bluuebunny/crossref_metadata_2025 huggingface dataset:
lin-12 and glp-1 are required zygotically for early embryonic cellular interactions and are regulated by maternal GLP-1 signaling in Caenorhabditis elegans
Authors: Moskowitz Ivan P. G. , Rothman Joel H.
Published: April 2021

Cell-cell interactions mediated by LIN-12 and GLP-1, members of the LNG (LIN-12, Notch, GLP-1) family of receptors, are required to specify numerous cell fates during development of the nematode Caenorhabditis elegans. Maternally expressed GLP-1 participates in two of at least four sequential inductive interactions that specify the fates of early embryonic descendants of the AB founder cell. We report that GLP-1 and LIN-12, and apparently their ligand, LAG-2, as well as a downstream component, LAG-1, are required in the latter two inductions. We find that LAG-2 is expressed in the signaling cells and LIN-12 is expressed in cells receiving the inductions, consistent with their proposed roles as ligand and receptor, respectively.

Furthermore, we report that maternal GLP-1 activity is required (1) to repress early zygotic lag-2 expression and (2) to activate zygotic lin-12 expression in the early embryo. The patterning of both receptor and ligand expression by maternal GLP-1 signaling establishes competence for the zygotic LNG-mediated cellular interactions and localizes these interactions to the appropriate cells. We propose that activation of maternal GLP-1 regulates zygotic lin-12 and lag-2 expression by a regulatory mechanism analogous to that described for the post-embryonic gonad.
cdh-3, a gene encoding a member of the cadherin superfamily, functions in epithelial cell morphogenesis in Caenorhabditis elegans
Authors: Pettitt Jonathan , Wood William B. , Plasterk Ronald H. A.
Published: April 2021

Several genes that encode members of the cadherin super-family have been identified in Caenorhabditis elegans. Based on the roles of cadherins in vertebrates and Drosophila, it is expected that they function in the control of epithelial morphogenesis, an event which is poorly understood at the molecular level in C. elegans. Reporter genes under the control of upstream sequences from one of these genes, cdh-3, are expressed in developing epithelial cells, but also in a number of neuroectodermal cells that extend processes along some of these epithelial cells. We generated a loss-of-function mutation in cdh-3 by transposon-mediated deletion mutagenesis. This mutation affects the morphogenesis of a single cell, hyp10, which forms the tip of the nematode tail. The lack of detectable defects associated with the other cells expressing cdh-3 reporter constructs hints at the existence of other genes that can compensate for cdh-3 loss of function.
Function of the Drosophila POU domain transcription factor Drifter as an upstream regulator of Breathless receptor tyrosine kinase expression in developing trachea
Authors: Anderson Michael G. , Certel Sarah J. , Certel Kaan , Lee Tzumin , Montell Denise J. , Johnson Wayne A.
Published: April 2021

Organogenesis of the Drosophila tracheal system involves extensive directed cell migrations leading to a stereotypic series of interconnected tubules. Although numerous gene products have been shown to be essential for tracheal morphogenesis, direct functional relationships between participants have not been previously established. Both the breathless gene, encoding a Drosophila fibroblast growth factor receptor tyrosine kinase homologue, and the POU-domain transcription factor gene, drifter, are expressed in all tracheal cells and are essential for directed cell migrations. We demonstrate here that ubiquitously expressed Breathless protein under control of a heterologous heat-shock promoter is able to rescue the severely disrupted tracheal phenotype associated with drifter loss-of-function mutations. In the absence of Drifter function, breathless expression is initiated normally but transcript levels fall drastically to undetectable levels as tracheal differentiation proceeds. In addition, breathless regulatory DNA contains seven high affinity Drifter binding sites similar to previously identified Drifter recognition elements. These results suggest that the Drifter protein, which maintains its own expression through a tracheal-specific autoregulatory enhancer, is not necessary for initiation of breathless expression but functions as a direct transcriptional regulator necessary for maintenance of breathless transcripts at high levels during tracheal cell migration. This example of a mechanism for maintenance of a committed cell fate offers a model for understanding how essential gene activities can be maintained throughout organogenesis.
Functional cooperation between the non-paralogous genes Hoxa-10 and Hoxd-11 in the developing forelimb and axial skeleton
Authors: Favier Bertrand , Rijli Filippo M. , Fromental-Ramain Catherine , Fraulob Valérie , Chambon Pierre , Dollé Pascal
Published: April 2021

The Abdominal B-related Hoxa-10 gene displays similar expression patterns in the differentiating forelimbs and hindlimbs of the mouse, with preferential expression around the humeral and femoral cartilages and more diffuse expression in distal regions. We found that a targeted disruption of Hoxa-10 has almost no effect in the forelimbs, while it affects the proximal hindlimb skeleton. The alterations were located along the dorsolateral side of the femur (labium laterale), with an enlargement and distal shift of the third trochanter, a misshapen lateral knee sesamoid, a supernumerary ‘ligament’ connecting these structures and an occasional duplication of the femoral trochlea. Some Hoxa-10−/− mutant mice developed severe degenerative alterations of the knee articulation upon ageing. Viable Hoxa-10/Hoxd-11 double mutant mice were produced by genetic intercrosses. The compound mutation resulted in synergistic forelimb phenotypic alterations, consisting of: (i) an exacerbation of Hoxd-11−/− phenotypic traits in the carpal and digital region, e.g. more pronounced truncations of the ulna styloid, pyramidal and pisiform bones and of some metacarpal and phalangeal bones and (ii) marked alterations in a more proximal region which is nearly unaffected in Hoxd-11−/− single mutants; the entire radius and ulna were truncated and thickened, with deformations of the ulna proximal extremity. Thus, functional redundancy can occur even between non-paralogous Abdominal B-related Hox genes. The double Hoxa-10/Hoxd-11 mutation also conferred full penetrance to the sacral and caudal vertebrae transforma-tions which are ∽50% penetrant in Hoxd-11−/− single mutants, revealing that functional cooperation can also occur between non-paralogous Hox gene products in axial skeleton patterning.
Specific and redundant functions of the paralogous Hoxa-9 and Hoxd-9 genes in forelimb and axial skeleton patterning
Authors: Fromental-Ramain Catherine , Warot Xavier , Lakkaraju Sudhakar , Favier Bertrand , Haack Herbert , Birling Céline , Dierich Andrée , Dollé Pascal , Chambon Pierre
Published: April 2021

Using gene targeting, we have produced mice with a disruption of Hoxa-9 or Hoxd-9, two paralogous Abdominal B-related genes. During embryogenesis, these genes are expressed in limb buds and along the vertebral axis with anterior expression boundaries at the level of prevertebra #20 for Hoxa-9 and #23 for Hoxd-9. Skeletal analysis revealed homeotic transformations corresponding to ante-riorisations of vertebrae #21 to #25 (L1 to L5) in the lumbar region of Hoxa-9−/− mutants; vertebrae #23 to #25 (L3 to L5) in the lumbar region together with vertebrae #28, #30 and #31 (S2, S4 and Ca1) in the sacrum and tail were ante-riorized in Hoxd-9−/− mutants. Thus, anteriorisation of vertebrae #23 to #25 were common to both phenotypes. Subtle forelimb (but not hindlimb) defects, corresponding to a reduction of the humerus length and malformation of its deltoid crest, were also observed in Hoxd-9−/−, but not in Hoxa-9−/−, mutant mice. By intercrosses between these two lines of mutant mice, we have produced Hoxa-9/Hoxd-9 double mutants which exhibit synergistic limb and axial malformations consisting of: (i) an increase of penetrance and expressivity of abnormalities present in the single mutants, and (ii) novel limb alterations at the level of the forelimb stylopod and additional axial skeleton transfor-mations. These observations demonstrate that the two paralogous genes Hoxa-9 and Hoxd-9 have both specific and redundant functions in lumbosacral axial skeleton pat-terning and in limb morphogenesis at the stylopodal level. Taken all together, the present and previously reported results show that disruption of different Hox genes can produce similar vertebral transformations, thus support-ing a combinatorial code model for specification of vertebral identity by Hox genes.
Interactions between rhombomeres modulate Krox-20 and follistatin expression in the chick embryo hindbrain
Authors: Graham Anthony , Lumsden Andrew
Published: April 2021

The rhombomeres of the embryonic hindbrain display compartment properties, including cell lineage restriction, genetic definition and modular anatomical phenotype. Consistent with the idea that rhombomeres are autonomous developmental units, previous studies have shown that certain aspects of rhombomere phenotype are determined early, at the time of rhombomere formation. By contrast, the apoptotic depletion of neural crest from rhombomeres 3 and 5 is due to an interaction with their neighbouring rhombomeres, involving the signalling molecule Bmp4. In this paper, we have examined whether inter-rhombomere interactions control further aspects of rhombomere phenotype. We find that the expression of Krox-20 and the repression of follistatin in r3 is dependent upon neighbour interaction, whereas these genes are expressed autonomously in r5. We further demonstrate that modulation of Krox-20 and follistatin expression is not dependent on Bmp4, indicating the existence of multiple pathways of interaction between adjacent rhombomeres. We also show that, although some phenotypic aspects of r3 are controlled by neighbour interactions, the axial identity of the segment is intrinsically determined.
Clonal analysis of the late flowering fca mutant of Arabidopsis thaliana: cell fate and cell autonomy
Authors: Furner I. J. , Ainscough J. F-X. , Pumfrey J. A. , Petty L. M.
Published: April 2021

Plants that are homozygous for the fca mutation bolt and flower later than wild-type (FCA) plants. The mutation has little or no effect on the fate map of the dry seed, except that the central cells give rise to further rosette leaves instead of the bolting stem, cauline leaves and inflorescence. The large and variable sectors affecting the late rosette leaves of fca plants were used to generate an abstract frequency-distance fate map of vegetative growth. The map relates the initiation of leaves in the plant apex to their final arrangements. The map was found to be a shallow dome with phyllotaxy superimposed on its surface.

X-irradiation was used to provoke loss of the FCA allele from cells in heterozygous seeds. The resulting fca sectors had no effect on the plant phenotype. Even when L2 and L3 cells at the centre of the meristem could not produce the FCA gene product, bolting and flowering was unaffected. The genotypically fca mutant tissue was incorporated into phenotypically normal stems, cauline leaves and flowers. Possible reasons for the non-autonomous behaviour of the trait are discussed.
A test for cell autonomy, based on di-cistronic messenger translation
Authors: Hart Karen , Bienz Mariann
Published: April 2021

We have devised a test for cell autonomy of a gene that is switched on ectopically in a clone of cells, allowing us to ask whether the wild-type activity of this gene can influence neighbouring cells. To switch on the test gene, we used the yeast FRT system, and marked the FRT-generated cell clone by co-expressing β-galactosidase. Co-expression is achieved by a stretch of 5′ untranslated mRNA from the homeotic gene Ultrabithorax (Ubx), which is inserted between the two coding sequences. We show that this Ubx sequence mediates efficient and reliable di-cistronic Mrna translation in wing imaginal discs of Drosophila. Applying our test to Ubx, we find that ectopic Ubx in wing discs strictly coincides with β-galactosidase expression. Consequently, wing cells are transformed into cells that appear to be intermediates between wing and haltere cells, contesting the view that homeotic genes act as binary switches.
hedgehog is required for the proliferation and specification of ovarian somatic cells prior to egg chamber formation in Drosophila
Authors: Forbes Alexandria J. , Lin Haifan , Ingham Philip W. , Spradling Allan C.
Published: April 2021

The hedgehog (hh) gene plays a role in regulating cell proliferation and specifying cell identity in diverse systems. We show that hh is expressed at the extreme apical end of Drosophila ovarioles in terminal filament cells and a newly identified group of associated somatic cells. Reducing or ectopically expressing hh affects somatic cells in region 2 of the germarium, 2-5 cells away from the cells in which Hh protein is detected. hh activity stimulates the proliferation of pre-follicle somatic cells, and promotes the specification of polar follicle cells. hh signalling during egg chamber assembly appears to be closely related to, or part of pathways involving the neurogenic genes.
Expression and regulation of Cek-8, a cell to cell signalling receptor in developing chick limb buds
Authors: Patel K. , Nittenberg R. , D’Souza D. , Irving C. , Burt D. , Wilkinson D. G. , Tickle C.
Published: April 2021

The Eph-related receptor tyrosine kinase gene, Cek-8, is expressed in mesenchyme at the tip of chick limb buds, with high levels of transcripts posteriorly and apically but fading out anteriorly. Expression of Cek-8 in distal mesenchyme is regulated by apical ridge- and FGF-polarising signals and retinoic acid, and is uniform across the antero- posterior axis in talpid3 mutants. These data indicate that Cek-8 expression responds to regulatory signals during limb patterning and suggest that this receptor tyrosine kinase may have a role in coordinating responses to signals in the progress zone of early buds. Later on in limb development, Cek-8 expression is associated with cell condensations that form tendons and their attachments to cartilage rudiments and then in developing feather buds.
A mutational analysis of the 5′ HoxD genes: dissection of genetic interactions during limb development in the mouse
Authors: Capecchi Allan Peter Davis and Mario R.
Published: April 2021

Using gene targeting in mice, we have undertaken a systematic mutational analysis of the homeobox-containing 5′ HoxD genes. In particular, we have characterized the limb defects observed in mice with independent targeted disruptions of hoxd-12 and hoxd-13. Animals defective for hoxd-12 are viable, fertile, and appear outwardly normal yet have minor autopodal defects in the forelimb which include a reduction in the bone length of metacarpals and phalanges, and a malformation of the distal carpal bone d4. The limb phenotypes observed in hoxd-13 mutant mice are more extensive, including strong reductions in length, complete absences, or improper segmentations of many metacarpal and phalangeal bones. Additionally, the d4 carpal bone is not properly formed and often produces an extra rudimentary digit. To examine the genetic interactions between the 5′ HoxD genes, we bred these mutant strains with each other and with our previously characterized hoxd-11 mouse to produce a series of trans-heterozygotes. Skeletal analyses of these mice reveal that these genes interact in the formation of the vertebrate limb, since the trans-heterozygotes display phenotypes not present in the individual heterozygotes, including more severe carpal, metacarpal and phalangeal defects. Some of these pheno- types appear to be accounted for by a delay in the ossification events in the autopod, which lead to either the failure of fusion or the elimination of cartilaginous elements. Characteristically, these mutations lead to the overall trun- cation of digits II and V on the forelimb. Additionally, some trans-animals show the growth of an extra postaxial digit VI, which is composed of a bony element resembling a phalange. The results demonstrate that these genes interact in the formation of the limb. In addition to the previously characterized paralogous interactions, a multitude of inter- actions between Hox genes is used to finely sculpt the forelimb. The 5′ Hox genes could therefore act as a major permissive genetic milieu that has been exploited by evolutionary adaptation to form the tetrapod limbs.
Specification of the anteroposterior axis in Caenorhabditis elegans
Authors: Goldstein Bob , Hird Steven N.
Published: April 2021

Anteroposterior asymmetries are apparent in C. elegans development before the first cell division. Here we identify the cue that specifies the anteroposterior axis, and investigate how this cue is interpreted to generate initial asymmetry. In C. elegans, the sperm normally enters the egg in an invariant position. We have found that causing fertilisation to occur in the abnormal end of the egg completely reverses the orientation of the anteroposterior axis, but gives otherwise normal development. This result suggests that a component of the sperm normally specifies the anteroposterior axis. We have found that a cytoplasmic rearrangement in the uncleaved zygote is directed by the sperm, suggesting a mechanism by which the sperm may specify the axis. The results additionally reveal that the C. elegans oocyte is constructed with no axis prespecified in the form of asymmetrically localised cytoplasmic determinants.
Functional domains in the Deformed protein
Authors: Zhu Aihua , Kuziora Michael A.
Published: April 2021

A chimeric protein consisting of Deformed with a substituted Abdominal-B homeodomain (Dfd/Abd-B) is used to identify protein domains outside the homeodomain that are required for regulatory activity in vivo. A series of deletion proteins were generated based on regions showing amino acid composition similar to known regulatory domains. Each mutant protein can influence regulation of homeotic genes in a manner distinct from the intact protein. Activity was also tested using promoter elements from empty spiracles and Distal-less, two genes known to be directly regulated by Abdominal-B. Removal of the acidic region and the C-tail region convert the chimera from a strong activator to a repressor of the Distal-less element, but had comparatively little effect on the activation of the empty spiracles element. Constructs without a third domain, the N domain, fail to show any regulatory activity. The N domain is the only domain of the Dfd/Abd-B protein which exhibits significant activation activity when fused to a heterologous DNA binding domain. Our results suggest transcriptional activity of the N domain can be modulated by the acidic and C-tail domains.
Surface tensions of embryonic tissues predict their mutual envelopment behavior
Authors: Foty Ramsey A. , Pfleger Cathie M. , Forgacs Gabor , Steinberg Malcolm S.
Published: April 2021

During embryonic development, certain tissues stream to their destinations by liquidlike spreading movements. According to the ‘differential adhesion hypothesis’, these movements are guided by cell-adhesion-generated tissue surface tensions (σs), operating in the same manner as surface tensions do in the mutual spreading behavior of immiscible liquids, among which the liquid of lower surface tension is always the one that spreads over its partner. In order to conduct a direct physical test of the ‘differential adhesion hypothesis’, we have measured the σ s of aggregates of five chick embryonic tissues, using a parallel plate compression apparatus specifically designed for this purpose, and compared the measured values with these tissues’ mutual spreading behaviors. We show that aggregates of each of these tissues behave for a time as elasticoviscous liquids with characteristic surface tension values. Chick embryonic limb bud mesoderm (σ = 20.1 dyne/cm) is enveloped by pigmented epithelium (σ = 12.6 dyne/cm) which, in turn, is enveloped by heart (σ = 8.5 dyne/cm) which, in turn, is enveloped by liver (σ = 4.6 dyne/cm) which, in turn, is enveloped by neural retina (σ = 1.6 dyne/cm). Thus, as predicted, the tissues’ surface tension values fall in the precise sequence required to account for their mutual envelopment behavior.
The Drosophila Polycomb group gene Sex comb on midleg (Scm) encodes a zinc finger protein with similarity to polyhomeotic protein
Authors: Bornemann Douglas , Miller Ellen , Simon Jeffrey
Published: April 2021

The Sex comb on midleg (Scm) gene is a member of the Polycomb group (PcG) of genes in Drosophila melanogaster. The PcG genes encode transcriptional repressors required for proper spatial expression of homeotic genes. We report the isolation of new Scm mutations and the molecular char- acterization of the Scm gene. Scm mRNA is expressed maternally, at peak levels in early embryos and then at lower levels throughout the remainder of development. Scm encodes a putative zinc finger protein of 877 amino acids. Scm protein is similar to polyhomeotic, another member of the PcG, both in the zinc finger region and in a separate C-terminal domain of 60 amino acids, which we term the SPM domain. Sequence analysis of an Scm mutant allele suggests a functional requirement for the SPM domain. Scm protein also bears homology in multiple domains to a mouse protein, Rae-28 (Nomura, M., Takihara, Y. and Shimada, K. (1994) Differentiation 57, 39-50) and to a fly tumor suppressor protein, the product of the lethal(3)malignant brain tumor gene (Wismar, J. et al., (1995) Mech. Dev. 53, 141-154). Possible functional rela- tionships among these proteins and potential biochemical roles for Scm protein in PcG repression are discussed.
Evidence from normal expression and targeted misexpression that Bone Morphogenetic Protein-4 (Bmp-4) plays a role in mouse embryonic lung morphogenesis
Authors: Bellusci Savério , Henderson Randall , Winnier Glenn , Oikawa Tsuyoshi , Hogan Brigid L. M.
Published: April 2021

Epithelial-mesenchymal interactions are critical for the branching and differentiation of the lung, but the mechanisms involved are still unclear. To investigate this problem in mouse embryonic lung, we have studied the temporal and spatial expression of genes implicated in the morpho-genesis of other organs. At 11.5 days p.c., hepatocyte nuclear factor-3β (Hnf-3β) is expressed uniformly through-out the epithelium, while Wnt-2 expression is confined to the distal mesenchyme. Sonic hedgehog (Shh) trancripts are found throughout the epithelium, with high levels in the distal tips of the terminal buds, while bone morphogenetic protein-4 (Bmp-4) transcripts are localized at high levels in the distal tips of the epithelium, with lower levels in the adjacent mesenchyme. Epithelial expression is also seen for Bmp-7, but transcripts are less dramatically upregulated at the distal tips. The Type I Bone morphogenetic protein receptor gene(Bmpr/Tfr-11/Brk-1) is expressed at low levels in the epithelium and in the distal mesenchyme. To investigate the role of Bmp-4 in lung development, we have mis-expressed the gene throughout the distal epithelium of transgenic lungs using a surfactant protein C enhancer/promoter. From 15.5 days p.c., transgenic lungs are smaller than normal, with grossly distended terminal buds and, at birth, contain large air-filled sacs which do not support normal lung function. Labeling with BrdU reveals an inhibition of epithelial proliferation in 15.5 days p.c. transgenic lungs. A small but significant stimulation of proliferation of mesenchymal cells is also observed, but this is accompanied by an increase in cell death. In situ hybridization with riboprobes for the proximal airway marker, CC10, and the distal airway marker, SP-C, shows normal differentiation of bronchiolar Clara cells but a reduction in the number of differentiated Type II cells in transgenic lungs. A model is proposed for the role of BMP4 and other signalling molecules in embryonic lung morphogenesis.
Involvement of FGF-8 in initiation, outgrowth and patterning of the vertebrate limb
Authors: Vogel Astrid , Rodriguez Concepción , Izpisúa-Belmonte Juan-Carlos
Published: April 2021

Fibroblast Growth Factors (FGFs) are signaling molecules that are important in patterning and growth control during vertebrate limb development. Beads soaked in FGF-1, FGF-2 and FGF-4 are able to induce additional limbs when applied to the flank of young chick embryos (Cohn, M.J., Izpisua-Belmonte, J-C., Abud, H., Heath, J. K., Tickle, C. (1995) Cell 80, 739-746). However, biochemical and expression studies suggest that none of these FGFs is the endogenous signal that initiates limb development. During chick limb development, Fgf-8 transcripts are detected in the intermediate mesoderm and subsequently in the prelimb field ectoderm prior to the formation of the apical ectodermal ridge, structures required for limb initiation and outgrowth, respectively. Later on, Fgf-8 expression is restricted to the ridge cells and expression disappears when the ridge regresses. Application of FGF-8 protein to the flank induces the development of additional limbs. Moreover, we show that FGF-8 can replace the apical ectodermal ridge to maintain Shh expression and outgrowth and patterning of the developing chick limb. Furthermore, continuous and widespread misexpression of FGF-8 causes limb truncations and skeletal alterations with phocomelic or achondroplasia phenotype. Thus, FGF-8 appears to be a key signal involved in initiation, outgrowth and patterning of the developing vertebrate limb.
Clustered arrangement of winged helix genes fkh-6 and MFH-1: possible implications for mesoderm development
Authors: Kaestner Klaus H. , Bleckmann Susanne C. , Monaghan A. Paula , Schlöndorff Johannes , Mincheva Antoaneta , Lichter Peter , Schütz Günther
Published: April 2021

The ‘winged helix’ or ‘forkhead’ transcription factor gene family is defined by a common 100 amino acid DNA binding domain which is a variant of the helix-turn-helix motif. Here we describe the structure and expression of the mouse fkh-6 and MFH-1 genes. Both genes are expressed in embryonic mesoderm from the headfold stage onward. Transcripts for both genes are localised mainly to mesenchymal tissues, fkh-6 mRNA is enriched in the mesenchyme of the gut, lung, tongue and head, whereas MFH-1 is expressed in somitic mesoderm, in the endocardium and blood vessels as well as the condensing mesenchyme of the bones and kidney and in head mesenchyme. Both genes are located within a 10 kb region on mouse chromosome 8 at 5.26±2.56 cM telomeric to Actsk1. The close physical linkage of these two winged helix genes is conserved in man, where the two genes map to chromosome 16q22-24. This tandem arrangement suggests the common use of regulatory mechanisms. The fkh-6/MFH-1 locus maps close to the mouse mutation amputated, which is characterised by abnormal development of somitic and facial mesoderm. Based on the expression patterns we suggest that a mutation in MFH-1, not fkh-6 is the possible cause for the amputated phenotype.
Activation of Fgf-4 and HoxD gene expression by BMP-2 expressing cells in the developing chick limb
Authors: Duprez Delphine M. , Kostakopoulou Konstadina , Francis-West Philippa H. , Tickle Cheryll , Brickell Paul M.
Published: April 2021

Bone morphogenetic protein-2 (BMP-2) has been implicated in the polarizing region signalling pathway, which specifies pattern across the antero-posterior of the developing vertebrate limb. Retinoic acid and Sonic Hedgehog (SHH) can act as polarizing signals; when applied anteriorly in the limb bud, they induce mirror-image digit duplications and ectopic Bmp-2 expression in anterior mesenchyme. In addition, the two signals can activate Fgf-4 expression in anterior ridge and HoxD expression in anterior mesenchyme. We tested the role of BMP-2 in this signalling cascade by ectopically expressing human BMP-2 (hBMP-2) at the anterior margin of the early wing bud using a replication defective retroviral vector, and found that ectopic expression of Fgf-4 was induced in the anterior part of the apical ectodermal ridge, followed later by ectopic expression of Hoxd-11 and Hoxd-13 in anterior mesenchyme. This suggests that BMP-2 is involved in regulating Fgf-4 and HoxD gene expression in the normal limb bud. Ectopically expressed hBMP-2 also induced duplication of digit 2 and bifurcation of digit 3, but could not produce the mirror-image digit duplications obtained with SHH-expressing cells. These results suggest that BMP-2 may be involved primarily in maintenance of the ridge, and in the link between patterning and outgrowth of the limb bud.
Alternatively spliced forms of the Drosophila αPS2 subunit of integrin are sufficient for viability and can replace the function of the αPS1 subunit of integrin in the retina
Authors: Roote Carol E. , Zusman Susan
Published: April 2021

The Drosophila inflated (if) gene encodes the αPS2 subunit of the PS family of integrins. The if transcript is spliced such that αPS2 is found in two alternative forms, αPS2(C) and αPS2(m8), which differ by 25 amino acid residues in a region shown to affect cation requirements and ligand specificity. In this study, we examine the functional significance of the protein isoforms of if by analyzing the ability of transgenes producing only one isoform to rescue developmental abnormalities associated with complete loss of PS2 integrin. We find that either form of αPS2 is sufficient to rescue if− animals to viability; however, the αPS2(C) form promotes higher survival of the organism. Furthermore, these studies suggest distinct roles for αPS2(C) and αPS2(m8) during development. When expressed in the developing wing, αPS2(m8) is more efficient at rescuing the if− wing blister phenotype than is αPS2(C). Expression of αPS2(C) in the eye produces dominant disruption of photoreceptor organization.

We have also examined the ability of αPS2 and αPS1 to maintain photoreceptor organization in the Drosophila retina. Clonal analysis of sectioned eyes suggests a requirement for αPS1, but not αPS2. However, ectopic expression of if(m8) or if(C) shows that either splice form of αPS2 can functionally replace αPS1 and rescue the mew eye phenotype.
Basic fibroblast growth factor induces notochord formation and the expression of As-T, a Brachyury homolog, during ascidian embryogenesis
Authors: Nakatani Yuki , Yasuo Hitoyoshi , Satoh Noriyuki , Nishida Hiroki
Published: April 2021

The tadpole larva of an ascidian develops 40 notochord cells in the center of its tail. Most of the notochord cells originate from the A-line precursors, among which inductive interactions are required for the subsequent differentiation of notochord. The presumptive-endoderm blastomeres or presumptive-notochord blastomeres themselves are inducers of notochord formation. Notochord induction takes place during the 32-cell stage. In amphibia, mesoderm induction is thought to be mediated by several growth factors, for example, activins and basic fibroblast growth factor (bFGF). In the ascidian, Halocynthia roretzi, treatment with bFGF of presumptive-notochord blastomeres that had been isolated at the early 32-cell stage promoted the formation of notochord at a low concentration of bFGF (0.02 ng/ml), while activin failed to induce notochord differentiation. The effect of bFGF reached a maximum at the end of the 32-cell stage and rapidly faded at the beginning of the subsequent cleavage, the time for full induction of notochord being at least 20 minutes.

The expression of As-T, a previously isolated ascidian homolog of the mouse Brachyury (T) gene, starts at the 64cell stage and is detectable exclusively in the presumptivenotochord blastomeres. The present study showed that presumptive-notochord blastomeres, isolated at the early 32-cell stage, neither differentiated into notochord nor expressed the As-T gene. However, when the presumptivenotochord blastomeres were coisolated or recombined with inducer blastomeres, transcripts of As-T were detected. When presumptive-notochord blastomeres were treated with bFGF, the expression of the As-T gene was also detected. These results suggest that inductive interaction is required for the expression of the As-T gene and that the expression of the As-T gene is closely correlated with the determined state of the notochord-precursor cells.
Modulation of Xenopus embryo mesoderm-specific gene expression and dorsoanterior patterning by receptors that activate the phosphatidylinositol cycle signal transduction pathway
Authors: Ault Katherine T. , Durmowicz Gerard , Galione Antony , Harger Patricia L. , Busa William B.
Published: April 2021

A role for the phosphatidylinositol (PI) cycle signal transduction pathway in Xenopus mesoderm induction has been revealed by observations of PI cycle activation coincident with this process, combined with the demonstration that Li+ (a PI cycle inhibitor) blocks this response and hyperdorsalizes mesoderm induction in intact embryos or augments growth factor-mediated induction in animal caps. It has been suggested that spatially restricted PI cycle activity in the marginal zone might modulate (but not, itself, activate) mesoderm induction. To better characterize the ability of PI cycle activity to modulate the pattern of mesoderm-specific gene expression elicited by mesoderm-inducing growth factors we have expressed in the embryo exogenous 5-hydroxytryptamine receptors that activate the PI cycle. In embryos, ventral expression and activation of these receptors during mesoderm induction are without obvious effect, whereas dorsal expression and activation yield dorsoanterior-deficient tadpoles. In animal caps induced with activin, simultaneous activation of exogenous 5-hydroxytryptamine receptors inhibits both convergent extension movements associated with dorsal mesoderm induction and the expression of goosecoid, a dorsal-specific gene, but is without effect on expression of a generic mesodermal marker, Xbra. All of these effects of a PI cycle-stimulating receptor are the opposites of those previously reported for the PI cycle inhibitor, Li+. PI cycle activity thus proves able to modulate the dorsal/ventral character of early mesodermal gene expression elicited by growth factor, suggesting a model for mesodermal patterning.
A novel regulatory mutation in the C. elegans sex determination gene tra-2 defines a candidate ligand/receptor interaction site
Authors: Kuwabara Patricia E.
Published: April 2021

Sex determination in the nematode C. elegans is dependent on cell-to-cell communication, which appears to be mediated by the predicted membrane protein TRA-2A and the secreted protein HER-1. In XO males, HER-1 is hypothesised to function as a repressive ligand that inactivates the TRA-2A receptor. In XX animals, HER-1 is absent and TRA-2A promotes hermaphrodite development by negatively regulating the FEM proteins. This paper describes the molecular and genetic characterisation of a novel class of feminising mutations called tra-2(eg), for enhanced gain-of-function. In XX animals, mutant tra2(eg) activity promotes entirely normal hermaphrodite development. However, the tra-2(eg) mutations generate an XO-specific gain-of-function phenotype, because they transform XO mutants from male into hermaphrodite.

Therefore, the tra-2(eg) mutations identify a major regulatory site, which may be the TRA-2A/HER-1 interaction site. All ten tra-2(eg) mutations encode identical missense changes in a predicted extracellular domain of TRA-2A, named the EG site. It is proposed that the tra-2(eg) mutation encodes a TRA-2A protein that functions constitutively in XO animals, because it is defective in HER-1 binding. Phenotypic characterisation of sexually transformed XO tra-2(eg) hermaphrodites reveals that their fertility is strongly affected by dosage compensation mutations, suggesting that dosage compensation plays a role in normal gametogenesis.
The vegetal determinants required for the Spemann organizer move equatorially during the first cell cycle
Authors: Sakai Masao
Published: April 2021

Embryos with no dorsal axis were obtained when more than 15% of the egg surface was deleted from the vegetal pole of the early 1-cell embryo of Xenopus laevis. The timing of the deletion in the first cell cycle was critical: dorsal-deficient embryos were obtained when the deletion began before time 0.5 (50% of the first cell cycle) whereas normal dorsal axis usually formed when the deletion was done later than time 0.8. The axis deficiency could be restored by lithium treatment and the injection of vegetal but not animal cytoplasm. Bisection of the embryo at the 2-cell stage, which is known to restore the dorsal structures in the UV-ventralized embryos, had no effect on the vegetal-deleted embryos. These results show clearly that, in Xenopus, (1) the dorsal determinants (DDs) localized in the vegetal pole region at the onset of development are necessary for dorsal axis development and (2) the DDs move from the vegetal pole to a subequatorial region where they are incorporated into gastrulating cells to form the future organizing center. A model for the early axis formation process in Xenopus is proposed.
Long-term in vitro culture and characterisation of avian embryonic stem cells with multiple morphogenetic potentialities
Authors: Pain B. , Clark M. E. , Shen M. , Nakazawa H. , Sakurai M. , Samarut J. , Etches R. J.
Published: April 2021

Petitte, J. N., Clarck, M. E., Verrinder Gibbins, A. M. and R. J. Etches (1990; Development 108, 185-189) demonstrated that chicken early blastoderm contains cells able to contribute to both somatic and germinal tissue when injected into a recipient embryo. However, these cells were neither identified nor maintained in vitro. Here, we show that chicken early blastoderm contains cells characterised as putative avian embryonic stem (ES) cells that can be maintained in vitro for long-term culture. These cells exhibit features similar to those of murine ES cells such as typical morphology, strong reactivity toward specific anti bodies, cytokine-dependent extended proliferation and high telomerase activity. These cells also present high capacities to differentiate in vitro into various cell types including cells from ectodermic, mesodermic and endodermic lineages. Production of chimeras after injection of the cultivated cells reinforced the view that our culture system maintains in vitro some avian putative ES cells.
The germ line regulates somatic cyst cell proliferation and fate during Drosophila spermatogenesis
Authors: Gönczy Pierre , DiNardo Stephen
Published: April 2021

Spermatogenesis relies on the function of germ-line stem cells, as a continuous supply of differentiated spermatids is produced throughout life. In Drosophila, there must also be somatic stem cells that produce the cyst cells that accompany germ cells throughout spermatogenesis. By lineage tracing, we demonstrate the existence of such somatic stem cells and confirm that of germ-line stem cells. The somatic stem cells likely correspond to the ultrastructurally described cyst progenitor cells. The stem cells for both the germ-line and cyst lineage are anchored around the hub of non-dividing somatic cells located at the testis tip. We then address whether germ cells regulate the behavior of somatic hub cells, cyst progenitors and their daughter cyst cells by analyzing cell proliferation and fate in testes in which the germ line has been genetically ablated. Daughter cyst cells, which normally withdraw from the cell cycle, continue to proliferate in the absence of germ cells. In addition, cells from the cyst lineage switch to the hub cell fate. Male-sterile alleles of chickadee and diaphanous, which are deficient in germ cells, exhibit similar cyst cell phenotypes. We conclude that signaling from germ cells regulates the proliferation and fate of cells in the somatic cyst lineage.
Oligodendrocytes and their precursors require phosphatidylinositol 3-kinase signaling for survival
Authors: Vemuri Geeta S. , McMorris F. Arthur
Published: April 2021

Signal transduction in response to several growth factors that regulate oligodendrocyte development and survival involves the activation of phosphatidylinositol 3-kinase, which we detect in oligodendrocytes and their precursors. To investigate the role of this enzyme activity, we analyzed cell survival in cultures of oligodendrocytes treated with wortmannin or LY294002, two potent inhibitors of phosphatidylinositol 3-kinase. Cell survival was inhibited by 6070% in these cultures within 24 hours, as quantitated by a tetrazolium staining assay for viable cells and by measurement of DNA content. Similar results were obtained with oligodendrocyte precursor cells. Nuclei of the dying cells contained fragmented DNA, as revealed by terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end-labeling assays, indicating that the cells were dying by apoptosis. Moreover, a significant increase in the number of cells with fragmented nuclear DNA was detected as early as 4 hours, well before any significant differences could be detected in glucose transport or cell viability. Exogenous addition of insulin-like growth factor-I, neurotrophin-3, platelet-derived growth factor, basic fibroblast growth factor, ciliary neurotrophic factor, N-acetyl cysteine, vitamin C, vitamin E, progesterone or serum did not prevent cell death in the presence of wortmannin or LY294002. These findings indicate that survival of oligodendrocytes and their precursors depends on a phosphatidylinositol 3-kinase mediated signaling pathway. Inhibition of this critical enzyme activity induces apoptotic cell death, even in the presence of exogenous growth factors or serum.

Abbreviations used in this paper: bFGF, basic fibroblast growth factor; CNP, 2′,3′-cyclic nucleotide 3′-phosphohydrolase; CNTF, ciliary neurotrophic factor; DMSO, dimethyl sulfoxide; EGF, epidermal growth factor; EGTA, ethylene glycol-bis(β-aminoethyl ether) N,N,N′,N′-tetraacetic acid; GC, galactocerebroside; HEPES, N-2Hydroxyethylpiperazine-N′-2-ethanesulphonic acid; IGF-I and II, insulin-like growth factor-I and II; IRS-1 and 2, insulin receptor substrate 1 and 2; MAP kinase, mitogen activated protein kinase; MTT, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide; NAC, N-acetyl cysteine; NT-3, neurotrophin-3; OM-5 and -6, oligodendroglial cell medium-5 and -6; PBS, phosphate buffered saline; PDGF, platelet-derived growth factor; PI(3)P, phosphatidylinositol(3)phosphate; PI(3,4)P2, phosphatidylinositol(3,4)bisphosphate; PI(3,4,5)P3, phosphatidylinositol(3,4,5)trisphosphate; PI(4)P, phosphatidylinositol(4)phosphate; PI, phosphatidylinositol; PI3-K, phosphatidylinositol 3-kinase; TCA, trichloroacetic acid; TLC, thin layer chromatography; TNF-α, tumor necrosis factor-α; TUNEL, terminal deoxynucleotidyl transferase (TdT)-mediated dUTP-biotin nick end-labeling assay.
A spinal cord fate map in the avian embryo: while regressing, Hensen’s node lays down the notochord and floor plate thus joining the spinal cord lateral walls
Authors: Catala Martin , Teillet Marie-Aimée , Robertis Edward M. De , Douarin Nicole M. Le
Published: April 2021

The spinal cord of thoracic, lumbar and caudal levels is derived from a region designated as the sinus rhomboidalis in the 6-somite-stage embryo. Using quail/chick grafts performed in ovo, we show the following. (1) The floor plate and notochord derive from a common population of cells, located in Hensen’s node, which is equivalent to the chordoneural hinge (CNH) as it was defined at the tail bud stage. (2) The lateral walls and the roof of the neural tube originate caudally and laterally to Hensen’s node, during the regression of which the basal plate anlage is bisected by floor plate tissue. (3) Primary and secondary neurulations involve similar morphogenetic movements but, in contrast to primary neurulation, extensive bilateral cell mixing is observed on the dorsal side of the region of secondary neurulation. (4) The posterior midline of the sinus rhomboidalis gives rise to somitic mesoderm and not to spinal cord. Moreover, mesodermal progenitors are spatially arranged along the rest of the primitive streak, more caudal cells giving rise to more lateral embryonic structures. Together with the results reported in our study of tail bud development (Catala, M., Teillet, M.-A. and Le Douarin, N.M. (1995). Mech. Dev. 51, 51-65), these results show that the mechanisms that preside at axial elongation from the 6-somite stage onwards are fundamentally similar during the complete process of neurulation.
SHORT INTEGUMENT (SIN1), a gene required for ovule development in Arabidopsis, also controls flowering time
Authors: Ray Animesh , Lang Jean D. , Golden Teresa , Ray Sumita
Published: April 2021

The short integument (sin1) mutation causes a femalespecific infertility, and a defect in the control of time to flowering in Arabidopsis. Female sterility of Sin− plants is due to abnormal ovule integument development and aberrant differentiation of the megagametophyte in a subset of ovules. An additional defect of sin1 mutants is the production of an increased number of vegetative leaf and inflorescence primordia leading to delayed flowering. The delayed flowering phenotype of sin1-1 is not due to a defect in the perception of day length periodicity or in gibberellic acid metabolism. Phenotypes of double mutant combina- tions of sin1 with terminal flower (tfl1) indicate that SIN1 activity is required for precocious floral induction typical in a tfl1 mutant. Unexpectedly, sin1-1 tfl1-1 plants do not make pollen, thus revealing a novel role for TFL1 in the anther. Early flowers of sin1-1 ap1-1 double mutants are transformed to long inflorescence-like shoots. A genetic model for the role of SIN1 in flowering time control is proposed.
Mutations affecting the development of the embryonic zebrafish brain
Authors: Schier Alexander F. , Neuhauss Stephan C. F. , Harvey Michele , Malicki Jarema , Solnica-Krezel Lilianna , Stainier Didier Y. R. , Zwartkruis Fried , Abdelilah Salim , Stemple Derek L. , Rangini Zehava , Yang Hong , Driever Wolfgang
Published: April 2021

In a large scale mutagenesis screen for embryonic mutants in zebrafish, we have identified 63 mutations in 24 loci affecting the morphogenesis of the zebrafish brain. The expression of marker genes and the integrity of the axonal scaffold have been studied to investigate abnormalities in regionalization, neurogenesis and axonogenesis in the brain. Mutants can be broadly classified into two groups, one affecting regionalization along the anterior-posterior or dorsal-ventral axis, and the other affecting general features of brain morphology. The first group includes one locus that is required to generate the anlage of the midbrain-hindbrain boundary region at the beginning of somitogenesis. Four loci were identified that affect dorsalventral patterning of the brain, including the previously described cyclops locus. Mutant embryos of this class show a reduction of ventral neuroectodermal structures and variable fusion of the eyes. The second group includes a large class of mutations affecting the formation of brain ventricles. Analysis of this class reveals the requirement of a functional cardiovascular system for ventricle enlargement during embryogenesis. Mutations in one locus lead to the formation of supernumerary primary neurons, a phenotype reminiscent of neurogenic mutants in Drosophila. Other mutant phenotypes described here range from abnormalities in the fasciculation and outgrowth of axons to defects in the diameter of the neural tube. The identified loci establish the genetic foundation for a further analysis of the development of the zebrafish embryonic brain.