bluuebunny/crossref_metadata_2025 huggingface dataset:
Developmental expression of the mouse c-rel proto-oncogene in hematopoietic organs
Authors: Carrasco Daniel , Weih Falk , Bravo Rodrigo
Published: April 2021

We have studied the expression of the c-rel proto-oncogene during mouse embryonic development and adult animals using in situ hybridization and immunocytochemical analysis. c-rel transcripts were detected late in development with an expression pattern that parallels the emergence and diversification of hematopoietic cells. In the embryo, c-rel is expressed first in the mesoderm-derived hematopoi-etic cells of the liver and later also in other hematopoietic tissues such as thymus and spleen. This correlation between c-rel expression and places of hematopoietic infiltration is conserved in the postnatal period, with expression of c-rel mRNA in the medullary region of the thymus and in splenic B cell areas, including the marginal zone and the outer region of the periarterial sheath. High levels of c-rel transcripts were also detected in the splenic germinal centers, lymph nodes and Peyer’s patches. Using double immuno-fluorescence and cell preparations from different embryonic and adult hematopoietic organs, we have defined the pattern and cell types of c-rel expression in different hematopoietic cell lineages and in the stromal cell content of the thymus. By using electrophoretic mobility shift assays, we have also correlated c-Rel expression in spleen with κB-binding activity in the form of c-Rel/p50 and c-Rel/p52 heterodimers. The timing and pattern of expression of the c-rel proto-oncogene in the different cell lineages suggest that temporally regulated changes in c-Rel expression may be required for vertebrate hematopoiesis.
Autonomous concentration-dependent activation and repression of Krüppel by hunchback in the Drosophila embryo
Authors: Schulz Cordula , Tautz Diethard
Published: April 2021

The subdivision of the anterior-posterior axis in Drosophila is achieved by a cascade of spatially regulated transcription factors which form short-range gradients at the syncytial blastoderm stage. These factors are assumed to have concentration-dependent regulatory effects on their target genes. However, there is so far little direct in vivo evidence that a single factor can autonomously activate and repress a given target gene. We have analysed here the regulatory capabilities of the gap gene hunchback by creating an artificial gradient of hunchback in the early embryo. This was achieved by providing the maternally expressed mRNA of hunchback with the anterior localization signal of the bicoid RNA. The effects of this artificial hunchback gradient were then studied in different types of mutant background. We show that under these conditions hb is autonomously capable of activating the target gene Krüppel at low concentrations and repressing it at high concentrations. In addition, we show that the artificially created hunchback gradient can organize a large part of the segment pattern, although it is expressed at a different position and in a different shape than the wild-type gradient of hunchback.
Mouse embryonic germ (EG) cell lines: transmission through the germline and differences in the methylation imprint of insulin-like growth factor 2 receptor (Igf2r) gene compared with embryonic stem (ES) cell lines
Authors: Labosky Patricia A. , Barlow Denise P. , Hogan Brigid L. M.
Published: April 2021

Primordial germ cells of the mouse cultured on feeder layers with leukemia inhibitory factor, Steel factor and basic fibroblast growth factor give rise to cells that resemble undifferentiated blastocyst-derived embryonic stem cells. These primordial germ cell-derived embryonic germ cells can be induced to differentiate extensively in culture, form teratocarcinomas when injected into nude mice and contribute to chimeras when injected into host blastocysts. Here, we report the derivation of multiple embryonic germ cell lines from 8.5 days post coitum embryos of C57BL/6 inbred mice. Four independent embryonic germ cell lines with normal male karyotypes have formed chimeras when injected into BALB/c host blastocysts and two of these lines have transmitted coat color markers through the germline. We also show that pluripotent cell lines capable of forming teratocarcinomas and coat color chimeras can be established from primordial germ cells of 8.0 days p.c. embryos and 12.5 days p.c. genital ridges. We have examined the methylation status of the putative imprinting box of the insulin-like growth factor type 2 receptor gene (Igf2r) in these embryonic germ cell lines. No correlation was found between methylation pattern and germline competence. A significant difference was observed between embryonic stem cell and embryonic germ cell lines in their ability to maintain the methylation imprint of the Igf2r gene in culture. This may illustrate a fundamental difference between these two cell types.
Glia in the fly wing are clonally related to epithelial cells and use the nerve as a pathway for migration
Authors: Giangrande Angela
Published: April 2021

The Drosophila major wing nerve collects axons from the anterior margin sensory organs. Using the flp recombinase to make clones, I show that all glia present on this nerve are clonally related to wing epithelial cells. Glial cells arise only from regions that also give rise to sensory organs and migrate along the nerve following the direction taken by axons. As in vertebrates, wing glial cells start migrating at a stage at which axons are growing. The migration of wing glial cells is affected by two mutations altering axonogenesis, fused and Notch, which suggests that the two processes are tightly associated.
The sevenless signalling cassette mediates Drosophila EGF receptor function during epidermal development
Authors: Diaz-Benjumea Fernando J. , Hafen Ernst
Published: April 2021

In Drosophila, Drk, an SH2 adaptor protein, Sos, a putative activator of Ras1, Ras1, raf and rolled/MAP kinase have been shown to be required for signalling from the sevenless and the torso receptor tyrosine kinase. From these studies, it was unclear whether these components act in a single linear pathway as suggested by the genetic analysis or whether different components serve to integrate different signals. We have analyzed the effects of removing each of these components during the development of the adult epidermal structures by generating clones of homozygous mutant cells in a heterozygous background. Mutations in each of these signalling components produce a very similar set of phenotypes. These phenotypes resemble those caused by loss-of-function mutations in the Drosophila EGF receptor homolog (DER). It appears that these components form a signalling cassette, which mediates all aspects of DER signalling but that is not required for other signalling processes during epidermal development.
Sex-determining genes in the homosporous fern Ceratopteris
Authors: Banks Jo Ann
Published: April 2021

Haploid Ceratopteris gametophytes are either hermaphroditic or male. The determinate of sex type is the pheromone antheridiogen (ACE) which is secreted by the meristic hermaphrodite and promotes ameristic male development of sexually undetermined gametophytes. Several mutations effecting the sex of the haploid gametophyte have been isolated and are described. The hermaphroditic (her) mutants are insensitive to ACE and develop as meristic hermaphrodites. These mutations effect ameristic male development in the presence of ACE but have no effect on hermaphroditic development. While most her mutations also have no effect on diploid sporophyte development, some partially ACE-insensitive her mutations have profound effects on sporophyte development. The transformer (tra) mutation effects both meristem and archegonia formation and causes the gametophyte to be an ameristic male under conditions that promote hermaphroditic development. The feminization (fem) mutation effects antheridia development in both male and hermaphroditic gametophytes and causes the gametophyte to develop as a meristic female in the absence or presence of the pheromone. The her1 tra1 double mutant is male in the absence or presence of ACE, indicating that tra1 is epistatic to her1. The phenotypes of her1, tra1 and fem1 single gene mutant phenotypes and the her1 tra1 double mutant phenotype are used to deduce a model suggesting how the products of these genes might interact in a regulatory pathway to control sex determination.
Stage-specific requirement for myosin II during Dictyostelium development
Authors: Springer M. L. , Patterson B. , Spudich J. A.
Published: April 2021

Dictyostelium cells that lack a functional myosin II heavy chain are motile and are capable of aggregation, but fail to undergo further multicellular development. We have used a Dictyostelium mutant expressing a cold-sensitive myosin heavy chain to examine the requirement for myosin throughout the course of development. The loss of myosin function upon cooling is rapid and reversible. Tempera-ture-shift experiments reveal that myosin is essential during two different stages of development. During aggregation, myosin function appears to be necessary for cells to sort correctly in a way that allows further development to occur. During the final stage of development, it is required for the formation of a complete stalk and the raising of the spore head. Development between those stages, however, proceeds normally in the absence of myosin function. Aggregates at non-permissive temperature undergo an aberrant form of development resulting in a ball of cells. Calcofluor staining and reporter gene fusions reveal that these structures contain defective spores and a miniature stalk.
Promoter-specific IGF2 imprinting status and its plasticity during human liver development
Authors: Ekström Tomas J. , Cui Hengmi , Li Xuri , Ohlsson Rolf
Published: April 2021

IGF2 has been shown to be expressed preferentially from the paternally derived allele, although the maternal allele can be found active during both prenatal and postnatal development as well as in neoplastic tumours in humans. We addressed here whether or not the biallelic expression patterns that can be seen during postnatal human liver development reflected a coordinated change in the activities of the four promoters of human IGF2. We show here that the P2, P3 and P4 promoters, but not the P1 promoter, display monoallelic activity in embryonic, neonatal and younger infant liver specimens. The P2, P3 and P4 promoters can, however, be found active either monoallelically or biallelically or even monoallelically on opposite parental alleles in older infant and adult liver specimens. In contrast, H19, which is closely linked to IGF2, is monoallelically expressed in all postnatal liver samples analysed. We conclude that the functional imprinting status of IGF2 during postnatal liver development appears to be promoter/enhancer-specific and either partly or completely independent of H19.
Invagination centers within the Drosophila stomatogastric nervous system anlage are positioned by Notch-mediated signaling which is spatially controlled through wingless
Authors: González-Gaitán Marcos , Jäckle Herbert
Published: April 2021

The gut-innervating stomatogastric nervous system of Drosophila, unlike the central and the peripheral nervous system, derives from a compact, single layered epithelial anlage. Here we report how this anlage is initially defined during embryogenesis by the expression of proneural genes of the achaete-scute complex in response to the maternal terminal pattern forming system. Within the stomatogastric nervous system anlage, the wingless-dependent inter-cellular communication system adjusts the cellular range of Notch-dependent lateral inhibition to single-out three achaete-expressing cells. Those cells define distinct invagination centers which orchestrate the behavior of neighboring cells to form epithelial infoldings, each headed by an achaete-expressing tip cell. Our results suggest that the wingless pathway acts not as an instructive signal, but as a permissive factor which coordinates the spatial activity of morphoregulatory signals within the stomatogastric nervous system anlage.
The existence of inositol 1,4,5-trisphosphate and ryanodine receptors in mature bovine oocytes
Authors: Yue Caiping , White Kenneth L. , Reed William A. , Bunch Thomas D.
Published: April 2021

Intracellular Ca2+ (Ca2+i) transients during fertilization are critical to the activation of eggs in all species studied. Activation of both the inositol 1,4,5-trisphosphate (IP3) receptor (IP3R) and ryanodine receptor (RYR) are responsible for the calcium oscillations during fertilization in sea urchin eggs. Using in vitro matured bovine oocytes loaded with Fura-2 AM ester as Ca2+i indicator, we addressed whether IP3Rs and RYRs coexist in mammalian eggs. Our results indicate that microinjection of 50-250 nM IP3 or 10-20 mM caffeine, 100-200 μM ryanodine and 4-8 μM cyclic ADP-ribose all induced Ca2+i release. The Ca2+i release induced by 250 nM IP3 could only be inhibited by prior injection of 1 mg/ml heparin which was overcome by continuous injection of IP3 to 1 μM. Prior injection of either 50 μM ruthenium red, 50 μM procaine or 1 % vehicle medium (VM) did not affect the Ca2+i release induced by IP3. Prior injection of heparin or VM did not affect the Ca2+i release induced by 10-20 mM caffeine or 200 μM ryanodine, but prior injection of 50 μM ruthenium red or procaine completely inhibited the effect of 10-20 mM caffeine. In addition, continuous injection of caffeine up to 40 mM overcame the inhibitory effect of ruthenium red or procaine. The same 50 μM concentration of ruthenium red or procaine only partially blocked the effect of 200 μM ryanodine, but 200 μM ruthenium red or procaine completely blocked the effect of 200 μM ryanodine. Oocytes were refractory for 15 minutes to further injections of IP3 after the initiation of Ca2+i release induced by 200 nM IP3; a 10 minute refractory period was observed for 10-20 mM caffeine and 200 μM ryanodine; either caffeine or ryanodine can desensitize RYRs to the other. However, the desensitization of IP3Rs by 200 nM IP3 does not abolish the effect of 200 μM ryanodine, as effectively as the reciprocal treatment. Prior injection of a subthreshold concentration of ryanodine itself only induced a slight increase in Ca2+i level, but it sensitized the RYR to a subsequent injection of a subthreshold concentration of caffeine. Similar results were obtained when ryanodine was first injected followed by injection of caffeine Based on these results, we conclude that independent IP3Rs and RYRs exist in mature bovine oocytes.
Localization of oskar RNA regulates oskar translation and requires Oskar protein
Authors: Rongo Christopher , Gavis Elizabeth R. , Lehmann Ruth
Published: April 2021

The site of oskar RNA and protein localization within the oocyte determines where in the embryo primordial germ cells form and where the abdomen develops. Initiation of oskar RNA localization requires the activity of several genes. We show that ovaries mutant for any of these genes lack Oskar protein. Using various transgenic constructs we have determined that sequences required for oskar RNA localization and translational repression map to the oskar 3′UTR, while sequences involved in the correct temporal activation of translation reside outside the oskar 3′UTR. Upon localization of oskar RNA and protein at the posterior pole, Oskar protein is required to maintain local- ization of oskar RNA throughout oogenesis. Stable anchoring of a transgenic reporter RNA at the posterior pole is disrupted by oskar nonsense mutations. We propose that initially localization of oskar RNA permits translation into Oskar protein and that subsequently Oskar protein regulates its own RNA localization through a positive feedback mechanism.
four-jointed is required for intermediate growth in the proximal-distal axis in Drosophila
Authors: Villano J. Lee , Katz Flora N.
Published: April 2021

Genes capable of translating positional information into regulated growth lie at the heart of morphogenesis, yet few genes with this function have been identified. Mutants in the Drosophila four-jointed (fj) gene show reduced growth and altered differentiation only within restricted sectors of the proximal-distal (PD) axis in the leg and wing, thus fj is a candidate for a gene with this coordination function. Consistent with a position-sensitive role, we show that fj is expressed in a regional pattern in the developing leg, wing, eye and optic lobe. The fj gene encodes a novel type II membrane glycoprotein. When the cDNA is translated in an in vitro translation system in the presence of exogenous microsomal membranes, the intralumenal portion of some of the molecules is cleaved, yielding a secreted C-terminal fragment. We propose that fj encodes a secreted signal that functions as a positive regulator of regional growth and differentiation along the PD axis of the imaginal discs.
Mutations affecting development of the zebrafish inner ear and lateral line
Authors: Whitfield Tanya T. , Granato Michael , Eeden Fredericus J. M. van , Schach Ursula , Brand Michael , Furutani-Seiki Makoto , Haffter Pascal , Hammerschmidt Matthias , Heisenberg Carl-Philipp , Jiang Yun-Jin , Kane Donald A. , Kelsh Robert N. , Mullins Mary C. , Odenthal Jörg , Nüsslein-Volhard Christiane
Published: April 2021

Mutations giving rise to anatomical defects in the inner ear have been isolated in a large scale screen for mutations causing visible abnormalities in the zebrafish embryo (Haffter, P., Granato, M., Brand, M. et al. (1996)Development 123, 1-36). 58 mutants have been classified as having a primary ear phenotype; these fall into several phenotypic classes, affecting presence or size of the otoliths, size and shape of the otic vesicle and formation of the semicircular canals, and define at least 20 complementation groups. Mutations in seven genes cause loss of one or both otoliths, but do not appear to affect development of other structures within the ear. Mutations in seven genes affect morphology and patterning of the inner ear epithelium, including formation of the semicircular canals and, in some, development of sensory patches (maculae and cristae). Within this class, dog-eared mutants show abnormal development of semicircular canals and lack cristae within the ear, while in van gogh, semicircular canals fail to form altogether, resulting in a tiny otic vesicle containing a single sensory patch. Both these mutants show defects in the expression of homeobox genes within the otic vesicle. In a further class of mutants, ear size is affected while patterning appears to be relatively normal; mutations in three genes cause expansion of the otic vesicle, while in little ears and microtic, the ear is abnormally small, but still contains all five sensory patches, as in the wild type. Many of the ear and otolith mutants show an expected behavioural phenotype: embryos fail to balance correctly, and may swim on their sides, upside down, or in circles. Several mutants with similar balance defects have also been isolated that have no obvious structural ear defect, but that may include mutants with vestibular dysfunction of the inner ear (Granato, M., van Eeden, F. J. M., Schach, U. et al. (1996)Development, 123, 399-413,). Mutations in 19 genes causing primary defects in other structures also show an ear defect. In particular, ear phenotypes are often found in conjunction with defects of neural crest derivatives (pigment cells and/or cartilaginous elements of the jaw). At least one mutant, dog-eared, shows defects in both the ear and another placodally derived sensory system, the lateral line, while hypersensitive mutants have additional trunk lateral line organs.
Left-right pattern of cardiac BMP4 may drive asymmetry of the heart in zebrafish
Authors: Chen Jau-Nian , Eeden Fredericus J. M. van , Warren Kerri S. , Chin Alvin , Nüsslein-Volhard Christiane , Haffter Pascal , Fishman Mark C.
Published: April 2021

The first evident break in left-right symmetry of the primitive zebrafish heart tube is the shift in pattern of BMP4 expression from radially symmetric to left-predominant. The midline heart tube then ‘jogs’ to the left and subsequently loops to the right. We examined 279 mutations, affecting more than 200 genes, and found 21 mutations that perturb this process. Some cause BMP4 to remain radially symmetric. Others randomize the asymmetric BMP4 pattern. Retention of BMP4 symmetry is associated with failure to jog: right-predominance of the BMP4 pattern is associated with reversal of the direction of jogging and looping. Raising BMP4 diffusely throughout the heart, via sonic hedgehog injection, or the blocking of its action by injection of a dominant negative BMP4 receptor, prevent directional jogging or looping. The genes crucial to directing cardiac asymmetry include a subset of those needed for patterning the dorsoventral axis and for notochord and ventral spinal cord development. Thus, the pattern of cardiac BMP4 appears to be in the pathway by which the heart interprets lateralizing signals from the midline.
MyoD and Myf-5 differentially regulate the development of limb versus trunk skeletal muscle
Authors: Kablar Boris , Krastel Kirsten , Ying Chuyan , Asakura Atsushi , Tapscott Stephen J. , Rudnicki Michael A.
Published: April 2021

The myogenic progenitors of epaxial (paraspinal and intercostal) and hypaxial (limb and abdominal wall) musculature are believed to originate in dorsal-medial and ventral-lateral domains, respectively, of the developing somite. To investigate the hypothesis that Myf-5 and MyoD have different roles in the development of epaxial and hypaxial musculature, we further characterized myogenesis in Myf-5- and MyoD-deficient embryos by several approaches. We examined expression of a MyoD-lacZ transgene in Myf-5 and MyoD mutant embryos to characterize the temporal-spatial patterns of myogenesis in mutant embryos. In addition, we performed immunohistochemistry on sectioned Myf-5 and MyoD mutant embryos with antibodies reactive with desmin, nestin, myosin heavy chain, sarcom-eric actin, Myf-5, MyoD and myogenin. While MyoD-- embryos displayed normal development of paraspinal and intercostal muscles in the body proper, muscle development in limb buds and brachial arches was delayed by about 2.5 days. By contrast, Myf-5−/− embryos displayed normal muscle development in limb buds and brachial arches, and markedly delayed development of paraspinal and intercostal muscles. Although MyoD mutant embryos exhibited delayed development of limb musculature, normal migration of Pax-3-expressing cells into the limb buds and normal subsequent induction of Myf-5 in myogenic precursors was observed. These results suggest that Myf-5 expression in the limb is insufficient for the normal progression of myogenic development. Taken together, these observations strongly support the hypothesis that Myf-5 and MyoD play unique roles in the development of epaxial and hypaxial muscle, respectively.
Establishing primordia in the Drosophila eye-antennal imaginal disc: the roles of decapentaplegic, wingless and hedgehog
Authors: Royet Julien , Finkelstein Robert
Published: April 2021

The eye-antennal imaginai discs of Drosophila melanogaster form the head capsule of the adult fly. Unlike the limb primordia, each eye-antennal disc gives rise to morphologically and functionally distinct structures. As a result, these discs provide an excellent model system for determining how the fates of primordia are specified during development.

In this study, we investigated how the adjacent primordia of the compound eye and dorsal head vertex are specified. We show that the genes wingless (wg) and orthodenticle (otd) are expressed throughout the entire second instar eye-antennal disc, conferring a default fate of dorsal vertex cuticle. Activation of decapentaplegic (dpp) expression in the posterior eye disc eliminates wg and otd expression, thereby permitting eye differentiation. We also demonstrate that otd is activated by wg in the vertex primordium. Finally, we show that early activation of dpp depends on hedgehog (hh) expression in the eye anlage prior to morphogenetic furrow formation.
Post-transcriptional regulation of gurken by encore is required for axis determination in Drosophila
Authors: Hawkins Nancy C. , Buskirk Cheryl Van , Grossniklaus Ueli , Schüpbach Trudi
Published: April 2021

Establishment of anterior-posterior and dorsal-ventral polarity within the Drosophila egg chamber requires signaling between the germline and the somatic cells of the ovary. The gene gurken (grk) encodes a TGFoi-like protein that is localized within the developing oocyte and is thought to locally activate torpedo/Egfr (top/Egfr), the Drosophila homolog of the EGF receptor, which is expressed throughout the follicular epithelium surrounding the oocyte. grk-Egfr signaling is required early in oogenesis for specification of posterior follicle cell fate and later in oogenesis for dorsal follicle cell fate determination, thus establishing the axes of the egg shell and embryo.

Previous studies have shown that these patterning processes are highly sensitive to changes in the levels and localization of grk mRNA. Here we show that post-transcriptional regulation of Grk protein levels is required for correct pattern formation. encore (enc), a gene that functions in the regulation of germline mitosis and maintenance of oocyte identity, is also required for the accumulation of Grk protein during oogenesis. We present evidence that enc regulates Grk post-transcriptionally to ensure adequate levels of signaling for establishment of the anterior-posterior and dorsal-ventral axes.
Spitz and Wingless, emanating from distinct borders, cooperate to establish cell fate across the Engrailed domain in the Drosophila epidermis
Authors: O’Keefe Louise , Dougan Scott T. , Gabay Limor , Raz Erez , Shilo Ben-Zion , DiNardo Stephen
Published: April 2021

A key step in development is the establishment of cell type diversity across a cellular field. Segmental patterning within the Drosophila embryonic epidermis is one paradigm for this process. At each parasegment boundary, cells expressing the Wnt family member Wingless confront cells expressing the homeoprotein Engrailed. The Engrailed-expressing cells normally differentiate as one of two alternative cell types. In investigating the generation of this cell type diversity among the 2-cell-wide Engrailed stripe, we previously showed that Wingless, expressed just anterior to the Engrailed cells, is essential for the specification of anterior Engrailed cell fate. In a screen for additional mutations affecting Engrailed cell fate, we identified anterior open/yan, a gene encoding an inhibitory ETS-domain transcription factor that is negatively regulated by the Ras1-MAP kinase signaling cascade. We find that Anterior Open must be inactivated for posterior Engrailed cells to adopt their correct fate. This is achieved by the EGF receptor (DER), which is required autonomously in the Engrailed cells to trigger the Ras1-MAP kinase pathway. Localized activation of DER is accomplished by restricted processing of the activating ligand, Spitz. Processing is confined to the cell row posterior to the Engrailed domain by the restricted expression of Rhomboid. These cells also express the inhibitory ligand Argos, which attenuates the activation of DER in cell rows more distant from the ligand source. Thus, distinct signals flank each border of the Engrailed domain, as Wingless is produced anteriorly and Spitz posteriorly. Since we also show that En cells have the capacity to respond to either Wingless or Spitz, these cells must choose their fate depending on the relative level of activation of the two pathways.
Roles of the imprinted gene Igf2 and paternal duplication of distal chromosome 7 in the perinatal abnormalities of androgenetic mouse chimeras
Authors: McLaughlin K. John , Kochanowski Helga , Solter Davor , Schwarzkopf Georg , Szabó Piroska E. , Mann Jeffrey R.
Published: April 2021

Mouse chimeras made with androgenetic (two paternal genomes) ova or embryonic stem cells frequently die at the perinatal stage and exhibit a range of defects, the most noticeable being a pronounced overgrowth of rib cartilage. Excess concentrations of IGFII, a potent mitogen, has been suggested to play a major role in these defects, as andro-genetic cells possess two active paternal copies of the imprinted Igf2 gene, rather than one inactive maternal and one active paternal copy as in normal cells. Here, we show that chimeras made with androgenetic embryonic stem cells, homozygous for an Igf2 null mutation, do not develop rib cartilage hyperplasia, demonstrating the dependence of this defect on Igf2 activity produced by androgenetic cells. In contrast, in these same chimeras, many other defects, including whole body overgrowth and perinatal death, are still prevalent, indicating that the abnormal expression of one or more imprinted genes, other than Igf2, is also capable of inducing most of the defects of androgenetic chimeras. Many of these genes may reside on distal chromosome 7, as we also show that perinatal chimeras made with embryonic stem cells possessing paternal duplication of distal chromosome 7 exhibit a range of defects similar to those of androgenetic chimeras. The relevance of these findings for the human imprinting-related disorder, Beckwith-Wiedemann syndrome, is discussed.
FIGα, a germ cell specific transcription factor involved in the coordinate expression of the zona pellucida genes
Authors: Liang Li-fang , Soyal Selma M. , Dean Jurrien
Published: April 2021

The mouse zona pellucida is composed of three glycoproteins, ZP1, ZP2 and ZP3, encoded by single-copy genes whose expression is temporally and spatially restricted to oocytes. All three proteins are required for the formation of the extracellular zona matrix and female mice with a single disrupted zona gene lack a zona and are infertile. An E-box (CANNTG), located approximately 200 bp upstream of the transcription start sites of Zp1, Zp2 and Zp3, forms a protein-DNA complex present in oocytes and, to a much lesser extent, in testes. It has been previously shown that the integrity of this E-box in Zp2 and Zp3 promoters is required for expression of luciferase reporter genes microinjected into growing oocytes. The presence of the ubiquitous transcription factor E12 in the complex was used to identify a novel basic helix-loop-helix protein, FIGot (Factor In the Germline alpha) whose expression was limited to oocytes within the ovary. The ability of FIGa to transactivate reporter genes coupled to each of the three mouse zona promoters in heterologous embryonic fibroblasts suggests a role in coordinating the expression of the three zona pellucida genes during oogenesis.
Early specification and autonomous development of cortical fields in the mouse hippocampus
Authors: Tole Shubha , Christian Caprice , Grove Elizabeth A.
Published: April 2021

Studies of the specification of distinct areas in the developing cerebral cortex have until now focused mainly on neocortex. We demonstrate that the hippocampus, an archicortical structure, offers an elegant, alternative system in which to explore cortical area specification. Individual hippocampal areas, called CA fields, display striking molecular differences in maturity. We use these distinct patterns of gene expression as markers of CA field identity, and show that the two major hippocampal fields, CA1 and CA3, are specified early in hippocampal development, during the period of neurogenesis. Two field-specific markers display consistent patterns of expression from the embryo to the adult. Presumptive CA1 and CA3 fields (Pca1, Pca3) can therefore be identified between embryonic days 14.5 and 15.5 in the mouse, a week before the fields are morphologically distinct. No other individual cortical areas have been detected by gene expression as early in development. Indeed, other features that distinguish between the CA fields appear after birth, indicating that mature CA field identity is acquired over at least 3 weeks. To determine if Pca1 and Pca3 are already specified to acquire mature CA field identities, the embryonic fields were isolated from further potential specification cues by maintaining them in slice culture. CA field development proceeds in slices of the entire embryonic hippocampus. More strikingly, slices restricted to Pca1 or Pca3 alone also develop appropriate mature features of CA1 or CA3. Pca1 and Pca3 are therefore able to develop complex characteristics of mature CA field identity autonomously, that is, without contact or innervation from other fields or other parts of the brain. Because Pca1 and Pca3 can be identified before major afferents grow into the hippocampus, innervation may also be unnecessary for the initial division of the hippocampus into separate fields. Providing a clue to the source of the true specifying signals, the earliest field markers appear first at the poles of the hippocampus, then progress inwards. General hippocampal development does not follow this pronounced pattern. We suggest that the sources of signals that specify hippocampal field identity lie close to the hippocampal poles, and that the signals operate first on cells at the poles, then move inwards.
Determination of the migratory capacity of embryonic cortical cells lacking the transcription factor Pax-6
Authors: Carić Damira , Gooday Douglas , Hill Robert E. , McConnell Susan K. , Price David J.
Published: April 2021

The cerebral cortex forms by the orderly migration and subsequent differentiation of neuronal precursors generated in the proliferative ventricular zone. We studied the role of the transcription factor Pax-6, which is expressed in the ventricular zone, in cortical development. Embryos homozygous for a mutation of Pax-6 (Small eye; Sey) had abnormalities suggesting defective migration of late-born cortical precursors. When late-born Sey/Sey precursors were transplanted into wild-type embryonic rat cortex, they showed similar integrative, migrational and differentiative abilities to those of transplanted wild-type mouse precursors. These results suggest that postmitotic cortical cells do not need Pax-6 to acquire the capacity to migrate and differentiate, but that Pax-6 generates a cortical environment that permits later-born precursors to express their full developmental potential.
Sectors expressing the homeobox gene liguleless3 implicate a timedependent mechanism for cell fate acquisition along the proximal-distal axis of the maize leaf
Authors: Muehlbauer Gary J. , Fowler John E. , Freeling Michael
Published: April 2021

The longitudinal axis of the maize leaf is composed of, in proximal to distal order, sheath, ligule, auricle and blade. The semidominant Liguleless3-O (Lg3-O) mutation disrupts leaf development at the ligular region of the leaf midrib by transforming blade to sheath. In a previous study, we showed that leaf sectors of Lg3 mutant activity are cell nonautonomous in the transverse dimension and can confer several alternative developmental fates (Fowler, Muehlbauer and Freeling (1996)Genetics 143, 489-503). In our present study we identify five Lg3 sector types in the leaf: sheath-like with displaced ligule (sheath-like), sheathlike with ectopic ligule (ectopic ligule), auricle-like, macrohairless blade and wild-type blade. The acquisition of a specific sector fate depends on the timing of Lg3 expression. Early Lg3 expression results in adoption of the sheath-like phenotype at the ligule position (a proximal cell fate), whereas later Lg3 expression at the same position results in one of the more distal cell fates. Furthermore, sheathlike Lg3 sectors exhibit a graded continuum of phenotypes in the transformed blade region from the most proximal (sheath) to the most distal (wild-type blade), suggesting that cell fate acquisition is a gradual process. We propose a model for leaf cell fate acquisition based on a timing mechanism whereby cells of the leaf primordium progress through a maturation schedule of competency stages which eventually specify the cell types along the proximal to distal axis of the leaf. In addition, the lateral borders between Lg3 ‘on’ sectors and wild-type leaf sometimes provide evidence of no spreading of the transformed phenotype. In these cases, competency stages are inherited somatically.
Functional analysis of an ascidian homologue of vertebrate Bmp-2/Bmp-4 suggests its role in the inhibition of neural fate specification
Authors: Miya Takahito , Morita Kiyokazu , Suzuki Atsushi , Ueno Naoto , Satoh Noriyuki
Published: April 2021

The ascidian tadpole larva is thought to be close to a prototype of the ancestral chordate. The vertebrate body plan is established by a series of inductive cellular interactions, whereas ascidians show a highly determinate mode of development. Recent studies however, suggest some roles of cell-cell interaction during ascidian embryogenesis. To elucidate the signaling molecules responsible for the cellular interaction, we isolated HrBMPb, an ascidian homologue of the vertebrate bone morphogenetic protein (BMP) gene, from Halocynthia roretzi. The amino acid sequence of HrBMPb closely resembled those of vertebrate BMP-2 and BMP-4 and of Drosophila Decapentaplegic (DPP). In addition to the sequence similarity, HrBMPb overexpression induced the ventralization of Xenopus embryos, suggesting functional conservation. The zygotic expression of HrBMPb was first detected around gastrulation. HrBMPb expression was maintained in some cells at the lateral edges of the neural plate through gastrulation to neurulation, although that in the presumptive muscle cells was downregulated. HrBMPb was not expressed in the presumptive epidermis during gastrulation. When HrBMPb mRNA was injected into fertilized Halocynthia eggs, cells that normally give rise to the neural tissue differentiated into epidermis, causing a loss of anterior neural tissue in the larva. In addition, HrBMPb might function synergistically with HrBMPa, an ascidian homologue of BMPs-5 to 8. However, HrBMPb overexpression did not affect differentiation of the notochord and muscle cells. These results suggest that HrBMPb functions as a neural inhibitor and as an epidermal inducer but not as a ventralizing agent in ascidian development.
Axial specification for sensory organs versus non-sensory structures of the chicken inner ear
Authors: Wu Doris K. , Nunes Fabio D. , Choo Daniel
Published: April 2021

A mature inner ear is a complex labyrinth containing multiple sensory organs and nonsensory structures in a fixed configuration. Any perturbation in the structure of the labyrinth will undoubtedly lead to functional deficits. Therefore, it is important to understand molecularly how and when the position of each inner ear component is determined during development. To address this issue, each axis of the otocyst (embryonic day 2.5, E2.5, stage 16-17) was changed systematically at an age when axial information of the inner ear is predicted to be fixed based on gene expression patterns. Transplanted inner ears were analyzed at E4.5 for gene expression of BMP4 (bone morphogenetic protein), SOHo-1 (sensory organ homeobox-1), Otx1 (cognate of Drosophila orthodenticle gene), p75NGFR (nerve growth factor receptor) and Msx1 (muscle segment homeobox), or at E9 for their gross anatomy and sensory organ formation. Our results showed that axial specification in the chick inner ear occurs later than expected and patterning of sensory organs in the inner ear was first specified along the anterior/posterior (A/P) axis, followed by the dorsal/ventral (D/V) axis. Whereas the A/P axis of the sensory organs was fixed at the time of transplantation, the A/P axis for most non-sensory structures was not and was able to be re-specified according to the new axial information from the host. The D/V axis for the inner ear was not fixed at the time of transplantation. The asynchronous specification of the A/P and D/V axes of the chick inner ear suggests that sensory organ formation is a multi-step phenomenon, rather than a single inductive event.
Cell lineage in marine nematode Enoplus brevis
Authors: Voronov Dmitrii A. , Panchin Yuri V.
Published: April 2021

Early cleavages of the marine nematode Enoplus brevis are symmetrical and occur in synchrony. At the 2- to 16-cell stages, blastomeres are indistinguishable. The progeny of blastomeres was investigated by intracellular injections of fluorescent dyes and horse radish peroxidase. One blastomere of the 2-cell embryo gives rise to a compact group of cells occupying about half of an embryo. The border between labeled and unlabeled cells differs in each embryo dividing it to anterior-posterior, left-right or intermediate parts. At the 8-cell stage, one blastomere gives rise to only endoderm, whereas the other blastomeres produce progeny that form multiple cell types, including nerve, muscle and hypoderm cells, in various proportions. Thus the fates of the blastomeres of early E. brevis embryos, with the exception of the endoderm precursor, are not determined. The process of gastrulation in E. brevis is very similar to that in Caenorhabditis elegans and other nematodes. At the beginning of gastrulation, the 2-celled endoderm precursor lies on the surface of embryo and then sinks inwards. After labeling of cells on the ventral side (near endoderm precursor) at the beginning of gastrulation, their progeny differentiate predominantly into body muscles or pharyngeal cells of the first stage larva. Cells that are located more laterally give rise mainly to neurons. The dorsal blastomeres differentiated principally into hypoderm cells. Our study suggests that a precise cell lineage is not a necessary attribute of nematode development.
δEF1, a zinc finger and homeodomain transcription factor, is required for skeleton patterning in multiple lineages
Authors: Takagi Tsuyoshi , Moribe Hiroki , Kondoh Hisato , Higashi Yujiro
Published: April 2021

δEF1 is a DNA binding protein containing a homeodomain and two zinc finger clusters, and is regarded as a vertebrate homologue of zfh-1 (zinc finger homeodomain-containing factor-1) in Drosophila. In the developing embryo, δEF1 is expressed in the notochord, somites, limb, neural crest derivatives and a few restricted sites of the brain and spinal cord. To elucidate the regulatory function of δEF1 in mouse embryogenesis, we generated δEF1 null mutant (δEF1null(lacZ)) mice. The δEF1null(lacZ) homozygotes developed to term, but never survived postnatally. In addition to severe T cell deficiency of the thymus, the δEF1null(lacZ) homozygotes exhibited skeletal defects of various lineages. (1) Craniofacial abnormalities of neural crest origin: cleft palate, hyperplasia of Meckel’s cartilage, dysplasia of nasal septum and shortened mandible. (2) Limb defects: shortening and broadening of long bones, fusion of carpal/tarsal bone and fusion of joints. (3) Fusion of ribs. (4) Sternum defects: split and asymmetric ossification pattern of the sternebrae associated with irregular sternocostal junctions. (5) Hypoplasia of intervertebral discs. These results indicate that δEF1 has an essential role in regulating development of these skeletal structures. Since the skeletal defects were not observed in δEF1ΔC727 mice, δEF1 bears distinct regulatory activities which are dependent on different domains of the molecule.
TGFβ3 promotes transformation of chicken palate medial edge epithelium to mesenchyme in vitro
Authors: Sun Dazhong , Vanderburg Charles R. , Odierna Gregory S. , Hay Elizabeth D.
Published: April 2021

Epithelial-mesenchymal transformation plays an important role in the disappearance of the midline line epithelial seam in rodent palate, leading to confluence of the palate. The aim of this study was to test the potential of the naturally cleft chicken palate to become confluent under the influence of growth factors, such as TGFβ3, which are known to promote epithelial-mesenchymal transformation. After labeling medial edge epithelia with carboxyfluorescein, palatal shelves (E8-9) with or without beak were dissected and cultured on agar gels. TGFβ1, TGFβ2 or TGFβ3 was added to the chemically defined medium. By 24 hours in culture, medial edge epithelia form adherent midline seams in all paired groups without intact beaks. After 72 hours, seams in the TGFβ3 groups disappear and palates become confluent due to epithelial-mesenchymal transformation, while seams remain mainly epithelial in control, TGFβ1 and TGFβ2 groups. Epithelium-derived mesenchymal cells are identified by carboxyfluorescein fluorescence with confocal microscopy and by membrane-bound carboxyfluorescein isolation bodies with electron microscopy. Labeled fibroblasts completely replace the labeled epithelia of origin in TGFβ3-treated palates without beaks. Single palates are unable to undergo transformation, and paired palatal shelves with intact beaks do not adhere or undergo transformation, even when treated with TGFβ3. Thus, physical contact of medial edge epithelia and formation of the midline seam are necessary for epithelial-mesenchymal transformation to be triggered. We conclude that there may be no fundamental difference in developmental potential of the medial edge epithelium for transformation to mesenchyme among reptiles, birds and mammals. The bird differs from other amniotes in having developed a beak and associated craniofacial structures that seemingly keep palatal processes separated in vivo. Even control medial edge epithelia partly transform to mesenchyme if placed in close contact. However, exogenous TGFβ3 is required to achieve complete confluence of the chicken palate.
The Drosophila sanpodo gene controls sibling cell fate and encodes a tropomodulin homolog, an actin/tropomyosin-associated protein
Authors: Dye Catherine A. , Lee Ja-Kyeong , Atkinson Richard C. , Brewster Rachel , Han Pyung-Lim , Bellen Hugo J.
Published: April 2021

Notch signaling is required in many invertebrate and vertebrate cells to promote proper cell fate determination. Mutations in sanpodo cause many different neuronal peripheral nervous system precursor cells to generate two identical daughter neurons, instead of a neuron and sibling cell. This phenotype is similar to that observed when Notch function is lost late in embryonic development and opposite to the numb loss-of-function phenotype. Genetic interaction studies show that sanpodo is epistatic to numb. Sanpodo encodes a homolog of tropomodulin, an actin/tropomyosin-associated protein. Loss of sanpodo leads to an aberrant F-actin distribution and causes differentiation defects of actin-containing sensory structures. Our data suggest that an actin-based process is involved in Notch signaling.