bluuebunny/crossref_metadata_2025 huggingface dataset:
Developmental regulation of creatine kinase gene expression by myogenic factors in embryonic mouse and chick skeletal muscle
Authors: Lyons Gary E. , Mühlebach Stephan , Moser Andrea , Masood Rizwan , Paterson Bruce M. , Buckingham Margaret E. , Perriard Jean-Claude
Published: April 2021

The B isoform of creatine kinase (BCK), which is expressed at a high level in embryonic neural tissues, is also expressed abundantly in developing striated muscle and is an early marker for skeletal myogenesis. Using isoform-specific 35S-labeled antisense cRNA probes for in situ hybridization, we have detected BCK mRNAs in embryonic mouse and chick myotomes, the first skeletal muscle masses to form in developing embryos. These transcripts are detectable, as soon as myotomes are morphologically distinguishable. BCK is expressed at high levels in both skeletal and cardiac muscle in mouse and chick embryos. In the mouse, BCK transcript levels fall off rapidly in striated muscle shortly after the onset of MCK gene expression.

The M isoform of creatine kinase (MCK), the striated muscle-specific isoform, is expressed later than BCK. In the mouse, BCK transcripts are expressed in myotomes at 8.5 days post coitum (p.c.), but MCK transcripts are not detected before 13 days p.c. In the chick, BCK mRNAs are present at Hamburger-Hamilton stage 13, but MCK mRNAs are not detected before stage 19.

We have compared the patterns of expression of the CK genes with those of myogenic differentiation factor genes, which are thought to regulate skeletal musclespecific gene expression. In the chick, both CMD1, first detected at stage 13, and myogenin, first detected at stage 15, are present prior to MCK, which begins to be expressed at stage 19. Unlike the mouse embryo, CMD1, the chick homologue of MyoDl, is expressed before chick myogenin. In the mouse, myogenin, first detected at 8.5 daysp.c., is expressed at the same time as BCK in myotomes. Both myogenin and MyoDl, which begins to be detected two days later than myogenin, are expressed at least two days before MCK.

It has been proposed that the myogenic factors, MyoDl and myogenin, directly regulate MCK gene expression in the mouse by binding to its enhancer. However, our results show that MCK transcripts are not detected until well after MyoDl and myogenin mRNAs are expressed, suggesting that these factors by themselves are not sufficient to initiate MCK gene expression.
Biologically active kit ligand growth factor is produced by mouse Sertoli cells and is defective in Sld mutant mice
Authors: Tajima Youichi , Onoue Hitoshi , Kitamura Yukihiko , Nishimune Yoshhake
Published: April 2021

In order to define the role of Kit ligand (KL) growth factor encoded at the mouse steel (SI) locus in spermatogenesis, we have examined its production in Sertoli cells. As a measure KL growth factor bioactivity, the ability to support proliferation and maintenance of mast cells was used in co-culture with primary mouse Sertoli cells. On the rertoli cells derived from +/+ and Wv/Wv mice, +/+ mast cells proliferated and were supported for more than 2 weeks, but not W/Wv mast cells. In contrast, Sertoli cells from Sld/Sld mice could not support + /+ mast cell proliferation under similar conditions. The supportive effect required close-range interaction of Sertoli cells with cultured mast cells. These results indicate that Sertoli cells derived from +/+ and Wv/Wv but not Sld/Sld mutant mice produce biologically active KL growth factor as a membranebound form. The biologically active KL of Sertoli cells may also play an important role in germ cell growth and differentiation.
The beginning of pattern formation in theDrosophilacompound eye: the morphogenetic furrow and the second mitotic wave
Authors: Wolff Tanya , Ready Donald F.
Published: April 2021

Events in the morphogenetic furrow set the stage for all subsequent compound eye development in Drosophila. The periodic pattern of the adult eye begins in the furrow with the spaced initiation of ommatidial rudi- ments, the preclusters. A wave of mitosis closely follows the furrow. A cell-by-cell analysis reveals details of these events.

Early stages of ommatidial assembly can be resolved using a lead sulfide stain. Overt ommatidial organization begins in the morphogenetic furrow as cells gather into periodically spaced concentric aggregates. A stereotyped sequence of cell rearrangements converts these aggre- gates into preclusters. In the furrow, new rows of ommatidia are initiated at the equator and grow as new clusters are added to the peripheral ends.

Mitotic labeling using BrdU feeds shows that all cells not incorporated into a precluster divide. BrdU injec- tions show that cells divide roughly simultaneously between two adjacent rows of ommatidia.
Clonal analysis of epiblast fate during germ layer formation in the mouse embryo
Authors: Lawson Kirstie A. , Meneses Juanito J. , Pedersen Roger A.
Published: April 2021

The fate of cells in the epiblast at prestreak and early primitive streak stages has been studied by injecting horseradish peroxidase (HRP) into single cells in situ of 6.7-day mouse embryos and identifying the labelled descendants at midstreak to neural plate stages after one day of culture.

Ectoderm was composed of descendants of epiblast progenitors that had been located in the embryonic axis anterior to the primitive streak. Embryonic mesoderm was derived from all areas of the epiblast except the distal tip and the adjacent region anterior to it: the most anterior mesoderm cells originated posteriorly, traversing the primitive streak early; labelled cells in the posterior part of the streak at the neural plate stage were derived from extreme anterior axial and paraxial epiblast progenitors; head process cells were derived from epiblast at or near the anterior end of the primitive streak. Endoderm descendants were most frequently derived from a region that included, but extended beyond, the region producing the head process: descendants of epiblast were present in endoderm by the midstreak stage, as well as at later stages. Yolk sac and amnion mesoderm developed from posterolateral and posterior epiblast. The resulting fate map is essentially the same as those of the chick and urodele and indicates that, despite geometrical differences, topological fate relationships are conserved among these vertebrates. Clonal descendants were not necessarily confined to a single germ layer or to extraembryonic mesoderm, indicating that these lineages are not separated at the beginning of gastrulation.

The embryonic axis lengthened up to the neural plate stage by (1) elongation of the primitive streak through progressive incorporation of the expanding lateral and initially more anterior regions of epiblast and, (2) expansion of the region of epiblast immediately cranial to the anterior end of the primitive streak. The population doubling time of labelled cells was 7.5 h; a calculated 43 % were in, or had completed, a 4th cell cycle, and no statistically significant regional differences in the number of descendants were found. This clonal analysis also showed that (1) growth in the epiblast was noncoherent and in most regions anisotropic and directed towards the primitive streak and (2) the midline did not act as a barrier to clonal spread, either in the epiblast in the anterior half of the axis or in the primitive streak. These results taken together with the fate map indicate that, while individual cells in the epiblast sheet behave independently with respect to their neighbours, morphogenetic movement during germ layer formation is coordinated in the population as a whole.
Blastocoel expansion in the preimplantation mouse embryo: stimulatory effect of TGF-a and EGF
Authors: Dardik Alan , Schultz Richard M.
Published: April 2021

The factors that promote blastocoel expansion in the preimplantation mouse embryo are not well understood. Since cAMP stimulates the rate of blastocoel expansion and, in other systems, EGF can elevate intracellular cAMP levels, we investigated the ability of either TGF-α or EGF to stimulate the rate of blastocoel expansion in the mouse. Picomolar concentrations of either TGF-α or EGF stimulate the rate of blastocoel expansion in a concentration-dependent manner, and the continual presence of the growth factor is required to observe the stimulatory effect. Neutralizing antibodies to either TGF-α or EGF inhibit the TGF-α or EGF stimulatory effect, respectively. An antibody to the extracellular domain of the EGF receptor stimulates the rate of blastocoel expansion in a concentration-dependent manner, whereas an antibody to the cytoplasmic domain of the receptor does not. Tyrphostin RG 50864, which inhibits the EGF receptor kinase activity, inhibits the TGF-α stimulation of the rate of blastocoel expansion in a concentration-dependent manner; the less active tyrphostin, RG 50862, has no inhibitory effect. In addition, TGF-α does not stimulate a precocious onset of cavitation. The stimulatory effect on the rate of blastocoel expansion elicited by TGF-α or EGF is observed in 70 % of the embryos (responders). Responders and nonresponders have similar intracellular ATP levels and cell numbers. Whereas TGF-α stimulates the uptake of [35S]methionine into the acid-soluble and acid-insoluble pools in the responders, TGF-α has no stimulatory effect in the nonresponders. Results of these experiments suggest that an initial differentiative function of the first mammalian epithelium -fluid transport -is sensitive to peptide growth factor modulation.
Laser microsurgery demonstrates that cytoplasmic strands anchoring the nucleus across the vacuole of premitotic plant cells are under tension. Implications for division plane alignment
Authors: Goodbody Kim C. , Venverloo Catharina J. , Lloyd Clive W.
Published: April 2021

In epidermal cells of the plant Nautilocalyx lynchii, induced to divide by explantation, the nucleus undergoes a series of movements, on cytoplasmic strands, leading to construction of a division plane across the vacuole. In the early stage, the nucleus separates from the cortex, occupying an eccentric position in the cell, suspended across the vacuole by few thin strands. In the central stage, the nucleus occupies a central position anchored to the cortex by more numerous, thicker strands. Finally, the phragmosome forms as a coalescence of cytoplasmic strands across the cell, constituting the division plane within which mitosis and cytokinesis take place. The behaviour and alignment of these strands is therefore important since some are precursors of the division plane.

In a previous paper (Flanders et al (1990)J. Cell Biol. 110,1111 – 1122), it was pointed out that the alignment of cytoplasmic strands showed features common to a variety of elements under tension. That is, provided they are free to move relative to the cortex, strands radiating from the nucleus should tend to seek short rather than long routes to the cortex. In this way, strands under tension would move away from distant cell corners where two of the cell’s edges make a three-way junction with a neighbouring wall. This provides a basis for the avoidance of four-way junctions and the maintenance of three-way junctions which are a characteristic feature of most plant tissues. In addition, tensile elements such as soap bubble walls contact rigid surfaces perpendicularly. Perpendicular attachment of the cell plate to the side wall is embodied in Sachs’ rule of cell division and the existence of the premitotic strands in a state of tension would provide a basis for this rule.

In this study, laser microsurgery has been used to confirm that strands connecting the premitotic nucleus to the cortex are under tension since the severed ends retract immediately upon severance. However,. the response of the nucleus to the breaking of a cytoplasmic strand by laser depends upon the particular stage. The nucleus is most likely to move during the early stage when it is eccentrically placed and has few cytoplasmic strands. In the middle and phragmosomal stages the nucleus is more resistant. Computer-aided image reconstruction of anti-tubulin-labelled confocal sections indicates that the middle stage is characterised by thick microtubule (MT) bundles, which radiate from the nucleus and which seem to account for. the increasing stabilization of the nucleus. Although such strands are more difficult to sever with the laser, their ends still display elastic properties when cut. It is proposed that tension is likely to influence the alignment of strands as well as the position of the nucleus during division plane formation in vacuolated cells.
Developmental potential of parthenogenetic cells: role of genotype-specific modifiers
Authors: Fundele Reinald , Howlett Sarah K. , Kothary Rashmi , Norris Michael L. , Mills Walter E. , Surani M. Azim
Published: April 2021

The developmental potential of parthenogenetic cells derived from different mouse strains was investigated by examining their distribution in various tissues of adult aggregation chimeras. Using GPI-1 allozymes as marker, no striking differences were observed between chimeras whose parthenogenetic cells were derived from activated oocytes isolated from females of different genetic backgrounds, (C57BL/6 × CBA/J) F1, CFLP, 129, and SWR. In all the combinations tested, parthenogenetic cells were consistently absent from skeletal muscle, but there were varying contributions to most other tissues. These results suggest that the maternal duplication of chromosomes containing imprinted gene(s) responsible for the systematic elimination of parthenogenetic cells from skeletal muscle, are not subject to a pronounced influence of genotype-specific modifiers. However, the contribution of parthenogenetic cells to the brain does appear to be influenced by strain background, since a marked improvement in the survival of CFLP, 129 and perhaps SWR parthenogenetic cells in chimeric brains was observed compared with F2 cells.
Establishment of rostrocaudal polarity in tectal primordium: engrailed expression and subsequent tectal polarity
Authors: Itasaki Nobue , Ichijo Hiroyuki , Hama Chihiro , Matsuno Toru , Nakamura Harukazu
Published: April 2021

In the E4 (embryonic day 4) chick tectal primordium, engrailed expression is strong at the caudal end and gradually weakens toward the rostral end. We used quail-chick chimeric tecta to investigate how the caudorostral gradient of engrailed expression is established and whether it is correlated with the subsequent rostrocaudal polarity of tectal development.

To examine the positional value of the tectal primordium, we produced ectopic tecta in the diencephalon by transplanting a part of the mesencephalic alar plate heterotopically. In the ectopic tectum, the gradient of the engrailed expression reversed and the strength of the expression was dependent on the distance from the mes-diencephalon junction; the nearer the ectopic tectum was to the junction, the weaker the expression was. Consequently, the pattern of the engrailed expression in the host and ectopic tecta was nearly a mirror image, suggesting the existence of a repressive influence around the mes-diencephalon junction on the engrailed expression.

We examined cytoarchitectonie development in the ectopic tecta, which normally proceeds in a gradient along the rostrocaudal axis; the rostral shows more advanced lamination than the caudal. In contrast, the caudal part of the ectopic tecta (near to the mesdiencephalon junction) showed more advanced lamination than the rostral. In both the host and ectopic tecta, advanced lamination was observed where the engrailed expression was repressed, and vice versa.

Next we studied the correlation between engrailed expression and retinotectal projection from a view of plasticity and rigidity of rostrocaudal polarity in the tectum. We produced ectopic tecta by anisochronal transplantations between E3 host and E2 donor, and showed that there is little repressive influence at E3 around the mes-diencephalon junction. We then made chimeric double-rostral tectum (caudal half of it was replaced by rostral half of the donor tectum) or double-caudal tectum at E3. The transplants kept their original staining pattern in hosts. Consequently, the chimeric tecta showed wholly negative or positive staining of engrailed protein on the grafted side. In such tecta retinotectal projection pattern was disturbed as if the transplants retained their original position-specific characters.

We propose from these heterotopic and anisochronal experiments that the engrailed expression can be a marker for subsequent rostrocaudal polarity in the tectum, both as regards cytoarchitectonie development and retinotectal projection.
A G-string positive cis-regulatory element in the LpS1 promoter binds two distinct nuclear factors distributed non-uniformly in Lytechinus pictus embryos
Authors: Xiang Mengqing , Lu Szu-Yao , Musso Marco , Karsenty Gerard , Klein William H.
Published: April 2021

The LpS1 α and β genes of Lytechinus pictus are activated at the late cleavage stage of embryogenesis, with LpS1 mRNAs accumulating only in lineages contributing to aboral ectoderm. We had shown previously that 762 bp of 5’ flanking DNA from the LpS1/β gene was sufficient for proper temporal and aboral ectoderm specific expression. In the present study, we identified a strong positive cis-regulatory element at −70 bp to −75 bp in the LpS1/β promoter with the sequence (G)6 and a similar, more distal cis-element at −721 bp to −726 bp. The proximal ‘G-string’ element interacted with two nuclear factors, one specific to ectoderm and one to endoderm/mesoderm nuclear extracts, whereas the distal G-string element interacted only with the ectoderm factor. The ectoderm and endoderm/mesoderm G-string factors were distinct based on their migratory behavior in electrophoretic mobility shift assays, binding site specificities, salt optima and EDTA sensitivity. The proximal G-string element shared homology with a binding site for the mammalian transcription factor IF1, a protein that binds to negative cis-regulatory elements in the mouse al(I) and a2(I) collagen gene promoters. Competition experiments using wild-type and mutant oligonucleotides indicated that the ectoderm G-string factor and IF1 have similar recognition sites. Partially purified IF1 specifically bound to an oligonucleotide containing the proximal G-string of LpS1/β. From our results, we suggest that the ectoderm G-string factor, a member of the G-rich DNA-binding protein family, activates the LpS1 gene in aboral ectoderm cells by binding to the LpS1 promoter at the proximal G-string site.
Specification of cell fate in the developing eye of Drosophila
Authors: Hafen Ernst , Basler Konrad
Published: April 2021

Determination of cell fate in the developing eye of Drosophila depends on cellular interactions. In the eye imaginai disc, an initially unpatterned epithelial sheath of cells, single cells are specified in regular intervals to become the R8 photoreceptor cells. Genes such as Notch and scabrous participate in this process suggesting that specification of ommatidial founder cells and the formation of bristles in the adult epidermis involve a similar mechanism known as lateral inhibition. The subsequent steps of ommatidial assembly involve a different mechanism: undetermined cells read their position based on the contacts they make with neighbors that have already begun to differentiate. The development of the R7 photoreceptor cell is best understood. The key role seems to be played by sevenless, a receptor tyrosine kinase on the surface of the R7 precursor. It transmits the positional information - most likely encoded by boss on the neighboring R8 cell membrane - into the cell via its tyrosine kinase that activates a signal transduction cascade. Two components of this cascade - Sos and sina - have been identified genetically, sina encodes a nuclear protein whose expression is not limited to R7. Constitutive activation of the sevenless kinase by overexpression results in the diversion of other ommatidial cells into the R7 pathway, suggesting that activation of the sevenless signalling pathway is sufficient to specify R7 development.
The regulation of motoneuron survival and differentiation by putative muscle-derived neurotrophic agents: neuromuscular activity and innervation
Authors: Houenou Lucien J. , Haverkamp Lanny J. , McManaman James L. , Oppenheim Ronald W.
Published: April 2021

The chronic blockade of neuromuscular activity is known to promote the survival of developing motoneurons in vivo in the chick, mouse and rat embryo. Increased survival in this situation may reflect an activity-dependent mechanism for the regulation of trophic factor production by target cells. To test this notion, we have examined motoneuron survival in vivo and choline acetyltransferase (ChAT) development in vitro following treatment of chick embryos and rat spinal cord cultures with partially purified skeletal muscle extracts derived from normally active, chronically paralyzed and aneural embryos, and from denervated postnatal chickens. Extracts from active and paralyzed chick embryos were equally effective in promoting motoneuron survival and ChAT activity. Aneural embryonic muscle extracts were slightly less effective in promoting motoneuron survival in vivo, but were not significantly different from control extracts in the in vitro ChAT assay. Denervated postnatal muscle extracts, however, were more effective in enhancing both motoneuron survival and ChAT activity. These data indicate that: (1) the promotion of motoneuron survival in vivo by activity blockade may not be mediated by an up-regulation of trophic factor synthesis in target cells; (2) postnatal innervation may regulate the production of putative muscle-derived neurotrophic factors; and (3) the synthesis or availability of trophic agents may be regulated differently in embryonic and postnatal muscle.
Origin of segmental identity in the development of the leech nervous system
Authors: Shankland Marty , Martindale Mark Q. , Nardelli-Haefliger Denise , Baxter Euan , Price David J.
Published: April 2021

The leech embryo develops its segmental body plan by means of a stereotyped cell lineage. Each hemilateral segment arises from a small set of embryonic blast cells via a comparable sequence of formative cell divisions, and for the most part, lineally homologous cells manifest similar patterns of differentiation in the various hemisegments. Nonetheless, some identified central neurons undergo segment-specific or laterally asymmetric patterns of neuropeptide expression and/or cell death. Certain aspects of this regional diversification result from competitive cell interactions which occur at the level of the postmitotic neuron. However, the neuron’s segmental identity is lineally determined, being inherited from its blast cell progenitor over several intervening rounds of mitosis. To learn more about the molecular basis of this phenomenon, we have isolated and begun to characterize leech homeobox genes which are related to the genes that govern segmental identity in other organisms.
Alternating patterns of cell surface properties and neural crest cell migration during segmentation of the chick hindbrain
Authors: Lumsden Andrew , Guthrie Sarah
Published: April 2021

The developing chick hindbrain is transiently divided into a series of repeating units or rhombomeres. Recent work has shown that an alternating periodicity exists both in the cell surface properties of rhombomeres and in the segmental origin of hindbrain neural crest cells. Experiments in which rhombomeres from different axial levels were confronted in the absence of an inter-rhombomere boundary showed that odd-numbered segments 3 and 5 combined without generating a boundary, as did even-numbered segments 2, 4 and 6. When rhombomeres originating from adjacent positions, or three rhombomeres distant from one another were combined, a new boundary was regenerated.

Mapping of the migration pathways of neural crest cells showed that odd-numbered and even-numbered rhombomeres share properties with respect to the production of neural crest cells. In the hindbrain region the neural crest is segregated into streams. Neural crest cells migrating from rhombomeres 1 and 2, rhombomere 4 and rhombomere 6 respectively populate distinct cranial nerve ganglia and branchial arches. In contrast, rhombomeres 3 and 5 are free of neural crest cells.
A nude mouse xenograft model of fetal intestine development and differentiation
Authors: Buono Raffaele Del , Fleming Kenneth A. , Morey Adrienne L. , Hall Peter A. , Wright Nicholas A.
Published: April 2021

This report describes a novel in vivo model of intestinal differentiation. Fourteen day, undifferentiated fetal rat small intestine, stripped of the major part of its mesenchyme, suspended in a type I collagen gel and then xenografted into a nude mouse, undergoes small intestinal morphogenesis and cytodifferentiation. All four major epithelial lineages, namely Paneth, goblet, columnar and endocrine are present. Double-label nonisotopic in situ hybridization, employing biotinylated and digoxigenin-labelled whole rat DNA and whole mouse DNA probes, was performed to distinguish donor cells from host cell types. The outer longitudinal smooth muscle layer, and the major part of the lamina propria, including pericryptal fibroblasts, are of host mouse origin; the inner circular smooth muscle layer is of donor rat origin. Cells of the muscularis propria and lamina propria acquired smooth muscle α-actin, presumably under the influence of the donor endoderm. Furthermore, this xenograft develops a host vascular network, and cells with the morphological appearance of lymphocytes are present within the intestinal epithelium. The production of chemotactic factors by the endoderm is postulated because grafting of collagen gel alone results in a minimal invasion by stromal cells which do not express smooth muscle α-actin.
Two myogenic lineages within the developing somite
Authors: Ordahl C.P. , Le Douarin N.M.
Published: April 2021

It is well known that the muscles of the vertebrate body are derived from the somite. Precursor cells within the somite proper form the back or axial muscles while other precursor cells migrate away from the somite to populate the muscle of the limbs and ventral body wall. Although both types of muscle are generally thought of as arising from a common progenitor population, the myotome, recent evidence points to developmental differences in these two groups of muscles which may reflect different developmental lineages. To test the lineage hypothesis, we used microsurgery and the chick-quail nucleolar marker system to follow the developmental fate of the lateral and medial halves of somites at the wing level. The results showed that the structures of the mature somite (myotome and sclerotome) are derived virtually exclusively from cells residing in the medial half of the newly formed somite. On the other hand, virtually all of the cells residing in the lateral half of the newly formed somite are destined to leave the somite proper and populate the limb muscle and, probably, other somite-derived mesenchymal structures in the limb and ventral body wall. Switch-graft experiments show that the two halves of newly formed somites are largely interchangeable demonstrating that their ultimate developmental fate is position-dependent and that it becomes fixed as a result of extrinsic influences which act during later stages of somitogenesis. We conclude that at least two distinct myogenic lineages exist in the somite; one giving rise to the muscles of the back and the other giving rise to the limb musculature.
Disruption of pioneer growth cone guidance in vivo by removal of glycosyl phosphatidylinositol-anchored cell surface proteins
Authors: Chang Wesley S. , Serikawa Kyle , Allen Karen , Bentley David
Published: April 2021

Cell surface proteins anchored to membranes via covalently attached glycosyl-phosphatidylinositol (GPI) have been implicated in neuronal adhesion, promotion of neurite outgrowth and directed cell migration. Treatment of grasshopper embryos with bacterial phosphatidylinositol-specific phospholipase C (PI-PLC), an enzyme that cleaves the GPI anchor, often induced disruptions in the highly stereotyped migrations of peripheral pioneer growth cones and afferent neuron cell bodies. In distal limb regions of embryos treated with PI-PLC at early stages of pioneer axon outgrowth, growth cones lost their proximal orientation toward the central nervous system (CNS) and turned distally. Pioneer growth cones in treated limbs also failed to make a characteristic ventral turn along the trochanter-coxa (Tr-Cx) segment boundary, and instead continued to grow proximally across the boundary. Treatment at an earlier stage of development caused pre-axonogenesis Cxi neurons to abandon their normal circumferential migration and reorient toward the CNS. None of these abnormal phenotypes were observed in limbs of untreated embryos or embryos exposed to other phospholipases that do not release GPI-anchored proteins. Incubation of embryos with PI-PLC effectively removed immunoreactivity for fasciclin I, a GPI-anchored protein expressed on a subset of neuronal surfaces. These results suggest that cell surface GPI-anchored proteins are involved in pioneer growth cone guidance and in pre-axonogenesis migration of neurons in the grasshopper limb bud in vivo.
Molecular analysis of the deletion mutants in the E homeotic complex of the silkworm Bombyx mori
Authors: Ueno Kohji , Hui Chi-Chung , Fukuta Masakazu , Suzuki Yoshiaki
Published: April 2021

The E loci in Bombyx mori are expected to contain a homeotic gene complex specifying the identities of the larval abdominal segments. However, the molecular structure of this complex remains to be determined. We have started to analyze the structural changes in the E complex mutations. We used three newly isolated Bombyx homeobox genes as probes. These genes are probably homologues of the Ultrabiihorax (Ubx), ab-dominal-A (abd-A) and Abdominal-B (Abd-B) in the Drosophila bithorax complex, because the amino-acid sequences of the homeobox regions in these Bombyx genes are almost identical to those of Drosophila genes. We found that the Bombyx Ubx and abd-A genes are deleted in the Eca chromosome, and the Bombyx abd-A gene is deleted in the chromosome. From these results, we conclude that the Bombyx E complex consists of the Ubx, abd-A and possibly Abd-B genes, which may play similar roles to their homologues in the Drosophila bithorax complex.
Motor neuron pathfinding following rhombomere reversals in the chick embryo hindbrain
Authors: Guthrie Sarah , Lumsden Andrew
Published: April 2021

Motor neurons are segmentally organised in the developing chick hindbrain, with groups of neurons occupying pairs of hindbrain segments or rhombomeres. The branchiomotor nucleus of the trigeminal nerve occupies rhombomeres 2 and 3 (r2 and r3), that of the facial nerve r4 and r5, and that of the glossopharyngeal nerve r6 and r7. Branchiomotor neuron cell bodies lie within the basal plate, forming columns on either side of the ventral midline floor plate. Axons originating in rhombomeres 2, 4 and 6 grow laterally (dorsally) towards the exit points located in the alar plates of these rhombomeres, while axons originating in odd-numbered rhombomeres 3 and 5 grow laterally and then rostrally, crossing a rhombomere boundary to reach their exit point. Examination of the trajectories of motor axons in odd-numbered segments at late stages of development (19-25) showed stereotyped pathways, in which axons grew laterally before making a sharp turn rostrally. During the initial phase of outgrowth (stage 14-15), however, axons had meandering courses and did not grow in a directed fashion towards their exit point. When r3 or r5 was transplanted with reversed rostrocaudal polarity prior to motor axon outgrowth, the majority of axons grew to their appropriate, rostral exit point, despite the inverted neuroepithelial polarity. In r3 reversals, however, there was a considerable increase in the normally small number of axons that grew out via the caudal, r4 exit point. These findings are discussed with relevance to the factors involved in motor neuron specification and axon outgrowth in the developing hindbrain.
Early mRNAs, spatially restricted along the animal-vegetal axis of sea urchin embryos, include one encoding a protein related to tolloid and BMP-1
Authors: Reynolds Susan D. , Angerer Lynne M. , Palis James , Nasir Adnan , Angerer Robert C.
Published: April 2021

The cloning and characterization of cDNAs representing four genes or small gene families that are coordinately expressed in a spatially restricted pattern during the very early blastula (VEB) stage of sea urchin development are presented. The VEB genes encode multiple transcripts that are expressed transiently in embryos of Strongylocentrotus purpuratus between 16-cell stage and hatching, with peak abundance 12 to 15 hours postfertilization (∼150-250 cells). The VEB transcripts share the same spatial pattern in the early blastula embryo: they are asymmetrically distributed along the animalvegetal axis but their distribution around this axis is uniform. Thus, the VEB transcripts are the earliest messages to reveal asymmetry along the primary axis in the sea urchin embryo. The temporal and spatial patterns of VEB transcript accumulation are not consistent with involvement of these gene products in cell division or in tissue-specific functions. Furthermore, VEB messages cannot be detected in either ovary or adult tissues, suggesting that these genes function exclusively during embryogenesis. We suggest that the VEB genes function in constructing the early blastula. Two VEB genes encode metalloendoproteases: one (SpHE) is hatching enzyme and the other (SpAN) is similar to bone morphogenetic protein-1 (BMP-1; Wozney et al., Science 242: 1528-1534, 1988) and the Tolloid gene product (tld) (Shimell et al., Cell 67: 459482, 1991). Several lines of evidence suggest that the VEB genes are regulated directly by factors or regulatory activities localized along the maternally specificed animal-vegetal axis.
Effects of W (c-kit) gene mutation on gametogenesis in male mice: agametic tubular segments in Wf/Wf testes
Authors: Koshimizu Uichi , Watanabe Daisuke , Tajima Youichi , Nishimune Yoshitake
Published: April 2021

Mutations of the W (e-kit) gene, which encodes a transmembrane tyrosine kinase receptor, affect the development and differentiation of many types of stem cell. Most homozygous W mutant mice are sterile, due to a lack of germ cells arising during embryonic development, but one of the notable exceptions is mice, which are fully fertile in both sexes. In order to elucidate the effects of the Wf mutation on spermatogenesis, postnatal spermatogenesis in Wf/Wf mice was histologically examined. The number of gonocytes at birth was significantly reduced and small portions of agametic seminiferous tubule segments were observed in mutant mice. It is suggested that this is due to a deficiency of primordial germ cells (PGC). Other than the agametic tubules, there was no evidence of reduced spermatogenesis after birth. These results indicate that the function of the W (e-kit) gene is more necessary for the development of PGC than for postnatal germ cells.
The mouse has a Polycomb-Wke chromobox gene
Authors: Pearce Jonathan J. H. , Singh Prim B. , Gaunt Stephen J.
Published: April 2021

The Drosophila gene Polycomb (Pc) has been implicated in the clonal inheritance of determined states and is a trans-regulator of the Antennapedia-like homeobox genes. Pc shares a region of homology (the chromobox) with the Drosophila gene Heterochromatin Protein 1 (HP1), a component of heterochromatin. The Pc chromobox has been used to isolate a mouse chromobox gene, M33, which encodes a predicted 519 amino acid protein. The M33 chromodomain is more similar to that in the Pc protein, than that in the HP1 protein. In addition to the chromodomain, the M33 and Pc proteins also share a region of homology at their C termini. The temporal and spatial expression patterns of M33 have been studied by in situ hybridization and northern analysis. During the final 10 days of embryonic development, M33 expression mirrors that of the cellcycle-specific cyclin B gene. It is therefore suggested that the rate of cellular proliferation controls M33 expression. From comparisons of the characteristics of M33 with those of Pc it is proposed that M33 is a Pc-like chromobox gene. The roles of M33 and Pc in models of cellular memory are examined and implications of the memory models addressed.
Afferent influences on cell death and birth during development of a cortical nucleus necessary for learned vocal behavior in zebra finches
Authors: Johnson Frank , Bottjer Sarah W.
Published: April 2021

Forebrain nuclei that control learned vocal behavior in zebra finches are anatomically distinct and interconnected by a simple pattern of axonal pathways. In the present study, we examined afferent regulation of neuronal survival during development of the robust nucleus of the archistriatum (RA). RA projection neurons form the descending motor pathway of cortical vocal-control regions and are believed to be directly involved in vocal production. RA receives afferent inputs from two other cortical regions, the lateral magnocellular nucleus of the anterior neostriatum (lMAN) and the higher vocal center (HVC). However, because the ingrowth of HVC afferent input is delayed, lMAN projection neurons provide the majority of afferent input to RA during early vocal learning. lMAN afferent input to RA is of particular interest because lMAN is necessary for vocal learning only during a restricted period of development. By making lesions of lMAN in male zebra finches at various stages of vocal development (20-60 days of age) and in adults (>90-days old), we asked whether the survival of RA neurons depends on lMAN afferent input, and if so whether such dependence changes over the course of vocal learning. The results showed that removal of lMAN afferent input induced the loss of over 40% of RA neurons among birds in early stages of vocal development (20 days of age). However, lMAN lesions lost the ability to induce RA neuron death among birds in later stages of vocal development (40 days of age and older). These findings indicate that many RA neurons require lMAN afferent input for their survival during early vocal learning, whereas the inability of lMAN lesions to induce RA neuron death in older birds may indicate a reduced requirement for afferent input or perhaps the delayed ingrowth of HVC afferent input (at approx. 35 days of age) provides an alternate source of afferent support. Removal of lMAN afferent input also dramatically increased the incidence of mitotic figures in RA, but only among 20-day-old birds at 2 days post-lesion. The early, acute nature of the mitotic events raises the possibility that cell division in RA may be regulated by lMAN afferent input.
Ciliary neurotrophic factor and leukemia inhibitory factor promote the generation, maturation and survival of oligodendrocytes in vitro
Authors: Mayer Margot , Bhakoo Kishore , Noble Mark
Published: April 2021

We have found that CNTF and LIF are pleiotropic modulators of development in the O-2A lineage. Both molecules enhanced the generation of oligodendrocytes in cultures of dividing O-2A progenitors. CNTF and LIF also promoted oligodendrocyte maturation, as determined by expression of myelin basic protein, and could promote oligodendrocyte survival to an extent comparable with insulin-like growth factor-1 or insulin. In addition, LIF and CNTF both promoted the differentiation of O-2A progenitors into type-2 astrocytes but only when applied in the presence of extra-cellular matrix (EnMx) derived from cultures of endo-thelial cells. The ability of CNTF and LIF to enhance differentiation of O-2A progenitors along either of the alternative pathways of oligodendrocyte and astrocyte differentiation suggests that these proteins are able to enhance the process of differentiation per se, while the actual path of differentiation promoted is determined by the presence or absence of additional molecules in the extracellular environment.
A novel member of the transmembrane serine/threonine kinase receptor family is specifically expressed in the gonads and in mesenchymal cells adjacent to the müllerian duct
Authors: Baarends Willy M. , Helmond Marjolein J. L. van , Post Miriam , Schoot Pieter J. C. M. van der , Hoogerbrugge Jos W. , Winter Johan P. de , Uilenbroek Jan Th. J. , Karels Bas , Wilming Laurens G. , Meijers J. H. Carel , Themmen Axel P. N. , Grootegoed J. Anton
Published: April 2021

The activin and TGF-β type II receptors are members of a separate subfamily of transmembrane receptors with intrinsic protein kinase activity, which also includes the recently cloned TGF-β type I receptor. We have isolated and characterized a cDNA clone (C14) encoding a new member of this subfamily. The domain structure of the C14-encoded protein corresponds with the structure of the other known transmembrane serine/threonine kinase receptors. It also contains the two inserts in the kinase domain that are characteristic for this subfamily. Using in situ hybridization, C14 mRNA was detected in the mesenchymal cells located adjacent to the müllerian ducts of males and females at day 15 (E15) of embryonic development. Marked C14 mRNA expression was also detected in the female gonads. In female E16 embryos, the C14 mRNA expression pattern remained similar to that in E15 embryos. However, in male E16 embryos C14 mRNA was detected in a circular area that includes the degenerating müllerian duct. The expression of C14 mRNA was also studied using RNase protection assays. At E15 and E16, C14 mRNA is expressed in the female as well as in the male urogenital ridge. However, at E19, a high C14 mRNA level in the female urogenital ridge contrasts with a lack of C14 mRNA in the male urogenital ridge. This correlates with the almost complete degeneration of the müllerian ducts in male embryos at E19. C14 mRNA expression was also detected in embryonic testes at E15, E16 and E19 using RNase protection assays, but at much lower levels than those found in the developing ovaries. In eleven other tissues no C14 mRNA was observed.

The results point to anti-müllerian hormone (AMH) being the most likely candidate ligand for C14. The embryonic C14 mRNA expression pattern in the urogenital ridge correlates with the expected site of AMH action, and C14 mRNA expression in the fetal ovary is in agreement with known effects of AMH on gonadal differentiation.

Postnatal C14 mRNA expression in rats was found to be confined mainly to the gonads. In the testis, C14 mRNA expression occurs in Sertoli cells. This testicular expression markedly increases during the first 3 weeks after birth, concurrent with the onset of spermatogenesis.
Neurogenic expression of snail is controlled by separable CNS and PNS promoter elements
Authors: Tony Ip Y. , Levine Michael , Bier Ethan
Published: April 2021

The Drosophila snail (sna) gene is first expressed in cells giving rise to mesoderm and is required for mesoderm formation. sna is subsequently expressed in the developing nervous system. sna expression during neurogenesis evolves from segmentally repeated neuroectodermal domains to a pan-neural pattern. We have identified a 2.8kb regulatory region of the sna promoter that drives LacZ expression in a faithful neuronal pattern. Deletion analysis of this region indicates that the pan-neural element is composed of separable CNS and PNS components. This finding is unexpected since all known genes controlling early neurogenesis, including the proneural genes (i.e. da and AS-C), are expressed in both the CNS and PNS. We also show that expression of sna during neurogenesis is largely independent of the proneural genes da and AS-C. The separate control of CNS and PNS sna expression and independence of proneural gene regulation add to a growing body of evidence that current genetic models of neurogenesis are substantially incomplete.
Bone morphogenetic proteins and a signalling pathway that controls patterning in the developing chick limb
Authors: Francis Philippa H. , Richardson Michael K. , Brickell Paul M. , Tickle Cheryll
Published: April 2021

We show here that bone morphogenetic protein 2 (BMP-2) is involved in patterning the developing chick limb. During early stages of limb development, mesenchymal expression of the Bmp-2 gene is restricted to the posterior part of the bud, in a domain that colocalizes with the polarizing region. The polarizing region is a group of cells at the posterior margin of the limb bud that can respecify the anteroposterior axis of the limb when grafted anteriorly and can activate expression of genes of the HoxD complex. We dissect possible roles of BMP-2 in the polarizing region signalling pathway by manipulating the developing wing bud. Retinoic acid application, which mimics the effects of polarizing region grafts, activates Bmp-2 gene expression in anterior cells. This shows that changes in anteroposterior pattern are correlated with changes in Bmp-2 expression. When polarizing region grafts are placed at the anterior margin of the wing bud, the grafts continue to express the Bmp-2 gene and also activate Bmp-2 expression in the adjacent anterior host mesenchyme. These data suggest that BMP-2 is part of the response pathway to the polarizing signal, rather than being the signal itself. In support of this, BMP-2 protein does not appear to have any detectable polarizing activity when applied to the wing bud. The pattern of Bmp-4 gene expression in the developing wing bud raises the possibility that BMP-2 and BMP-4 could act in concert. There is a close relationship, both temporal and spatial, between the activation of the Bmp-2 and Hoxd-13 genes in response to retinoic acid and polarizing region grafts, suggesting that expression of the two genes might be linked.
Expression of the zebrafish gene hlx-1 in the prechordal plate and during CNS development
Authors: Fjose Anders , Izpisúa-Belmonte Juan-Carlos , Fromental-Ramain Catherine , Duboule Denis
Published: April 2021

The zebrafish hlx-1 gene belongs to the H2.0 subfamily of homeobox genes and is closely related to the mouse Dbx gene with respect to both homeodomain homology (96.7%) and neural expression during embryogenesis. Analysis of hlx-1 expression by in situ hybridization reveals several particularly interesting features. In late gastrula embryos, hlx-1 transcripts are detected within a circular area in the region of the presumptive rostral brain. Subsequently, the expression domain becomes restricted to the hypoblast and undergoes dynamic changes involving conversion into a longitudinal stripe which elongates and retracts following a temporal sequence. The site of transient hlx-1 expression along the ventral midline of the rostral neurectoderm, which in part corresponds to the prechordal plate, suggests a role in the determination of head mesoderm as well as in patterning of the rostral brain. As the midline stripe gradually disappears, the hlx-1 gene becomes regionally expressed within the diencephalon and at a specific dorsoventral level along the hindbrain and spinal cord. In the hindbrain, expression is initiated in dorsoventrally restricted transversal stripes which correlate with the segmental pattern of rhombomeres. The stripes fuse into bilateral columns that are later converted to two series of paired transversal stripes at the rhombomere borders. This pattern is consistent with the proposed subdivision of hindbrain segments into rhombomere centers separated by border regions.