bluuebunny/crossref_metadata_2025 huggingface dataset:
Glued participates in distinct microtubule-based activities in Drosophila eye development
Authors: Fan Seng-Sheen , Ready Donald F.
Published: April 2021

A C-terminal truncation of Glued, the Drosophila homolog of the cytoplasmic dynein activating protein, dynactin, results in a severe and complex retinal phenotype, including a roughening of the facet array, malformation of the photosensitive rhabdomeres, and a general deficit and disorder of retinal cells. We have characterized the developmental phenotype in Glued1 and found defects in multiple stages of eye development, including mitosis, nuclear migration, cell fate determination, rhabdomere morphogenesis and cell death. Transgenic flies that express dominant negative Glued under heat-shock control reproduce distinct features of the original Glued1 phenotype depending on the stage of development. The multiple phenotypes effected by truncated Glued point to the multiple roles served by dynactin/dynein during eye development.
Differential response of fetal and adult fibroblasts to cytokines: cell migration and hyaluronan synthesis
Authors: Ellis I. , Banyard J. , Schor S. L.
Published: April 2021

Previous studies have indicated that fetal skin fibroblasts display an elevated level of migratory activity compared to adult cells and that this may result from inherent differences in the production of hyaluronan (HA) by these cells. Data presented in this communication indicate that the elevated level of fetal fibroblast migration into 3D-collagen gels and HA synthesis by these cells were not affected by epidermal growth factor (EGF), platelet-derived growth factor (PDGF), acidic fibroblast growth factor (aFGF) or basic fibroblast growth factor (bFGF). In contrast, both cell migration and HA synthesis by fetal fibroblasts were inhibited by transforming growth factor-beta1 (TGF-β1). Adult fibroblasts responded to these cytokines in a distinct fashion: i.e. cell migration and HA synthesis were stimulated by EGF, PDGF, aFGF and bFGF, but remained unaffected by TGF-β1. Gel-filtration chromatography revealed that these effects of cytokines on HA synthesis were predominantly confined to the production of high molecular mass (>106 kDa) species. Co-exposure of cells to both cytokines and Streptomyces hyaluronidase revealed that (1) the elevated migration of control fetal fibroblasts was inhibited by hyaluronidase, (2) this inhibition was partially restored by co-exposure to EGF, PDGF, aFGF and bFGF, but remained unaffected by TGF-β1, (3) the migration of control adult fibroblasts was unaffected by hyaluronidase and partially stimulated by EGF, aFGF and bFGF (when compared to the effects of these cytokines on cells cultured in the absence of hyaluronidase) and (4) neither PDGF nor TGF-β1 affected the migration of hyaluronidase-treated adult cells. Linear regression analysis revealed a significant correlation between cell migration and HA synthesis by both fetal and adult fibroblasts in the presence and absence of cytokines (r2=0.9277, P<0.0001), with the exception of adult fibroblasts exposed to PDGF. Taken together, these findings suggest that (1) the migration of fetal and adult fibroblasts is differentially modulated by exogenous cytokines and (2) with the possible exception of the effects of PDGF on adult fibroblasts, cytokine-induced modulation of cell migration appears to utilise both HA-dependent and HA-independent pathways.
Defects of urogenital development in mice lacking Emx2
Authors: Miyamoto Norimasa , Yoshida Michio , Kuratani Shigeru , Matsuo Isao , Aizawa Shinichi
Published: April 2021

The homeobox gene Emx2 is a mouse homologue of a Drosophila head gap gene empty spiracles (ems) and is essential for the development of dorsal telencephalon (Yoshida, M., Suda, Y., Matsuo, I., Miyamoto, N., Takeda, N., Kuratani, S. and Aizawa, S. (1997)Development 124, 101-111). At the same time, Emx2 is expressed in the epithelial components of the developing urogenital system and, in Emx2 mutant mice, the kidneys, ureters, gonads and genital tracts were completely missing. Pax-2 and c-ret expressions in the Wolffian duct and WT-1 and GDNF expressions in the metanephric blastema were initially normal in the mutant. The ureteric bud grew and invaded the metanephric mesenchyme where Pax-2 expression was normally induced. Subsequently, however, Pax-2, c-ret and Lim1 expressions in the ureteric bud and GDNF expression in the mesenchyme were greatly reduced. Wnt-4 expression was never found in the mesenchyme. The tip of the ureteric bud never dilated and branching of the bud did not occur.

Neither pretubular cell aggregates nor epithelialization were found in the mesenchyme. Instead the ureteric bud soon degenerated and apoptotic figures were prominent in mesenchymal cells. In explant culture, the mutant ureteric bud did not induce the epithelial transformation of the wild-type mesenchyme, and branching of the mutant ureteric bud was not induced by wild-type mesenchyme. In contrast, defects were not apparent in the mutant mesenchyme by co-culture with wild-type ureteric bud or spinal cord. These results suggest that, in metanephrogenesis, Emx2 is essential for the ureteric bud functions sub-sequent to Pax-2 induction in the metanephric mesenchyme. Degeneration of the Wolffian duct and mesonephric tubules was also abnormally accelerated without the formation of the Müllerian duct.
The single MyoD family gene of Ciona intestinalis encodes two differentially expressed proteins: implications for the evolution of chordate muscle gene regulation
Authors: Meedel Thomas H. , Farmer Steven C. , Lee James J.
Published: April 2021

A MyoD family gene was identified in the ascidian Ciona intestinalis and designated CiMDF (Ciona intestinalisMuscle Determination Factor). Expression of CiMDF was restricted to the muscle cells of the developing embryo and the body-wall muscle of adults. Northern blots showed that two differentially regulated CiMDF transcripts were expressed during development. A 1.8 kb transcript (CiMDFa) appeared first and was gradually replaced by a 2.7 kb transcript (CiMDFb). These transcripts encoded essentially identical MyoD family proteins with the exception of a 68 amino acid C-terminal sequence present in CiMDFb that was absent from CiMDFa. Although both CiMDFa and CiMDFb contained the cysteine-rich/basichelix loop helix domain (Cys-rich/bHLH) present in all MyoD family proteins, only CiMDFb contained the region near the C terminus (Domain III) characteristic of this gene family. Genomic Southern blots showed that C. intestinalis has only one MyoD family gene, suggesting that CiMDFa and CiMDFb result from differential processing of primary transcripts. The existence of two MyoD family proteins that are differentially expressed during ascidian embryogenesis has novel parallels to vertebrate muscle development and may reflect conserved myogenic regulatory mechanisms among chordates.
A nanos homolog in leech
Authors: Pilon Marc , Weisblat David A.
Published: April 2021

From the glossiphoniid leech Helobdella robusta, we have cloned and determined the complete coding sequence of Hro-nos, a gene homologous to the nanos gene from Drosophila melanogaster. Developmental northern blots show that Hro-nos, like nanos, is a maternal transcript that decays rapidly during early development. A polyclonal antiserum raised against the HRO-NOS protein was used in developmental western blots and for immunostaining leech embryos of different developmental stages. The HRO-NOS protein is first detectable in 2-cell embryos (4-6 hours of development) and exhibits a transient expression peaking during fourth cleavage (9-12 cells; 8-14 hours of development). The HRO-NOS protein exhibits a graded distribution along the primary embryonic axis and is partitioned unequally between the sister cells DNOPQ and DM, progeny of macromere D′ at fourth cleavage: DNOPQ is the segmental ectoderm precursor cell and exhibits levels of HRO-NOS protein that are at least two-fold higher than in cell DM, the segmental mesoderm precursor cell. The observed expression pattern suggests that Hro-nos plays a role in the decision between ectodermal and mesodermal cell fates in leech.
The nuclear receptor SF-1 mediates sexually dimorphic expression of Müllerian Inhibiting Substance, in vivo
Authors: Giuili Galicia , Shen Wen-Hui , Ingraham Holly A.
Published: April 2021

Müllerian Inhibiting Substance (MIS) functions to promote regression of the Müllerian duct during male development. Maintaining the sexually dimorphic pattern of MIS expression is essential for proper mammalian reproductive tract development. Here, we show that the intricate spatial and temporal pattern of MIS expression is directed by a remarkably small proximal promoter of only 180 base pairs in length. Expression of the MIS-human growth hormone transgene (MIS/GH) is restricted to Sertoli cells in embryonic testis and to granulosa cells of postnatal ovary, consistent with the known MIS expression pattern. The proximal MIS promoter is therefore sufficient to direct the initiation and the maintenance of MIS gene expression in both sexes. Moreover, in vivo MIS promoter activity requires an intact binding site for the orphan nuclear receptor SF-1. Taken together, these data strongly suggest that SF-1 directly activates MIS in embryonic and postnatal gonads. Consistent with the proposed role of SF-1 in mammalian sex-determination, our study provides physiological evidence that a SF-1 binding site is essential for gene activation of an embryonic testis-specific marker.
A role for the Drosophila Toll/Cactus pathway in larval hematopoiesis
Authors: Qiu Ping , Pan Pang Chu , Govind Shubha
Published: April 2021

In the Drosophila larva, blood cells or hemocytes are formed in the lymph gland. The major blood cell type, called plasmatocyte, is small, non-adhesive and phagocytic. Plasmatocytes differentiate into adhesive lamellocytes to form multilayered capsules around foreign substances or, in mutant melanotic tumor strains, around self tissue. Mutations in cactus or Toll, or constitutive expression of dorsal can induce lamellocyte differentiation and cause the formation of melanotic capsules. As maternally encoded proteins, Toll, Cactus and Dorsal, along with Tube and Pelle, participate in a common signal transduction pathway to specify the embryonic dorsal-ventral axis. Using the maternal pathway as a paradigm, we investigated if these proteins have additional roles in larval hemocyte formation and differentiation. Analysis of cactus mutants that lack Cactus protein revealed that almost all of these animals have an overabundance of hemocytes, carry melanotic capsules and die before reaching pupal stages. In addition, the lymph glands of cactus larvae are considerably enlarged. The number of mitotic cells in the cactus and TollD hemolymph is higher than that in the wild-type hemolymph. The hemocyte density of mutant Toll, tube or pelle hemolymph is significantly lower than that of the wild type. Lethality of mutant cactus animals could be rescued either by the selective expression of wild-type Cactus protein in the larval lymph gland or by the introduction of mutations in Toll, tube or pelle. Cactus, Toll, Tube and Pelle proteins are expressed in the nascent hemocytes of the larval lymph gland. Our results suggest that the Toll/Cactus signal transduction pathway plays a significant role in regulating hemocyte proliferation and hemocyte density in the Drosophila larva. These findings are discussed in light of similar hematopoietic functions of Rel/IĸB-family proteins in mice.
Patterns and control of cell motility in the Xenopus gastrula
Authors: Wacker Stephan , Brodbeck Anja , Lemaire Patrick , Niehrs Christof , Winklbauer Rudolf
Published: April 2021

By comparing cells with respect to several motility-related properties and the ability to migrate on fibronectin, three cell types can be distinguished in the Xenopus gastrula. These occur in a distinct spatial pattern, thus defining three motility domains which do not correspond to the prospective germ layers. Migratory behavior is confined to a region encompassing the anterior mesoderm and endoderm. When stationary animal cap cells are induced to migrate by treatment with activin, cells become adhesive at low concentrations of fibronectin, show polarized protrusive activity, and form lamellipodia. Adhesion and polarization, but not lamellipodia formation, are mimicked by the immediate early response gene Mix.1. Goosecoid, another immediate early gene, is without effect when expressed alone in animal cap cells, but it acts synergistically with Mix.1 in the control of adhesion, and antagonistically in the polarization of protrusive activity. bFGF also induces migration, lamellipodia formation and polarization in animal cap cells, but has no effect on adhesion. By the various treatments of animal cap cells, new combinations of motile properties can be generated, yielding cell types which are not found in the embryo.
The RXRα gene functions in a non-cell-autonomous manner during mouse cardiac morphogenesis
Authors: Tran Chanh M. , Sucov Henry M.
Published: April 2021

Germline mutation in mice of the retinoic acid receptor gene RXRα results in a proliferative failure of cardiomyocytes, which leads to an underdeveloped ventricular chamber and midgestation lethality. Mutation of the cell cycle regulator N-myc gene also leads to an apparently identical phenotype. In this study, we demonstrate by chimera analysis that the cardiomyocyte phenotype in RXRα−/−embryos is a non-cell-autonomous phenotype. In chimeric embryos made with embryonic stem cells lacking RXRα, cardiomyocytes deficient in RXRα develop normally and contribute to the ventricular chamber wall in a normal manner. Because the ventricular hypoplastic phenotype reemerges in highly chimeric embryos, we conclude that RXRα functions in a non-myocyte lineage of the heart to induce cardiomyocyte proliferation and accumulation, in a manner that is quantitatively sensitive. We further show that RXRα is not epistatic to N-myc, and that RXRα and N-myc regulate convergent obligate pathways of cardiomyocyte maturation.
A role for SOX1 in neural determination
Authors: Pevny Larysa H. , Sockanathan Shantini , Placzek Marysia , Lovell-Badge Robin
Published: April 2021

In vertebrates, the delineation of the neural plate from a region of the primitive ectoderm is accompanied by the onset of specific gene expression which in turn promotes the formation of the nervous system. Here we show that SOX1, an HMG-box protein related to SRY, is one of the earliest transcription factors to be expressed in ectodermal cells committed to the neural fate: the onset of expression of SOX1 appears to coincide with the induction of neural ectoderm. We demonstrate a role for SOX1 in neural determination and differentiation using an inducible expression P19 cell system as an in vitro model of neurogenesis. Misexpression of SOX1 can substitute for the requirement of retinoic acid to impart neural fate to competent ectodermal P19 cells. Using a series of antigenic markers which identify early neural cell types in combination with BrdU labeling, we demonstrate a temporal and spatial correlation between the differentiation of cell types along the dorsoventral axis of the neural tube and the downregulation of SOX1 expression. SOX1, therefore, defines the dividing neural precursors of the embryonic central nervous system (CNS).
Determination of sclerotome to the cartilage fate
Authors: Dockter Jennifer L. , Ordahl Charles P.
Published: April 2021

When the somite first forms the cells appear to be equivalent in potential. In order to understand the lineage diversification of the somite, the determination of sclerotome cells to the cartilage fate was tested using an in vivo challenge assay in which quail sclerotome fragments were grafted into a dorsal position in a chick host. Grafts containing undetermined cells were expected to differentiate into other tissues while grafts containing determined chondrocyte precursors were expected to consistently give rise to cartilage. We found that grafted sclerotome fragments from somite stages V-XX were capable of giving rise to integrated muscle and dermis and that it was not until fragments from stage XII somites were grafted that cartilage was consistently produced in the assay. Sclerotomal tissue from embryonic day 4-6 embryos remained as morphologically unintegrated mesenchyme when grafted into an embryonic day 2 host, but formed only cartilage when placed into an identically aged host. Vertebral body cartilage from embryonic day 7 and embryonic day 8 embryos formed exclusively ectopic cartilage in an embryonic day 2 host. We conclude that cells determined to the cartilage fate do not appear until somite stage XII, but that not all sclerotome cells are determined at this time. The effect of host age on the differentiation and morphogenetic behavior of sclerotome fragment grafts in this assay indicate the existence of developmental eras within the embryo.
Drosophila βHeavy-spectrin is essential for development and contributes to specific cell fates in the eye
Authors: Thomas Claire M. , Zarnescu Daniela C. , Juedes Amy E. , Bales Mark A. , Londergan Amy , Korte Carol C. , Kiehart Daniel P.
Published: April 2021

The spectrin membrane skeleton is a ubiquitous cytoskeletal structure with several cellular roles, including the maintenance of cell integrity, determination of cell shape and as a contributor to cell polarity. We have isolated mutations in the gene encoding βHeavy-spectrin in Drosophila, and have named this essential locus karst. karst mutant individuals have a pleiotropic phenotype characterized by extensive larval lethality and, in adult escapers, rough eyes, bent wings, tracheal defects and infertility. Within karst mutant eyes, a significant number of ommatidia specifically lack photoreceptor R7 alongside more complex morphological defects.

Immunolocalization of βHeavy-spectrin in wild-type eye-antennal and wing imaginal discs reveals that βHeavy-spectrin is present in a restricted subdomain of the membrane skeleton that colocalizes with DE-cadherin. We propose a model where normal levels of Sevenless signaling are dependent on tight cell-cell adhesion facilitated by the βHeavy-spectrin membrane skeleton.

Immunolocalization of βHeavy-spectrin in the adult and larval midgut indicates that it is a terminal web protein, but we see no gross morphological defects in the adult apical brush border in karst mutant flies. Rhodamine phalloidin staining of karst mutant ovaries similarly reveals no conspicuous defect in the actin cytoskeleton or cellular morphology in egg chambers. This is in contrast to mutations in α-spectrin, the molecular partner of βHeavy-spectrin, which affect cellular structure in both the larval gut and adult ovaries.

Our results emphasize the fundamental contribution of the spectrin membrane skeleton to normal development and reveals a critical interplay between the integrity of a cell’s membrane skeleton, the structure of cell-cell contacts and cell signaling.
Combinatorial Gli gene function in floor plate and neuronal inductions by Sonic hedgehog
Authors: Altaba A. Ruiz i
Published: April 2021

Within the developing vertebrate nervous system, it is not known how progenitor cells interpret the positional information provided by inducing signals or how the domains in which distinct groups of neural cells differentiate are defined. Gli proteins may be involved in these processes. In the frog neural plate, we have previously shown that the zinc finger transcription factor Gli1 is expressed in midline cells and mediates the effects of Shh inducing floor plate differentiation. In contrast, Gli2 and Gli3 are expressed throughout the neural plate except for the midline. Here, it is shown that Gli3 and Shh repress each other whereas Gli2, like Gli1, is a target of Shh signaling. However, only Gli1 can induce the differentiation of floor plate cells. In addition, Gli2 and Gli3 repress the ectopic induction of floor plate cells by Gli1 in co-injection assays and inhibit endogenous floor plate differentiation. The definition of the floor plate domain, therefore, appears to be defined by the antagonizing activities of Gli2 and Gli3 on Gli1 function. Because both Gli1 and Gli2 are induced by Shh, these results establish a regulatory feedback loop triggered by Shh that restricts floor plate cells to the midline. We have also previously shown that the Gli genes induce neuronal differentiation and here it is shown that there is specificity to the types of neurons the Gli proteins induce. Only Gli1 induces Nkx2.1/TTF-1+ ventral forebrain neurons. Moreover, Gli2 and Gli3 inhibit their differentiation. In contrast, the differentiation of spinal motor neurons can be induced by the two ventrally expressed Gli genes, Gli1 and Gli2, suggesting that Gli2 directly mediates induction of motor neurons by Shh. In addition, Gli3 inhibits motor neuron differentiation by Gli2. Thus, combinatorial Gli function may pattern the neural tube, integrating positional information and cell type differentiation.
Mice lacking the homeodomain transcription factor Nkx2.2 have diabetes due to arrested differentiation of pancreatic β cells
Authors: Sussel L. , Kalamaras J. , Hartigan-O’Connor D. J. , Meneses J. J. , Pedersen R. A. , Rubenstein J. L. R. , German M. S.
Published: April 2021

The endocrine pancreas is organized into clusters of cells called islets of Langerhans comprising four well-defined cell types: α, β, δ and PP cells. While recent genetic studies indicate that islet development depends on the function of an integrated network of transcription factors, the specific roles of these factors in early cell-type specification and differentiation remain elusive. Nkx2.2 is a member of the mammalian NK2 homeobox transcription factor family that is expressed in the ventral CNS and the pancreas. Within the pancreas, we demonstrate that Nkx2.2 is expressed in α, β and PP cells, but not in δ cells. In addition, we show that mice homozygous for a null mutation of Nkx2.2 develop severe hyperglycemia and die shortly after birth.

Immunohistochemical analysis reveals that the mutant embryos lack insulin-producing β cells and have fewer glucagon-producing α cells and PP cells. Remarkably, in the mutants there remains a large population of islet cells that do not produce any of the four endocrine hormones. These cells express some β cell markers, such as islet amyloid polypeptide and Pdx1, but lack other definitive β cell markers including glucose transporter 2 and Nkx6.1. We propose that Nkx2.2 is required for the final differentiation of pancreatic β cells, and in its absence, β cells are trapped in an incompletely differentiated state.
The hem of the embryonic cerebral cortex is defined by the expression of multiple Wnt genes and is compromised in Gli3-deficient mice
Authors: Grove Elizabeth A. , Tole Shubha , Limon Julie , Yip Lin-wah , Ragsdale Clifton W.
Published: April 2021

In the developing vertebrate CNS, members of the Wnt gene family are characteristically expressed at signaling centers that pattern adjacent parts of the neural tube. To identify candidate signaling centers in the telencephalon, we isolated Wnt gene fragments from cDNA derived from embryonic mouse telencephalon. In situ hybridization experiments demonstrate that one of the isolated Wnt genes, Wnt7a, is broadly expressed in the embryonic telencephalon. By contrast, three others, Wnt3a, 5a and a novel mouse Wnt gene, Wnt2b, are expressed only at the medial edge of the telencephalon, defining the hem of the cerebral cortex.

The Wnt-rich cortical hem is a transient, neuron-containing, neuroepithelial structure that forms a boundary between the hippocampus and the telencephalic choroid plexus epithelium (CPe) throughout their embryonic development. Indicating a close developmental relationship between the cortical hem and the CPe, Wnt gene expression is upregulated in the cortical hem both before and just as the CPe begins to form, and persists until birth. In addition, although the cortical hem does not show features of differentiated CPe, such as expression of transthyretin mRNA, the CPe and cortical hem are linked by shared expression of members of the Bmp and Msx gene families.

In the extra-toesJ(XtJ) mouse mutant, telencephalic CPe fails to develop. We show that Wnt gene expression is deficient at the cortical hem in XtJ/XtJmice, but that the expression of other telencephalic developmental control genes, including Wnt7a, is maintained. The XtJmutant carries a deletion in Gli3, a vertebrate homolog of the Drosophila gene cubitus interruptus (ci), which encodes a transcriptional regulator of the Drosophila Wnt gene, wingless. Our observations indicate that Gli3 participates in Wnt gene regulation in the vertebrate telencephalon, and suggest that the loss of telencephalic choroid plexus in XtJmice is due to defects in the cortical hem that include Wnt gene misregulation.
Control of floral homeotic gene expression and organ morphogenesis in Antirrhinum
Authors: McSteen Paula C. M. , Vincent Coral A. , Doyle Sandra , Carpenter Rosemary , Coen Enrico S.
Published: April 2021

The development of reproductive organs in Antirrhinum depends on the expression of an organ identity gene, plena, in the central domain of the floral meristem. To investigate the mechanism by which plena is regulated, we have characterised three mutants in which the pattern of plena expression is altered. In polypetala mutants, expression of plena is greatly reduced, resulting in a proliferation of petals in place of reproductive organs. In addition, polypetala mutants exhibit an altered pattern of floral organ initiation, quite unlike that seen in loss-of-function plena mutants. This suggests that polypetala normally has two roles in flower development: regulation of plena and control of organ primordia formation. In fistulata mutants, plena is ectopically expressed in the distal domain of petal primordia, resulting in the production of anther-like tissue in place of petal lobes. Flowers of fistulata mutants also show a reduced rate of petal lobe growth, even in a plena mutant background. This implies that fistulata normally has two roles in the distal domain of petal primordia: inhibition of plena expression and promotion of lobe growth. A weak allele of the floral meristem identity gene, floricaula, greatly enhances the effect of fistulata on plena expression, showing that floricaula also plays a role in repression of plena in outer whorls. Taken together, these results show that genes involved in plena regulation have additional roles in the formation of organs, perhaps reflecting underlying mechanisms for coupling homeotic gene expression to morphogenesis.
Chromatin silencing and the maintenance of a functional germline in Caenorhabditis elegans
Authors: Kelly William G. , Fire Andrew
Published: April 2021

The germline of the nematode Caenorhabditis elegans exhibits a remarkable ability to specifically silence transgenic DNA. We have shown that this silencing mechanism is disrupted in animals mutant for the maternal effect sterile genes mes-2, mes-3, mes-4 and mes-6. The proteins encoded by mes-2 and mes-6 have been shown to be related to the Polycomb Group of transcriptional repressors (Holdeman, R., Nehrt, S. and Strome, S. (1998). Development 125, 2457-2467; Korf, I., Fan, F. and Strome, S. (1998). Development 125, 2469-2478). These results suggest that a genetic silencing process is essential for sustained germline function, and that this silencing is mediated, at least in part, by Polycomb Group proteins.
GSK3β/shaggy mediates patterning along the animal-vegetal axis of the sea urchin embryo
Authors: Emily-Fenouil Françoise , Ghiglione Christian , Lhomond Guy , Lepage Thierry , Gache Christian
Published: April 2021

In the sea urchin embryo, the animal-vegetal axis is defined before fertilization and different embryonic territories are established along this axis by mechanisms which are largely unknown. Significantly, the boundaries of these territories can be shifted by treatment with various reagents including zinc and lithium. We have isolated and characterized a sea urchin homolog of GSK3β/shaggy, a lithium-sensitive kinase which is a component of the Wnt pathway and known to be involved in axial patterning in other embryos including Xenopus. The effects of overexpressing the normal and mutant forms of GSK3β derived either from sea urchin or Xenopus were analyzed by observation of the morphology of 48 hour embryos (pluteus stage) and by monitoring spatial expression of the hatching enzyme (HE) gene, a very early gene whose expression is restricted to an animal domain with a sharp border roughly coinciding with the future ectoderm / endoderm boundary. Inactive forms of GSK3β predicted to have a dominant-negative activity, vegetalized the embryo and decreased the size of the HE expression domain, apparently by shifting the boundary towards the animal pole. These effects are similar to, but even stronger than, those of lithium. Conversely, overexpression of wild-type GSK3β animalized the embryo and caused the HE domain to enlarge towards the vegetal pole. Unlike zinc treatment, GSK3β overexpression thus appeared to provoke a true animalization, through extension of the presumptive ectoderm territory.

These results indicate that in sea urchin embryos the level of GSKβ activity controls the position of the boundary between the presumptive ectoderm and endoderm territories and thus, the relative extent of these tissue layers in late embryos. GSK3β and probably other downstream components of the Wnt pathway thus mediate patterning both along the primary AV axis of the sea urchin embryo and along the dorsal-ventral axis in Xenopus, suggesting a conserved basis for axial patterning between invertebrate and vertebrate in deuterostomes.
Involvement of Sox1, 2 and 3 in the early and subsequent molecular events of lens induction
Authors: Kamachi Yusuke , Uchikawa Masanori , Collignon Jérôme , Lovell-Badge Robin , Kondoh Hisato
Published: April 2021

Activation of the first lens-specific gene of the chicken, δ1-crystallin, is dependent on a group of lens nuclear factors, δEF2, interacting with the δ1-crystallin minimal enhancer, DC5. One of the δEF2 factors was previously identified as SOX2. We show that two related SOX proteins, SOX1 and SOX3, account for the remaining members of δEF2. Activation of the DC5 enhancer is dependent on their C-terminal domains. Expression of Sox1-3 in the eye region during lens induction was studied in comparison with Pax6 and δ?-crystallin. Pax6, known to be required for the inductive response of the ectoderm, is broadly expressed in the lateral head ectoderm from before lens induction. After tight association of the optic vesicle (around stage 10-11, 40 hours after egg incubation), expression of Sox2 and Sox3 is activated in the vesicle-facing ectoderm at stage 12 (44 hours). These cells, expressing together Pax6 and Sox2/3, subsequently give rise to the lens, beginning with formation of the lens placode and expression of δ-crystallin at stage 13 (48 hours). Sox1 then starts to be expessed in the lens-forming cells at stage 14. When the prospective retina area of the neural plate was unilaterally ablated at stage 7, expression of Sox2/3 was lost in the side of lateral head ectoderm lacking the optic cup, implying that an inductive signal from the optic cup activates Sox2/3 expression. In the mouse embryonic lens, this subfamily of Sox genes is expressed in an analogous fashion, although Sox3 transcripts have not been detected and Sox2 expression is down-regulated when Sox1 is activated. In ectodermal tissues of the chicken embryo, δ-crystallin expression occurs in a few ectopic sites. These are always characterized by overlapping expression of Sox2/3 and Pax6. Thus, an essential molecular event in lens induction is the ‘turning on’ of the transcriptional regulators SOX2/3 in the Pax6-expressing ectoderm and these SOX proteins activate crystallin gene expression. Continued activity, especially of SOX1, is then essential for further development of the lens.
Hoxa-2 restricts the chondrogenic domain and inhibits bone formation during development of the branchial area
Authors: Kanzler Benoît , Kuschert Stephen J. , Liu Yi-Hsin , Mallo Moisés
Published: April 2021

In Hoxa-2−/− embryos, the normal skeletal elements of the second branchial arch are replaced by a duplicated set of first arch elements. We show here that Hoxa-2 directs proper skeletal formation in the second arch by preventing chondrogenesis and intramembranous ossification. In normal embryos, Hoxa-2 is expressed throughout the second arch mesenchyme, but is excluded from the chondrogenic condensations. In the absence of Hoxa-2, chondrogenesis is activated ectopically within the rostral Hoxa-2 expression domain to form the mutant set of cartilages. In Hoxa-2−/− embryos the Sox9 expression domain is shifted into the normal Hoxa-2 domain. Misexpression of Sox9 in this area produces a phenotype resembling that of the Hoxa-2 mutants. These results indicate that Hoxa-2 acts at early stages of the chondrogenic pathway, upstream of Sox9 induction. We also show that Hoxa-2 inhibits dermal bone formation when misexpressed in its precursors. Furthermore, molecular analyses indicate that Cbfa1 is upregulated in the second branchial arches of Hoxa-2 mutant embryos suggesting that prevention of Cbfa1 induction might mediate Hoxa-2 inhibition of dermal bone formation during normal second arch development. The implications of these results on the patterning of the branchial area are discussed.
SCL specifies hematopoietic mesoderm in Xenopus embryos
Authors: Mead Paul E. , Kelley Clair M. , Hahn Peter S. , Piedad Oscar , Zon Leonard I.
Published: April 2021

Targeted gene disruption experiments in the mouse have demonstrated an absolute requirement for several transcription factors for the development of hematopoietic progenitors during embryogenesis. Disruption of the basic helix-loop-helix gene SCL (stem cell leukemia) causes a block early in the hematopoietic program with defects in all hematopoietic lineages. To understand how SCL participates in the organogenesis of blood, we have isolated cDNAs encoding Xenopus SCL and characterized the function of SCL during embryogenesis. We demonstrate that SCL is expressed in ventral mesoderm early in embryogenesis. SCL expression is induced by BMP-4, and a dominant negative BMP-4 receptor inhibits SCL expression in the ventral region of the embryo. Expression of SCL in either bFGF-treated animal pole explants or dorsal marginal zone explants leads to the expression of globin protein. Furthermore, over-expression of SCL does not alter normal dorsal-ventral patterning in the embryo, indicating that SCL acts to specify mesoderm to a hematopoietic fate after inductive and patterning events have occurred. We propose that SCL is both necessary and sufficient to specify hematopoietic mesoderm, and that it has a similar role in specifying hematopoietic cell fate as MyoD has in specifying muscle cell fate.
Two domains in vertebral development: antagonistic regulation by SHH and BMP4 proteins
Authors: Watanabe Yuji , Duprez Delphine , Monsoro-Burq Anne-Hélène , Vincent Christine , Douarin Nicole M. Le
Published: April 2021

It has previously been shown that the notochord grafted laterally to the neural tube enhances the differentiation of the vertebral cartilage at the expense of the derivatives of the dermomyotome. In contrast, the dorsomedial graft of a notochord inhibits cartilage differentiation of the dorsal part of the vertebra carrying the spinous process. Cartilage differentiation is preceded by the expression of transcription factors of the Pax family (Pax1/Pax9) in the ventrolateral domain and of the Msx family in the dorsal domain. The proliferation and differentiation of Msx-expressing cells in the dorsal precartilaginous domain of the vertebra are stimulated by BMP4, which acts upstream of Msx genes. It has previously been shown that the SHH protein arising from the notochord (and floor plate) is necessary for the survival and further development of Pax1/Pax9-expressing sclerotomal cells. We show here that SHH acts antagonistically to BMP4. SHH-producing cells grafted dorsally to the neural tube at E2 inhibit expression of Bmp4 and Msx genes and also inhibits the differentiation of the spinous process. We present a model that accounts for cartilage differentiation in the vertebra.
Ecdysone pathway is required for furrow progression in the developing Drosophila eye
Authors: Brennan Catherine A. , Ashburner Michael , Moses Kevin
Published: April 2021

In Drosophila, secretion of the steroid hormone ecdysone from the prothoracic ring gland coordinates and triggers events such as molting and metamorphosis. In the developing Drosophila compound eye, pattern formation and cell-type specification initiate at a moving boundary known as the morphogenetic furrow. We have investigated the role of ecdysone in eye development and report here that the ecdysone signaling pathway is required for progression of the morphogenetic furrow in the eye imaginal disc of Drosophila. Genetic disruption both of the ecdysone signal in vivo with the ecdysoneless1 (ecd1) mutant and of ecdysone response with a Broad-Complex mutant result in disruption of morphogenetic furrow progression. In addition, we show that ecdysone-dependent gene expression, both of a reporter of transcriptional activity of the Ecdysone Receptor and of the Z1 isoform of the Broad Complex, are localized in and close to the furrow. These results suggest that, in the morphogenetic furrow, temporal hormonal signals are integrated into genetic pathways specifying spatial pattern.
Mutant Vg1 ligands disrupt endoderm and mesoderm formation in Xenopus embryos
Authors: Joseph Elaine M. , Melton Douglas A.
Published: April 2021

The Xenopus Vg1 gene, a TGFβ superfamily member, is expressed as a maternal mRNA localized to prospective endoderm, and mature Vg1 protein can induce both endodermal and mesodermal markers in embryonic cells. Most previous work on embryonic inducers, including activin, BMPs and Vg1, has relied on ectopic expression to assay for gene function. Here we employ a mutant ligand approach to block Vg1 signaling in developing embryos. The results indicate that Vg1 expression is essential for normal endodermal development and the induction of dorsal mesoderm in vivo.
TGF-β/BMP superfamily members, Gbb-60A and Dpp, cooperate to provide pattern information and establish cell identity in the Drosophila wing
Authors: Khalsa Ongkar , Yoon Jung-won , Torres-Schumann Sonia , Wharton Kristi A.
Published: April 2021

Within a developing organism, cells receive many signals which control their proliferation, fate specification and differentiation. One group of such proteins is the TGF-β/BMP class of related signaling molecules. Based on expression studies, multiple members of this class of ligands must impinge upon the same cells of a developing tissue; however, the role that multiple TGF-β/BMP ligands may play in directing the development of such a tissue is not understood. Here we provide evidence that multiple BMPs are required for growth and patterning of the Drosophila wing. The Drosophila BMP gene, gbb-60A, exhibits a requirement in wing morphogenesis distinct from that shown previously for dpp, a well-characterized Drosophila BMP member. gbb-60A mutants exhibit a loss of pattern elements from the wing, particularly those derived from cells in the posterior compartment, consistent with the gbb-60A RNA and protein expression pattern. Based on genetic analysis and expression studies, we conclude that Gbb-60A must signal primarily as a homodimer to provide patterning information in the wing imaginal disc. We demonstrate that gbb-60A and dpp genetically interact and that specific aspects of this interaction are synergistic while others are antagonistic. We propose that the positional information received by a cell at a particular location within the wing imaginal disc depends on the balance of Dpp to Gbb-60A signaling. Furthermore, the critical ratio of Gbb-60A to Dpp signaling appears to be mediated by both Tkv and Sax type I receptors.
Leafbladeless1 is required for dorsoventrality of lateral organs in maize
Authors: Timmermans Marja C. P. , Schultes Neil P. , Jankovsky Julia P. , Nelson Timothy
Published: April 2021

The maize leafbladeless1 (lbl1) mutant displays a variety of leaf and plant phenotypes. The most extreme manifestation in the leaf is the formation of radially symmetric, abaxialized leaves due to a complete loss of adaxial cell types. Less severe phenotypes, resulting from a partial loss of adaxial cell identity, include the formation of ectopic laminae at the boundary between abaxialized, mutant sectors on the adaxial leaf surface and the bifurcation of leaves. Ectopic laminae and bifurcations arise early in leaf development and result in an altered patterning of the leaf along the proximodistal axis, or in complete duplication of the developing organ. Leaf-like lateral organs of the inflorescences and flowers show similar phenotypes. These observations suggest that Lbl1 is required for the specification of adaxial cell identity within leaves and leaflike lateral organs. Lbl1 is also required for the lateral propagation of leaf founder cell recruitment, and plays a direct or indirect role in the downregulation of the homeobox gene, knotted1, during leaf development. Our results suggest that adaxial/abaxial asymmetry of lateral organs is specified in the shoot apical meristem, and that formation of this axis is essential for marginal, lateral growth and for the specification of points of proximodistal growth. Parallels between early patterning events during lateral organ development in plants and animals are discussed.
Transgene expression of steel factor in the basal layer of epidermis promotes survival, proliferation, differentiation and migration of melanocyte precursors
Authors: Kunisada Takahiro , Yoshida Hisahiro , Yamazaki Hidetoshi , Miyamoto Akitomo , Hemmi Hiroaki , Nishimura Emi , Shultz Leonard D. , Nishikawa Shin-Ichi , Hayashi Shin-Ichi
Published: April 2021

Mutations at the murine dominant white spotting (KitW) and steel (MgfSl) loci, encoding c-Kit receptor kinase and its ligand respectively, exert developmental defects on hematopoietic cells, melanocytes, germ cells and interstitial cells of Cajal. The expression patterns of steel factor (SLF) observed in the skin and gonads suggest that SLF mediates a migratory or a chemotactic signal for c-Kit-expressing stem cells (melanocyte precursors and primordial germ cells). By targeting expression of SLF to epidermal keratinocytes in mice, we observed extended distribution of melanocytes in a number of sites including oral epithelium and footpads where neither melanocytes nor their precursors are normally detected. In addition, enlarged pigmented spots of KitW and other spotting mutant mice were observed in the presence of the SLF transgene. These results provide direct evidence that SLF stimulates migration of melanocytes in vivo. We also present data suggesting that SLF does not simply support survival and proliferation of melanocytes but also promotes differentiation of these cells. Unexpectedly, melanocyte stem cells independent of the c-Kit signal were maintained in the skin of the SLF transgenic mice. After the elimination of c-Kit-dependent melanoblasts by function-blocking anti-c-Kit antibody, these stem cells continued to proliferate and differentiate into mature melanocytes. These melanoblasts are able to migrate to cover most of the epidermis after several months. The SLF transgenic mice described in this report will be useful in the study of melanocyte biology.
Localized changes in apoplastic and cytoplasmic pH are associated with root hair development in Arabidopsis thaliana
Authors: Bibikova Tatiana N. , Jacob Tobias , Dahse Ingo , Gilroy Simon
Published: April 2021

Morphogenesis in plants is characterized by highly regulated cell enlargement. However, the mechanisms controlling and localizing regions of growth remain essentially unknown. Root hair formation involves the induction of a localized cell expansion in the lateral wall of a root epidermal cell. This expanded region then enters a second phase of localized growth called tip growth. Root hair formation therefore provides a model in which to study the cellular events involved in regulating localized growth in plants. Confocal ratio imaging of the pH of the cell wall revealed an acidification at the root hair initiation site. This acidification was present from the first morphological indications of localized growth, but not before, and was maintained to the point where the process of root hair initiation ceased and tip growth began. Preventing the wall acidification with pH buffers arrested the initiation process but growth resumed when the wall was returned to an acidic pH. Cytoplasmic pH was found to be elevated from approximately 7.3 to 7.7 at the initiation site, and this elevation coincided with the acidification of the wall. Preventing the localized increase in cytoplasmic pH with 10 mM butyrate however did not inhibit either the wall acidification or the initiation process. In contrast, there was no detectable gradient in pH associated with the apex of tip growing root hairs, but both elevated apoplastic pH and butyrate treatment irreversibly inhibited the tip growth process. Thus the processes of tip growth and initiation of root hairs show differences in their pH requirements. These results highlight the role of localized control of apoplastic pH in the control of cell architecture and morphogenesis in plants.
Characterization of three novel members of the zebrafish Pax2/5/8 family: dependency of Pax5 and Pax8 expression on the Pax2.1 (noi) function
Authors: Pfeffer Peter L. , Gerster Thomas , Lun Klaus , Brand Michael , Busslinger Meinrad
Published: April 2021

The mammalian Pax2, Pax5 and Pax8 genes code for highly related transcription factors, which play important roles in embryonic development and organogenesis. Here we report the characterization of all members of the zebrafish Pax2/5/8 family. These genes have arisen by duplications before or at the onset of vertebrate evolution. Due to an additional genome amplification in the fish lineage, the zebrafish contains two Pax2 genes, the previously known Pax[b] gene (here renamed as Pax2.1) and a novel Pax2.2 gene. The zebrafish Pax2.1 gene most closely resembles the mammalian Pax2 gene in its expression pattern, as it is transcribed first in the midbrain-hindbrain boundary region, then in the optic stalk, otic system, pronephros and nephric ducts, and lastly in specific interneurons of the hindbrain and spinal cord. Pax2.2 differs from Pax2.1 by the absence of expression in the nephric system and by a delayed onset of transcription in other Pax2.1 expession domains. Pax8 is also expressed in the same domains as Pax2.1, but its transcription is already initiated during gastrulation in the primordia of the otic placode and pronephric anlage, thus identifying Pax8 as the earliest developmental marker of these structures. The zebrafish Pax5 gene, in contrast to its mouse orthologue, is transcribed in the otic system in addition to its prominent expression at the midbrain-hindbrain boundary. The no isthmus (noi) mutation is known to inactivate the Pax2.1 gene, thereby affecting the development of the midbrain-hindbrain boundary region, pronephric system, optic stalk and otic region. Although the different members of the Pax2/5/8 family may potentially compensate for the loss of Pax2.1 function, we demonstrate here that only the expression of the Pax2.2 gene remains unaffected in noi mutant embryos. The expression of Pax5 and Pax8 is either not initiated at the midbrain-hindbrain boundary or is later not maintained in other expression domains. Consequently, the noi mutation of zebrafish is equivalent to combined inactivation of the mouse Pax2 and Pax5 genes with regard to the loss of midbrain-hindbrain boundary development.