in the fetal pancreas and was coexpressed in each islet cell type during development. Peptide YY showed a high degree of co-localization with glucagon- and insulin-producing cells in early pancreatic development, but by adulthood, peptide YY was expressed in less than half of the α cells and was no longer expressed in β cells. Peptide YY was also coexpressed with somatostatin and pancreatic polypeptide when these cell types first appeared, but most δ and pancreatic polypeptide cells continued to express peptide YY throughout development. The use of conditions that distinguish peptide YY from the related peptides, pancreatic polypeptide and neuropeptide Y, as well as the ability of the peptide YY gene to direct expression of a reporter gene in islets of transgenic mice, establishes expression of peptide YY in the earliest pancreatic endocrine cells. Coexpression of peptide YY in all islet cell types as they first emerge suggests that the four established islet cell types may arise from a common, previously unrecognized peptide YY-producing progenitor cell.
A role for CdxA in gut closure and intestinal epithelia differentiation
Authors: Frumkin Ayala , Pillemer Graciela , Haffner Rebecca , Tarcic Nora , Gruenbaum Yosef , Fainsod Abraham
Published: April 2021

CdxA is a homeobox gene of the caudal type that was previously shown to be expressed in the endoderm-derived gut epithelium during early embryogenesis. Expression of the CDXA protein was studied during intestine morphogenesis from stage 11 (13 somites) to adulthood in the chicken. The CDXA protein can be detected during all stages of gut closure, from stage 11 to 5 days of incubation, and is mainly localized to the intestinal portals, the region where the splanchnopleure is undergoing closure. In this region, which represents the transition between the open and closed gut, the CDXA protein is restricted to the endodermderived epithelium. At about day 5 of incubation, the process of formation of the previllous ridges begins, which marks the beginning of the morphogenesis of the villi. From this stage to day 11 expression of CDXA is localized to the epithelial lining of the intestine. In parallel, a gradual increase in CDXA protein expression begins in the mesenchyme that is close in proximity to the CDXA-positive endoderm. Maximal CDXA levels in the mesenchyme are observed at day 9 of incubation. During days 10 and 11 CDXA levels in the mesenchyme remain constant, and by day 12 CDXA becomes undetectable in these cells and the epithelium again becomes the main site of expression. From day 12 of incubation until adulthood the CDXA protein is present in the intestinal epithelium. Until day 18 of incu-bation expression can be detected along the whole length of the villus with a stronger signal at the tip. With hatching the distribution along the villi changes so that the main site of CDXA protein expression is at the base of the villi and in the crypts. The transient expression of CDXA in the mesenchyme between days 5 and 11 may be related to the interactions taking place between the mesenchyme and the epithelium that ultimately result in the axial specification of the alimentary canal and the differentiation of its various epithelia. The main CDXA spatial distribution during morphogenesis suggests a tight linkage to the formation and differentiation of the intestinal epithelium itself. CDXA appears to play a role in the morphogenetic events leading to closure of the alimentary canal. During previllous ridge formation the CDXA protein is transiently expressed in the mesenchymal cells thought to provide instructive interactions for the regionalization and differentiation of the gut epithelium. Finally, CDXA is expressed, from hatching until adulthood, in the crypts and the base of the villi, in cells on their way to differentiate and replace those aged by digestive activity.
Spatially regulated expression of three receptor tyrosine kinase genes during gastrulation in the zebrafish
Authors: Xu Qiling , Holder Nigel , Patient Roger , Wilson Stephen W.
Published: April 2021

We describe the isolation and early developmental expression of three novel zebrafish genes (rtk1-3) that encode members of the eph family of receptor tyrosine kinases. At the onset of gastrulation, rtk1 is expressed in the shield region corresponding to the future dorsal side of the embryo. As gastrulation proceeds, both rtk1 and rtk2 are expressed within the axial hypoblast along the entire axis of the embryo. After the gastrula stage is complete, expression of both genes is maintained in precursor cells of the notochord in the tail bud but is downregulated in other regions of the axial hypoblast. rtk3 is expressed in anterior axial hypoblast including the ‘pillow’ at the anterior tip of the hypoblast and in paraxial tissue in posterior regions of the embryo.

We show that the precise spatial regulation of expression of rtk genes, ntl and goosecoid along the anteroposterior axis is maintained in embryos that have no dorsoventral axis. This indicates that the mechanisms that regulate gene expression along the anteroposterior and dorsoventral axes of the hypoblast may be independent.