bluuebunny/crossref_metadata_2025 huggingface dataset:
Hair matrix germinative epidermal cells confer follicle-inducing capabilities on dermal sheath and high passage papilla cells
Authors: Reynolds Amanda J. , Jahoda Colin A. B.
Published: April 2021

Low passage cultured dermal papilla cells from adult rats stimulate complete hair follicle neogenesis when re-implanted into heterotypic skin. In contrast, cultured sheath cells are non-inductive despite sharing other behav-ioural characteristics (a common lineage and in situ proximity) with papilla cells. However, since sheath cells can behave inductively in amputated follicles after regen-erating the papilla, this poses the question of what influ-ences the sheath to papilla cell transition? During recipro-cal tissue interactions specific epidermal cues are crucial to skin appendage development, and while in vivo assays to date have focussed on dermal interactive influence, our aim was to investigate epidermal potential. We have previously observed that hair follicle epidermal cells display excep-tional interactive behaviour when combined with follicle dermal cells in vitro. Thus in the present study, hair follicle germinative, outer root sheath or skin basal epidermal cells were separately combined with each of three non-inductive dermal cell types (high passage papilla, low passage sheath or fibroblast) and then implanted into small ear skin wounds. The sheath/germinative and papilla/germinative cell implants repeatedly induced giant vibrissa-type follicles and fibres. In complete contrast, any single cell type and all other forms of recombination were consistently non-inductive. Hence, the adult germinative epidermal cells enable non-inductive adult dermal cells to stimulate hair follicle neogenesis, effectively, by altering their ‘status’, causing the sheath cells to ‘specialise’ and the ‘aged’ papilla cells to ‘rejuvenate’.
Mice lacking tartrate-resistant acid phosphatase (Acp 5) have disrupted endochondral ossification and mild osteopetrosis
Authors: Hayman Alison R. , Jones Sheila J. , Boyde Alan , Foster Diane , Colledge William H. , Carlton Mark B. , Evans Martin J. , Cox Timothy M.
Published: April 2021

Mature osteoclasts specifically express the purple, band 5 isozyme (Acp 5) of tartrate-resistant acid phosphatase, a binuclear metalloenzyme that can generate reactive oxygen species. The function of Acp 5 was investigated by targeted disruption of the gene in mice. Animals homozygous for the null Acp 5 allele had progressive foreshortening and deformity of the long bones and axial skeleton but apparently normal tooth eruption and skull plate development, indicating a rôle for Acp 5 in endochondral ossification. Histomorphometry and mineralization density analysis of backscattered electron imaging revealed widened and dis-organized epiphyseal growth plates with delayed mineralization of cartilage in 6- to 8-week-old mutant mice. The membrane bones of the skull showed increased density at all ages examined, indicating defective osteoclastic bone turnover. Increased mineralization density was observed in the long bones of older animals which showed modelling deformities at their extremities: heterozygotes and homozygous Acp 5 mutant mice had tissue that was more mineralized and occupied a greater proportion of the bone in all regions. Thus the findings reflect a mild osteopetrosis due to an intrinsic defect of osteoclastic modelling activity that was confirmed in the resorption pit assay in vitro. We conclude that this bifunctional metalloprotein of the osteoclast is required for normal mineralization of cartilage in developing bones; it also maintains integrity and turnover of the adult skeleton by a critical contribution to bone matrix resorption.
A role for cadherins in tissue formation
Authors: Larue Lionel , Antos Christopher , Butz Stefan , Huber Otmar , Delmas Véronique , Dominis Mara , Kemler Rolf
Published: April 2021

We have produced null mutant mouse embryonic stem cells for the cell adhesion molecule E-cadherin. Such E-cadherin−/− ES cells are defective in cell aggregation; this defect can be corrected by transfection with cDNA for either E-cadherin or N-cadherin driven by a constitutive promoter. The presence (or absence) of E-cadherin regulates the expression of the transcription factor T-brachyury, indicating that cadherins play a role in linking cell surface receptors and gene expression. Comparative analysis of the parental and the genetically altered ES cell lines was performed to examine cell differentiation and the capability to form organized tissues. While differentiating E-cadherin−/− ES cells are still able to express various early and late differentiation markers, they show a clear-cut deficiency in forming organized structures. This phenotype can be rescued by constitutive expression of E-cadherin, which results exclusively in formation of epithelia. In contrast, rescue transfectants expressing N-cadherin show no epithelial structures, instead forming neuroepithelium and cartilage. These results provide the first evidence that specific cadherins directly stimulate differentiation into certain types of tissues.
The Drosophila fish-hook gene encodes a HMG domain protein essential for segmentation and CNS development
Authors: Nambu Patricia A. , Nambu John R.
Published: April 2021

We describe the isolation and analysis of the Drosophila fish-hook (fish) gene, which encodes a novel member of the SOX subgroup of High Mobility Group (HMG) domain proteins that exhibit similarity to the mammalian testis determining factor, SRY. The fish gene is initially expressed in a pair-rule-like pattern which is rapidly replaced by strong neuroectoderm expression. fish null mutants exhibit severe segmentation defects, including loss and/or fusion of abdominal denticle belts and stripe-specific defects in pair-rule and segment polarity gene expression. fish mutant embryos also exhibit loss of specific neurons, fusion of adjacent ventral nerve cord ganglia and aberrant axon scaffold organization. These results indicate an essential role for fish in anterior/posterior pattern formation and nervous system development, and suggest a potential function in modulating the activities of gap and pair-rule proteins.
Patterning the dorsal longitudinal flight muscles (DLM) of Drosophila: insights from the ablation of larval scaffolds
Authors: Fernandes Joyce J. , Keshishian Haig
Published: April 2021

The six Dorsal Longitudinal flight Muscles (DLMs) of Drosophila develop from three larval muscles that persist into metamorphosis and serve as scaffolds for the formation of the adult fibers. We have examined the effect of muscle scaffold ablation on the development of DLMs during metamorphosis. Using markers that are specific to muscle and myoblasts we show that in response to the ablation, myoblasts which would normally fuse with the larval muscle, fuse with each other instead, to generate the adult fibers in the appropriate regions of the thorax. The development of these de novo DLMs is delayed and is reflected in the delayed expression of erect wing, a transcription factor thought to control differentiation events associated with myoblast fusion. The newly arising muscles express the appropriate adult-specific Actin isoform (88F), indicating that they have the correct muscle identity.

However, there are frequent errors in the number of muscle fibers generated. Ablation of the larval scaffolds for the DLMs has revealed an underlying potential of the DLM myoblasts to initiate de novo myogenesis in a manner that resembles the mode of formation of the Dorso-Ventral Muscles, DVMs, which are the other group of indirect flight muscles. Therefore, it appears that the use of larval scaffolds is a superimposition on a commonly used mechanism of myogenesis in Drosophila. Our results show that the role of the persistent larval muscles in muscle patterning involves the partitioning of DLM myoblasts, and in doing so, they regulate formation of the correct number of DLM fibers.
The distal limb environment regulates MyoD accumulation and muscle differentiation in mouse-chick chimæric limbs
Authors: Robson Lesley G. , Hughes Simon M.
Published: April 2021

Differentiation of muscle and cartilage within developing vertebrate limbs occurs in a proximodistal progression. To investigate the cues responsible for regulating muscle pattern, mouse myoblasts were implanted into early chick wings prior to endogenous chick muscle differentiation. Fetal myogenic cells originating from transgenic mice carrying a lacZ reporter were readily detected in vivo after implantation and their state of differentiation determined with species-specific antibodies to MyoD and myosin heavy chain. When mouse myogenic cells are implanted at the growing tip of early stage 21 limbs MyoD expression is suppressed and little differentiation of the mouse cells is detected initially. At later stages ectopically implanted mouse cells come to lie within muscle masses, re-express MyoD and differentiate in parallel with differentiating chick myoblasts. However, if mouse cells are implanted either proximally at stage 21 or into the limb tip at stage 24, situations in which mouse cells encounter endogenous differentiating chick myoblasts earlier, MyoD suppression is not detected and a higher proportion of mouse cells dif- ferentiate. Mouse cells that remain distal to endogenous differentiating myogenic cells are more likely to remain undifferentiated than myoblasts that lie within differenti- ated chick muscle. Undifferentiated distal mouse cells are still capable of differentiating if explanted in vitro, sug- gesting that myoblast differentiation is inhibited in vivo. In vitro, MyoD is suppressed in primary mouse myoblasts by the addition of FGF2 and FGF4 to the culture media. Taken together, our data suggest that the inhibition of myogenic differentiation in the distal limb involves MyoD suppression in myoblasts, possibly through an FGF-like activity.
Genetic evidence for the subdivision of the arthropod limb into coxopodite and telopodite
Authors: González-Crespo Sergio , Morata Ginés
Published: April 2021

Arthropod appendages are thought to have evolved as out-growths from the body wall of a limbless ancestor. Snodgrass, in his Principles of Insect Morphology (1935), proposed that, during evolution, expansion of the body wall would originate the base of the appendages, or coxopodite, upon which the most distal elements that represent the true outer limb, or telopodite, would develop. The homeobox gene Distal-less (Dll), which is required in the Drosophila appendages for development of distal regions, has been proposed to promote formation of telopodite structures above the evolutionary ground-state of non-limb or body wall. Here, we present evidence that another homeobox gene, extradenticle (exd), which is required for appropriate development of the trunk and the proximal parts of the appendages, represents a coxopodite gene. We show that exd function is eliminated from the distal precursors in the developing limb and remains restricted to proximal precursors throughout development. This elimination is important because, when ectopically expressed, exd prevents distal development and gives rise to truncated appendages lacking distal elements. Moreover, the maintenance of exd expression during larval stages, contrary to Dll, does not require the hedgehog (hh) signaling pathway, suggesting that the proximal regions of the appendages develop independently of hh function. Finally, we show that in the crustacean Artemia, exd and Dll are expressed in comparable patterns as in Drosophila, suggesting a conserved genetic mechanism subdividing the arthropod limb.
Expression cloning of a Xenopus T-related gene (Xombi) involved in mesodermal patterning and blastopore lip formation
Authors: Lustig K. D. , Kroll K. L. , Sun E. E. , Kirschner M. W.
Published: April 2021

We have used a functional assay to identify a putative T-box transcription factor (Xombi) that has the ability to induce sites of invagination in the ectoderm of Xenopus embryos that resemble the blastopore lip. Maternal Xombi transcript is first localized to the oocyte’s vegetal cortex and cytoplasm, early sources of mesoderm and endoderm-inducing signals. Soon after zygotic transcription begins, there is a wave of Xombi expression, beginning in dorsal mesoderm and then extending to lateral and ventral mesoderm, that precedes and parallels blastopore lip formation at the border between the mesoderm and endoderm. Transcripts encoding brachyury, Xwnt8 and goosecoid colocalize with Xombi transcripts within the marginal zone; ectopic expression of Xombi induces expression of all three mesodermal genes. In ectodermal explants, Xombi expression is induced by the secreted mesoderm inducers activinA, activinB, Xnr1 and eFGF, and by brachyury, another Xenopus T-box containing gene. The time course and location of Xombi expression, its bio-logical activities and the partial dependence of Xombi expression and blastopore lip formation on fibroblast growth factor (FGF) signaling suggest that Xombi con-tributes to a traveling wave of morphogenesis and differ-entiation during Xenopus gastrulation.
Somatic sex-determining signals act on XX germ cells in Drosophila embryos
Authors: Staab Susanne , Heller Astrid , Steinmann-Zwicky Monica
Published: April 2021

In Drosophila, the enhancer-trap line mgm1 is already specifically expressed in male germ cells. Staining is first detected in 10-hour-old embryos and it is found in later stem cells. This line, which reveals the earliest sex-specific gene expression in the germline known so far, is a useful molecular marker to assess the sexual pathway that germ cells have entered before any overt sexual dimorphism is apparent. XY germ cells that develop in feminized animals express mgm1, which shows that this marker is autonomously expressed in XY germ cells. However, XX germ cells that develop in masculinized animals also express mgm1. Therefore, somatic sex-determining signals have already acted on XX germ cells in 10-hour-old embryos.
Compartments, wingless and engrailed: patterning the ventral epidermis of Drosophila embryos
Authors: Lawrence Peter A. , Sanson Bénédicte , Vincent Jean-Paul
Published: April 2021

Recent experiments on the wing disc of Drosophila have shown that cells at the interface between the anterior and posterior compartments drive pattern formation by becoming the source of a morphogen. Here we ask whether this model applies to the ventral embryonic epidermis. First, we show that interfaces between posterior (engrailed ON) and anterior (engrailed OFF) cells are required for pattern formation. Second, we provide evidence that Wingless could play the role of the morphogen, at least within part of the segmental pattern. We looked at the cuticular structures that develop after different levels of uniform Wingless activity are added back to unsegmented embryos (wingless−engrailed−). Because it is rich in landmarks, the T1 segment is a good region to analyse. There, we find that the cuticle formed depends on the amount of added Wingless activity. For example, a high concentration of Wingless gives the cuticle elements normally found near the top of the presumed gradient.

Unsegmented embryos are much shorter than wild type. If Wingless activity is added in stripes, the embryos are longer than if it is added uniformly. We suggest that the Wingless gradient landscape affects the size of the embryo, so that steep slopes would allow cells to survive and divide, while an even distribution of morphogen would promote cell death. Supporting the hypothesis that Wingless acts as a morphogen, we find that these stripes affect, at a distance, the type of cuticle formed and the planar polarity of the cells.
The three dominant female-sterile mutations of the Drosophila ovo gene are point mutations that create new translation-initiator AUG codons
Authors: Mével-Ninio Maryvonne , Fouilloux Eric , Guénal Isabelle , Vincent Alain
Published: April 2021

The Drosophila ovo gene, which encodes a putative transcription factor (Ovo) with TFIIIA-like zinc fingers, is required for female germline survival and proper oogenesis. Three dominant female-sterile ovoD mutations cause ovarian abnormalities that define an allelic series, with ovoD1 displaying the stronger phenotype and ovoD3 the weaker. We report here that all three ovoD mutations are point mutations that create new in-frame methionine codons in the 5′ part of ovo. There are two types of over-lapping ovo transcription units, ovoα and ovoβ. By using various ovo-lacZ reporter genes, we determined that the long Ovo isoforms starting at methionine M1, present in transcripts ovoα, are expressed at low levels only in mature oocytes. Short Ovo isoforms are translated from methionine M373, the first in-frame start codon present in tran-script ovoβ, and correspond to the activity defined by recessive loss of function ovo mutations. The new AUGs created in ovoD mutations all are located upstream of the M373 initiation site. Our results support the hypothesis that they can substitute for M373 as translation starts and initiate the synthesis of Ovo proteins that have extra amino acids at their N termini. We propose that premature expression of long Ovo protein isoforms occurs in ovoD mutants and interferes with wild-type Ovo function in con-trolling female germline differentiation.
Two distinct roles for Ras in a developmentally regulated cell migration
Authors: Lee Tzumin , Feig Larry , Montell Denise J.
Published: April 2021

Receptor tyrosine kinases have been shown to promote cell movement in a variety of systems. The Ras protein, a well-documented downstream effector for receptor tyrosine kinases, may contribute to receptor tyrosine kinase-mediated motility. In the present study, we have examined the role of Ras in the migration of a small subset of follicle cells, known as the border cells, during Drosophila oogenesis. A dominant-negative Ras protein inhibited cell migration when expressed specifically in border cells during the period when these cells normally migrate. When expressed prior to migration, dominant-negative Ras promoted premature initiation of migration. Conversely, expression of constitutively active Ras prior to migration resulted in a significant delay in the initiation step. Fur-thermore, the defect in initiation of border cell migration found in slbo1, a mutation at the locus that encodes Drosophila C/EBP, was largely rescued by reducing Ras activity in border cells prior to migration. Taken together, these observations indicate that Ras activity plays two distinct roles in the border cells: (1) reduction in Ras activity promotes the initiation of that migration process and (2) Ras activity is required during border cell migration. We further examined the possible involvement of two downstream effectors of Ras in border cell migration. Raf activity was dispensable to border cell migration while reduced Ral activity inhibited initiation. We therefore suggest that Ras plays a critical role in the dynamic regulation of border cell migration via a Raf-inde-pendent pathway.
Lineage-specific morphogenesis in the developing pancreas: role of mesenchymal factors
Authors: Gittes George K. , Galante Philip E. , Hanahan Douglas , Rutter William J. , Debas Haile T.
Published: April 2021

Pancreatic organogenesis has been a classic example of epitheliomesenchymal interactions. The nature of this interaction, and the way in which endocrine, acinar and ductal cell lineages are generated from the embryonic foregut has not been determined. It has generally been thought that mesenchyme is necessary for all aspects of pancreatic development. In addition islets have been thought to derive, at least in part, from ducts. We microdissected 11-day embryonic mouse pancreas and developed several culture systems for assays of differentiation: (i) on transparent filters; (ii) suspended in a collagen I gel; (iii) suspended in a basement membrane rich gel; (iv) under the renal capsule of an adult mouse. Epithelia were grown either with or without mesenchyme, and then assayed histologically and immunohistochemically. Epithelium with its mesenchyme (growth systems i-iv) always grew into fully differentiated pancreas (acinar, endocrine, and ductal elements). In the basement membrane-rich gel, epithelium without mesenchyme formed ductal structures. Under the renal capsule of the adult mouse the epithelium without mesenchyme exclusively formed clusters of mature islets. These latter results represent the first demonstration of pure islets grown from early pancreatic precursor cells. In addition, these islets seemed not to have originated from ducts.

We propose that the default path for growth of embryonic pancreatic epithelium is to form islets. In the presence of basement membrane constituents, however, the pancreatic anlage epithelium appears to be programmed to form ducts. Mesenchyme seems not to be required for all aspects of pancreatic development, but rather only for the formation of acinar structures. In addition, the islets seem to form from early embryonic epithelium (which only express non-acinar genes). This formation occurs without any specific embryonic signals, and without any clear duct or acinus formation.
The tangled-1 mutation alters cell division orientations throughout maize leaf development without altering leaf shape
Authors: Smith Laurie G. , Hake Sarah , Sylvester Anne W.
Published: April 2021

It is often assumed that in plants, where the relative positions of cells are fixed by cell walls, division orientations are critical for the generation of organ shapes. However, an alternative perspective is that the generation of shape may be controlled at a regional level independently from the initial orientations of new cell walls. In support of this latter view, we describe here a recessive mutation of maize, tangled-1 (tan-1), that causes cells to divide in abnormal orientations throughout leaf development without altering overall leaf shape. In normal plants, leaf cells divide either transversely or longitudinally relative to the mother cell axis; transverse divisions are associated with leaf elongation and longitudinal divisions with leaf widening. In tan-1 mutant leaves, cells in all tissue layers at a wide range of developmental stages divide transversely at normal frequencies, but longitudinal divisions are largely substituted by a variety of aberrantly oriented divisions in which the new cell wall is crooked or curved. Mutant leaves grow more slowly than normal, but their overall shapes are normal at all stages of their growth. These observations demonstrate that the generation of maize leaf shape does not depend on the precise spatial control of cell division, and support the general view that mechanisms independent from the control of cell division orientations are involved in the generation of shape during plant development.
Maternal mRNA encoding the orphan steroid receptor SpCOUP-TF is localized in sea urchin eggs
Authors: Vlahou Antonia , Gonzalez-Rimbau Maria , Flytzanis Constantin N.
Published: April 2021

The SpCOUP-TF gene is a highly conserved sea urchin homologue of the vertebrate COUP-TFs and the Drosophila seven up subfamily of transcription factors, which are members of the orphan steroid hormone receptors. Whole-mount in situ hybridization experiments, using three sea urchin species, detect the maternal SpCOUP-TF mRNA deposited unevenly in the oocytes, mature eggs and the blastomeres of the early embryo. The localization pattern indicates that, in all three sea urchin species, the maternal SpCOUP-TF mRNA is placed in the egg in a fixed position relative to the embryonic axes, i.e. lateral to the animal-vegetal and at 45° angle to the oralaboral (ventral-dorsal) axis. The embryonic expression of the SpCOUP-TF gene is spatially restricted in the oral ectoderm of the early embryo and, at later stages, in the cells of the ciliated band, the neurogenic cell lineage of the sea urchin embryo. The SpCOUP-TF mRNA, the first sea urchin maternal mRNA encoding a transcription factor that is specifically localized with respect to both embryonic axes in the egg, could be involved in early specification events in the sea urchin embryo.
TGF-β signals and a prepattern in Xenopus laevis endodermal development
Authors: Henry Gilbert L. , Brivanlou Iman H. , Kessler Daniel S. , Hemmati-Brivanlou Ali , Melton Douglas A.
Published: April 2021

We have analyzed two gene products expressed in the early endoderm of Xenopus laevis: Xlhbox-8, a pancreas-specific transcription factor and intestinal fatty acid binding protein (IFABP), a marker of small intestinal epithelium. Expression of the pancreas marker relies on cell signaling mediated by both the TGFβ and FGF classes of secreted peptide growth factors, whereas, expression of the more posterior small intestinal marker does not. Endodermal explants devoid of mesoderm express both markers in a regionalized manner. Cortical rotation is required for the expression of the more anterior marker, Xlhbox-8, but not for the small intestinal marker, IFABP. These findings suggest that endodermal patterning is dependent, in part, on the same events and signals known to play important roles in mesodermal development. Furthermore, inhibition of TGFβ signaling in the endoderm leads to ectopic expression of both mesodermal and ectodermal markers, suggesting that TGFβ signaling may play a general role in the segregation of the three embryonic germ layers.
Ectopic expression of Hoxa-1 in the zebrafish alters the fate of the mandibular arch neural crest and phenocopies a retinoic acid-induced phenotype
Authors: Alexandre Daniel , Clarke Jonathan D. W. , Oxtoby Elli , Yan Yi-Lin , Jowett Trevor , Holder Nigel
Published: April 2021

Considerable evidence has demonstrated that retinoic acid influences the formation of the primary body axis in vertebrates and that this may occur through the regulation of Hox gene expression. In this study, we show that the phenotype induced by exogenous retinoic acid in the zebrafish can also be generated by the overexpression of Hoxa-1 following injection of synthetic RNA into the fertilised egg. The isolation, sequence and expression pattern of the zebrafish Hoxa-1 gene is described. We show that exogenously applied retinoic acid causes the ectopic accumulation of Hoxa-1 message during gastrulation in the hypoblast in the head region. Overexpression of Hoxa-1 following injection of RNA causes abnormal growth of the anterior hindbrain, duplication of Mauthner neurons in rhombomere (r) 2 and fate changes of r2 mesenchymal and neurogenic neural crest. These results are discussed in terms of the role of Hoxa-1 in controlling anterior hindbrain patterning and the relationship between expression of Hoxa-1 and retinoic acid.
A muscle-specific variant of microtubule-associated protein 4 (MAP4) is required in myogenesis
Authors: Mangan Mary E. , Olmsted J. B.
Published: April 2021

Microtubule-associated protein 4 (MAP4) transcripts vary in different mouse tissues, with striated muscle (skeletal and cardiac) expressing 8and 9-kb transcripts preferentially to the more widely distributed 5.5and 6.5-kb transcripts (West, R. W., Tenbarge, K. M. and Olmsted, J. B. (1991). J. Biol. Chem. 266, 21886-21896). Cloning of the sequence unique to the muscle transcripts demonstrated that these mRNAs vary from the more ubiquitous ones by a single 3.2-kb coding region insertion within the projection domain of MAP4. During differentiation of the myogenic cell line, C2C12, muscle-specific MAP4 transcripts appear within 24 hours of growth in differentiation medium, and a larger MAP4 isotype (350×103Mr) accumulates to high levels by 48 hours of differentiation. In situ hybridization analyses of transcript distribution in mouse embryos demonstrated that muscle-specific transcripts appear early in myogenesis. To block the expression of the muscle-specific MAP4, stable lines of C2C12 cells were generated bearing an antisense construct with the musclespecific MAP4 sequence. Myoblast growth was unaffected whereas myotube formation was severely perturbed. Fusion occurred in the absence of the muscle MAP4 isotype, but the multinucleate syncytia were short and apolar, microtubules were disorganized and normal anisotropic myofibrils were absent. The patterns of expression of the muscle-specific transcripts and the antisense experiments indicate that this unique structural form of MAP4 plays a critical role in the formation and maintenance of muscle.
Polyembryonic development: insect pattern formation in a cellularized environment
Authors: Grbic Miodrag , Nagy Lisa M. , Carroll Sean B. , Strand Michael
Published: April 2021

The polyembryonic wasp Copidosoma floridanum produces up to 2000 individuals from a single egg. During the production of individual embryos the original anteroposterior axis of the egg is lost and axial patterning must subsequently be reestablished within each embryo. The mechanism by which this occurs is unknown. In most insects, egg polarity is established during oogenesis and early development takes place in a syncytium. In Drosophila melanogaster, the syncytium is considered essential for establishing the morphogenetic gradients that initiate segmental patterning. However, we found that development of C. floridanum occurs almost exclusively in a cellularized environment. To determine whether the D. melanogaster patterning cascade is conserved in the absence of a syncytium, we analyzed the expression of Even-skipped, Engrailed and Ultrabithorax/Abdominal-A during polyembryonic development. Here we show that in spite of the absence of a syncytium, the elements of the D. melanogaster segmentation hierarchy are conserved. The segment-polarity gene Engrailed and the homeotic genes Ultrabithorax/Abdominal-A are expressed in a conserved pattern relative to D. melanogaster. However, we detect an alteration in the expression of the Even-skipped antigen. Even-skipped is initially expressed in segmentally reiterated stripes and not in a pair-rule pattern as it is in D. melanogaster. We also observe that the expression of these regulatory proteins does not occur during the early proliferative phases of polyembryony. Our results indicate that a syncytium is not required for segmental patterning in this insect.
Regional specification of motoneurons along the anterior-posterior axis is independent of the notochord
Authors: Fukushima Mikiko , Nakamura Masaaki , Ohta Kunimasa , Okamura Ryouichi , Negi Akira , Tanaka Hideaki
Published: April 2021

Cek8 and low affinity NGF receptor (LNGFR) are expressed at high levels on the chick spinal motoneurons of the brachial and lumbar segments from embryonic day (E) 5 to E7, but weakly on the motoneurons of the non-limbinnervating segments. We determined by means of heterotopic neural tube transplantation, that the expression of these molecules was already intrinsically determined at E2. We used these spatiotemporal specific molecules as markers of motoneuron subpopulations. To analyze how motoneurons acquire regional specification along the anterior-posterior (A-P) axis and in the transverse plane, we observed the expression of these molecules on ectopic motoneurons induced by implanting a supernumerary notochord or floor plate at E2. The ectopic motoneurons induced by the graft obtained from either the thoracic or lumbar segments had the same expression profile as the normal motoneurons at each A-P level. These findings suggest that regional specification of motoneurons, at least of Cek8 and LNGFR expression, is independent of the notochord and the floor plate and that the whole neural tube appears to be committed to differentiate into the motoneuron subtypes along the A-P axis at the operative stages.
Gliolectin is a novel carbohydrate-binding protein expressed by a subset of glia in the embryonic Drosophila nervous system
Authors: Tiemeyer Michael , Goodman Corey S.
Published: April 2021

Interactions between embryonic neural cells generate the specific patterns of connectivity observed in nervous systems. Cell surface carbohydrates have been proposed to function in cellular recognition events guiding such interactions. Carbohydrate-binding proteins (lectins) that recognize specific oligosaccharide ligands in embryonic neural tissue provide a molecular mechanism for carbohydrate-mediated cell-cell interactions in neural development. Therefore, we have screened an embryonic Drosophila melanogaster cDNA library, expressed in COS1 cells, for carbohydrate-binding activity. COS1 cells expressing putative Drosophila lectins were identified and recovered based on their adhesion to immobilized preparations of neutral and zwitterionic glycolipids extracted from Drosophila embryos. We have identified an endogenous lectin expressed during Drosophila embryogenesis. The cloned lectin, designated ‘gliolectin’, possesses a novel protein sequence with a calculated molecular mass of 24,993. When expressed in Drosophila S2 cells, the lectin mediates heterophilic cellular aggregation. In embryos, gliolectin is expressed by a subset of glial cells found at the midline of the developing nervous system. Expression is highest during the formation of the Drosophila embryonic axonal commissures, a process requiring midline glial cell function. Immunoprecipitation with a monoclonal antibody against gliolectin yields a protein of Mr=46,600 from Drosophila embryonic membranes, suggesting that post-translational modification of gliolectin is extensive. Epitope-tagged chimeric proteins composed of the amino terminal one-half of gliolectin and the Fc region of human IgG bind a small subset of the total glycolipids extracted from Drosophila embryos, demonstrating that the lectin activity of gliolectin can discriminate between oligosaccharide structures. The presence of gliolectin in the developing Drosophila embryonic nervous system further supports a role for cell surface carbohydrates in cell-cell recognition and indicates that the molecular diversity of animal lectins is not yet completely defined.
The Drosophila genes crumbs and stardust are involved in the biogenesis of adherens junctions
Authors: Grawe Ferdi , Wodarz Andreas , Lee Bill , Knust Elisabeth , Skaer Helen
Published: April 2021

Morphogenetic movements of epithelia during development underlie the normal elaboration of the final body plan. The tissue integrity critical for these movements is conferred by anchorage of the cytoskeleton to the membrane at sites of cell adhesion, mediated by adherens junctions, initially spot and later belt-like, zonular structures, which encircle the apical side of the cell. Loss-offunction mutations in the Drosophila genes crumbs and stardust lead to the loss of cell polarity in most ectodermally derived epithelia, followed in some, such as the epidermis, by extensive apoptosis. Here we show that both mutants fail to establish proper zonulae adherentes in the epidermis. Our results suggest that the two genes are involved in different aspects of this process. Further, they are compatible with the hypothesis that crumbs delimits the apical border, where the zonula adherens usually forms and where Crumbs protein is normally most abundant. In contrast, stardust seems to be required at an earlier stage for the assembly of the spot adherens junctions. In both mutants, the defects observed at the ultrastructural level are preceded by a misdistribution of Armadillo and DEcadherin, the homologues of β-catenin and E-cadherin, respectively, which are two constituents of the vertebrate adherens junctions. Strikingly, expansion of the apical membrane domain in epidermal cells by overexpression of crumbs also abolishes the formation of adherens junctions and results in the disruption of tissue integrity, but without loss of membrane polarity. This result supports the view that membrane polarity is independent of the formation of adherens junctions in epidermal cells.
Initiation patterns of flower and floral organ development in Arabidopsis thaliana
Authors: Bossinger Gerd , Smyth David R.
Published: April 2021

Sector boundary analysis has been used to deduce the number and orientation of cells initiating flower and floral organ development in Arabidopsis thaliana. Sectors were produced in transgenic plants carrying the Ac transposon from maize inserted between the constitutive 35S promoter and the GUS reporter gene. Excision of the transposon results in a blue-staining sector. Plants were chosen in which an early arising sector passed from vegetative regions into the inflorescence and through a mature flower. The range of sector boundary positions seen in mature flowers indicated that flower primordia usually arise from a group of four cells on the inflorescence flank. The radial axes of the mature flower are apparently set by these cells, supporting the concept that they act as a structural template. Floral organs show two patterns of initiation, a leaf-like pattern with eight cells in a row (sepals and carpels), or a shoot-like pattern with four cells in a block (stamens). The petal initiation pattern involved too few cells to allow assignment. The numbers of initiating cells were close to those seen when organ growth commenced in each case, indicating that earlier specification of floral organ development does not occur. By examining sector boundaries in homeotic mutant flowers in which second whorl organs develop as sepal-like organs rather than petals, we have shown that their pattern of origin is position dependent rather than identity dependent.
The GAGA factor is required in the early Drosophila embryo not only for transcriptional regulation but also for nuclear division
Authors: Bhat Krishna Moorthi , Farkas Gabriella , Karch Francois , Gyurkovics Henrik , Gausz Janos , Schedl Paul
Published: April 2021

The GAGA protein of Drosophila was first identified as a stimulatory factor in in vitro transcription assays using the engrailed and Ultrabithorax promoters. Subsequent studies have suggested that the GAGA factor promotes transcription by blocking the repressive effects of histones; moreover, it has been shown to function in chromatin remodeling, acting together with other factors in the formation of nuclease hypersensitive sites in vitro. The GAGA factor is encoded by the Trithorax-like locus and in the studies reported here we have used the maternal effect allele Trl13C to examine the functions of the protein during embryogenesis. We find that GAGA is required for the proper expression of a variety of developmental loci that contain GAGA binding sites in their upstream regulatory regions. The observed disruptions in gene expression are consistent with those expected for a factor involved in chromatin remodeling. In addition to facilitating gene expression, the GAGA factor appears to have a more global role in chromosome structure and function. This is suggested by the spectrum of nuclear cleavage cycle defects observed in Trl13C embryos. These defects include asynchrony in the cleavage cycles, failure in chromosome condensation, abnormal chromosome segregation and chromosome fragmentation. These defects are likely to be related to the association of the GAGA protein with heterochromatic satellite sequences which is observed throughout the cell cycle.
The SH2-containing tyrosine phosphatase corkscrew is required during signaling by sevenless, Ras1 and Raf
Authors: Allard John D. , Chang Henry C. , Herbst Ronald , McNeill Helen , Simon Michael A.
Published: April 2021

The sevenless gene encodes a receptor tyrosine kinase which is required for the development of the R7 photoreceptor cell in each ommatidium of the Drosophila eye. We have previously used a sensitized genetic screen to identify mutations, designated Enhancers of sevenless (E(sev)), which affect genes that encode components of the sevenless signaling pathway. Here, we report that one of these mutations, E(sev)1Ae0P is a dominantly inhibiting allele of corkscrew, which encodes an SH2 domain-containing protein tyrosine phosphatase (Perkins et al., 1992). We show that corkscrew function is essential for sevenless signaling and that expression of a membrane-targeted form of corkscrew can drive R7 photoreceptor development in the absence of sevenless function. Furthermore, we have used the dominantly inhibiting corkscrew allele to examine the role of corkscrew during signaling by activated forms of Ras1 and Raf. Our analysis indicates that corkscrew function is still required during signaling by activated Ras1 and Raf proteins. These results define a function for corkscrew that is either downstream of Ras1 activation or in a parallel pathway that acts with activated Ras1/Raf to specify R7 photoreceptor development.
The Drosophila morphogenetic protein Bicoid binds DNA cooperatively
Authors: Ma Xiuguang , Yuan Dong , Diepold Katharina , Scarborough Tom , Ma Jun
Published: April 2021

The Drosophila morphogenetic protein Bicoid, encoded by the maternal gene bicoid, is required for the development of the anterior structures in the embryo. Bicoid, a transcriptional activator containing a homeodomain, is distributed in an anterior-to-posterior gradient in the embryo. In response to this gradient, the zygotic gene hunchback is expressed uniformly in the anterior half of the embryo in a nearly all- or-none manner. In this report we demonstrate that a recombinant Bicoid protein binds cooperatively to its sites within a hunchback enhancer element. A less than 4-fold increase in Bicoid concentration is sufficient to achieve an unbound/bound transition in DNA binding. Using various biochemical and genetic methods we further demonstrate that Bicoid molecules can interact with each other. Our results are consistent with previous studies performed in the embryo, and they suggest that one mechanism to achieve a sharp on/off switch of gene expression in response to a morphogenetic gradient is cooperative DNA binding facilitated by protein-protein interaction.
Conservation in hedgehog signaling: induction of a chicken patched homolog by Sonic hedgehog in the developing limb
Authors: Marigo Valeria , Scott Matthew P. , Johnson Ronald L. , Goodrich Lisa V. , Tabin Clifford J.
Published: April 2021

Hedgehog genes have been implicated in inductive signaling during development in a variety of organisms. A key element of the hedgehog signaling system is encoded by the gene patched. In Drosophila hedgehog regulates gene expression by antagonizing the action of patched. In addition, patched is itself a transcriptional target of hedgehog signaling.

We have isolated a chicken patched homolog and find it to be strongly expressed adjacent to all tissues where members of the hedgehog family are expressed. As in Drosophila, ectopic expression of Sonic hedgehog leads to ectopic induction of chicken Patched. Based on this regulatory conservation, vertebrate Patched is likely to be directly downstream of Sonic hedgehog signaling.

An important role of Sonic hedgehog is the regulation of anterior/posterior pattern in the developing limb bud. Since Patched is directly downstream of the hedgehog signal, the extent of high level Patched expression provides a measure of the distance that Sonic hedgehog diffuses and directly acts. On this basis, we find that Sonic hedgehog directly acts as a signal over only the posterior third of the limb bud.

During limb patterning, secondary signals are secreted in both the mesoderm (e.g. Bone Morphogenetic Protein-2) and apical ectodermal ridge (e.g. Fibroblast Growth Factor-4) in response to Sonic hedgehog. Thus knowing which is the direct target tissue is essential for unraveling the molecular patterning of the limb. The expression of Patched provides a strong indication that the mesoderm and not the ectoderm is the direct target of Sonic hedgehog signaling in the limb bud.

Finally we demonstrate that induction of Patched requires Sonic hedgehog but, unlike Bone Morphogenetic Protein-2 and Hox genes, does not require Fibroblast Growth Factor as a co-inducer. It is therefore a more direct target of Sonic hedgehog than previously reported pat-terning genes.
Segregation of germ granules in living Caenorhabditis elegans embryos: cell-type-specific mechanisms for cytoplasmic localisation
Authors: Hird Steven N. , Paulsen Janet E. , Strome Susan
Published: April 2021

Germ granules are ribonucleoprotein particles that are thought to function in germline specification in invertebrates and possibly in vertebrates. In Caenorhabditis elegans, these structures, termed P granules, are partitioned to the germline P cells during the early embryonic divisions. By injecting a fluorescently labelled anti-P-granule antibody into the C. elegans germline syncitium, we followed P-granule segregation in live embryos using laser-scanning confocal microscopy. We show that, in early P cells (P0 and P1), P-granule partitioning is achieved primarily by their migration through the cytoplasm towards the site of formation of the germline daughter cell. A different mechanism appears to operate in later P cells (P2 and P3): P granules associate with the nucleus and move with it toward the site of formation of the germline daughter cell, where they are then deposited. At each division, there is also disassembly or degradation of those P granules that remain in the cytoplasm destined for the somatic daughter cell. Microfilaments, microtubules and the product of the gene mes-1 are required for the normal pattern of P-granule segregation in P2.
Characterizing the zebrafish organizer: microsurgical analysis at the early-shield stage
Authors: Shih John , Fraser Scott E.
Published: April 2021

The appearance of the embryonic shield, a slight thickening at the leading edge of the blastoderm during the formation of the germ ring, is one of the first signs of dorsoventral polarity in the zebrafish embryo. It has been proposed that the shield plays a role in fish embryo patterning similar to that attributed to the amphibian dorsal lip. In a recent study, we fate mapped many of the cells in the region of the forming embryonic shield, and found that neural and mesodermal progenitors are intermingled (Shih, J. and Fraser, S. E. (1995) Development 121, 2755-2765), in contrast to the coherent region of mesodermal progenitors found at the amphibian dorsal lip. Here, we examine the fate and the inductive potential of the embryonic shield to determine if the intermingling reflects a different mode of embryonic patterning than that found in amphibians. Using the microsurgical techniques commonly used in amphibian and avian experimental embryology, we either grafted or deleted the region of the embryonic shield. Homotopic grafting experiments confirmed the fates of cells within the embryonic shield region, showing descendants in the hatching gland, head mesoderm, notochord, somitic mesoderm, endoderm and ventral aspect of the neuraxis. Heterotopic grafting experiments demonstrated that the embryonic shield can organize a second embryonic axis; however, contrary to our expectations based on amphibian research, the graft contributes extensively to the ectopic neuraxis. Microsurgical deletion of the embryonic shield region at the onset of germ ring formation has little effect on neural development: embryos with a well-formed and well-patterned neuraxis develop in the complete absence of notochord cells. While these results show that the embryonic shield is sufficient for ectopic axis formation, they also raise questions concerning the necessity of the shield region for neural induction and embryonic patterning after the formation of the germ ring.