bluuebunny/crossref_metadata_2025 huggingface dataset:
Wnt-7a in feather morphogenesis: involvement of anterior-posterior asymmetry and proximal-distal elongation demonstrated with an in vitro reconstitution model
Authors: Widelitz Randall B. , Jiang Ting-Xin , Janet Chen Chia-Wei , Stott N. Susan , Jung Han-Sung , Chuong Cheng-Ming
Published: April 2021

How do vertebrate epithelial appendages form from the flat epithelia? Following the formation of feather placodes, the previously radially symmetrical primordia become anterior-posterior (A-P) asymmetrical and develop a proximo-distal (P-D) axis. Analysis of the molecular heterogeneity revealed a surprising parallel of molecular profiles in the A-P feather buds and the ventral-dorsal (V-D) Drosophila appendage imaginal discs. The functional significance was tested with an in vitro feather reconstitution model. Wnt-7a expression initiated all over the feather tract epithelium, intensifying as it became restricted first to the primordia domain, then to an accentuated ring pattern within the primordia border, and finally to the posterior bud. In contrast, sonic hedgehog expression was induced later as a dot within the primordia. RCAS was used to overexpress Wnt-7a in reconstituted feather explants derived from stage 29 dorsal skin to further test its function in feather formation. Control skin formed normal elongated, slender buds with A-P orientation, but Wnt-7a overexpression led to plateau-like skin appendages lacking an A-P axis. Feathers in the Wnt-7a overexpressing skin also had inhibited elongation of the P-D axes. This was not due to a lack of cell proliferation, which actually was increased although randomly distributed. While morphogenesis was perturbed, differentiation proceeded as indicated by the formation of barb ridges. Wnt-7a buds have reduced expression of anterior (Tenascin) bud markers. Middle (Notch-1) and posterior bud markers including Delta-1 and Serrate-1 were diffusely expressed. The results showed that ectopic Wnt-7a expression enhanced properties characteristic of the middle and posterior feather buds and suggest that P-D elongation of vertebrate skin appendages requires balanced interactions between the anterior and posterior buds.
Axial mesendoderm refines rostrocaudal pattern in the chick nervous system
Authors: Rowan Autumn M. , Stern Claudio D. , Storey Kate G.
Published: April 2021

There has long been controversy concerning the role of the axial mesoderm in the induction and rostrocaudal patterning of the vertebrate nervous system. Here we investigate the neural inducing and regionalising properties of defined rostrocaudal regions of head process/prospective notochord in the chick embryo by juxtaposing these tissues with extraembryonic epiblast or neural plate explants. We localise neural inducing signals to the emerging head process and using a large panel of region-specific neural markers, show that different rostrocaudal levels of the head process derived from headfold stage embryos can induce discrete regions of the central nervous system. However, we also find that rostral and caudal head process do not induce expression of any of these molecular markers in explants of the neural plate. During normal development the head process emerges beneath previously induced neural plate, which we show has already acquired some rostrocaudal character. Our findings therefore indicate that discrete regions of axial mesendoderm at headfold stages are not normally responsible for the establishment of rostrocaudal pattern in the neural plate. Strikingly however, we do find that caudal head process inhibits expression of rostral genes in neural plate explants. These findings indicate that despite the ability to induce specific rostrocaudal regions of the CNS de novo, signals provided by the discrete regions of axial mesendoderm do not appear to establish regional differences, but rather refine the rostrocaudal character of overlying neuroepithelium.
her1, a zebrafish pair-rule like gene, acts downstream of notch signalling to control somite development
Authors: Takke Christina , Campos-Ortega José A.
Published: April 2021

During vertebrate embryonic development, the paraxial mesoderm becomes subdivided into metameric units known as somites. In the zebrafish embryo, genes encoding homologues of the proteins of the Drosophila NOTCH signalling pathway are expressed in the presomitic mesoderm and expression is maintained in a segmental pattern during somitogenesis. This expression pattern suggests a role for these genes during somite development. We misexpressed various zebrafish genes of this group by injecting mRNA into early embryos. RNA encoding a constitutively active form of NOTCH1a (notch1a-intra) and a truncated variant of deltaD [deltaD(Pst)], as well as transcripts of deltaC and deltaD, the hairy-E(spl) homologues her1 and her4, and groucho2 were tested for their effects on somite formation, myogenesis and on the pattern of transcription of putative downstream genes. In embryos injected with any of these RNAs, with the exception of groucho2 RNA, the paraxial mesoderm differentiated normally into somitic tissue, but failed to segment correctly. Activation of NOTCH results in ectopic activation of her1 and her4. This misregulation of the expression of her genes might be causally related to the observed mesodermal defects, as her1 and her4 mRNA injections led to effects similar to those seen with notch1a-intra. deltaC and deltaD seem to function after subdivision of the presomitic mesoderm, since the her gene transcription pattern in the presomitic mesoderm remains essentially normal after misexpression of delta genes. Whereas NOTCH signalling alone apparently does not affect myogenesis, zebrafish groucho2 is involved in differentiation of mesodermal derivatives.
Increased hemangioblast commitment, not vascular disorganization, is the primary defect in flt-1 knock-out mice
Authors: Fong Guo-Hua , Zhang Liyong , Bryce Dawn-Marie , Peng Jun
Published: April 2021

We previously demonstrated the essential role of the flt-1 gene in regulating the development of the cardiovascular system. While the inactivation of the flt-1 gene leads to a very severe disorganization of the vascular system, the primary defect at the cellular level was unknown. Here we report a surprising finding that it is an increase in the number of endothelial progenitors that leads to the vascular disorganization in flt-1−/− mice. At the early primitive streak stage (prior to the formation of blood islands), hemangioblasts are formed much more abundantly in flt-1−/− embryos. This increase is primarily due to an alteration in cell fate determination among mesenchymal cells, rather than to increased proliferation, migration or reduced apoptosis of flt-1−/− hemangioblasts. We further show that the increased population density of hemangioblasts is responsible for the observed vascular disorganization, based on the following observations: (1) both flt-1−/− and flt-1+/+ endothelial cells formed normal vascular channels in chimaeric embryos; (2) wild-type endothelial cells formed abnormal vascular channels when their population density was significantly increased; and (3) in the absence of wild-type endothelial cells, flt-1−/− endothelial cells alone could form normal vascular channels when sufficiently diluted in a developing embryo. These results define the primary defect in flt-1−/− embryos at the cellular level and demonstrate the importance of population density of progenitor cells in pattern formation.
Role of PDGF-B and PDGFR-β in recruitment of vascular smooth muscle cells and pericytes during embryonic blood vessel formation in the mouse
Authors: Hellström Mats , Kalén Mattias , Lindahl Per , Abramsson Alexandra , Betsholtz Christer
Published: April 2021

Development of a vascular system involves the assembly of two principal cell types – endothelial cells and vascular smooth muscle cells/pericytes (vSMC/PC) – into many different types of blood vessels. Most, if not all, vessels begin as endothelial tubes that subsequently acquire a vSMC/PC coating. We have previously shown that PDGF-B is critically involved in the recruitment of pericytes to brain capillaries and to the kidney glomerular capillary tuft. Here, we used desmin and α-smooth muscle actin (ASMA) as markers to analyze vSMC/PC development in PDGF-B−/− and PDGFR-β−/− embryos. Both mutants showed a site-specific reduction of desmin-positive pericytes and ASMA-positive vSMC. We found that endothelial expression of PDGF-B was restricted to immature capillary endothelial cells and to the endothelium of growing arteries. BrdU labeling showed that PDGFR-β-positive vSMC/PC progenitors normally proliferate at sites of endothelial PDGF-B expression. In PDGF-B−/− embryos, limb arterial vSMC showed a reduced BrdU-labeling index. This suggests a role of PDGF-B in vSMC/PC cell proliferation during vascular growth. Two modes of vSMC recruitment to newly formed vessels have previously been suggested: (1) de novo formation of vSMC by induction of undifferentiated perivascular mesenchymal cells, and (2) co-migration of vSMC from a preexisting pool of vSMC. Our data support both modes of vSMC/PC development and lead to a model in which PDGFR-β-positive vSMC/PC progenitors initially form around certain vessels by PDGF-B-independent induction. Subsequent angiogenic sprouting and vessel enlargement involves PDGF-B-dependent vSMC/PC progenitor co-migration and proliferation, and/or PDGF-B-independent new induction of vSMC/PC, depending on tissue context.
Characterization of the transvection mediating region of the Abdominal-B locus in Drosophila
Authors: Zhou Jumin , Ashe Hilary , Burks Christian , Levine Michael
Published: April 2021

Genetic studies have identified an unusual transvection process in the Abdominal-B (Abd-B) locus of Drosophila. In some cases distal infraabdominal (iab) regulatory domains continue to activate the Abd-B promoter even when translocated onto different chromosomes. Transvection depends on an approx. 10 kb genomic DNA sequence, termed the transvection mediating region (tmr), located immediately downstream of the Abd-B transcription unit. Here we report a detailed analysis of this region. Different DNA fragments from the tmr were inserted into a variety of P-transformation vectors. Analyses of reporter gene expression in transgenic embryos and adults identify at least three cis-regulatory elements, including two enhancers (IAB7 and IAB8) and a new insulator DNA (Frontabdominal-8, Fab-8). Evidence is also presented for a Polycomb Response Element (PRE) linked to the IAB8 enhancer, and an internal promoter in the iab-8 domain, which transcribes the iab-7 and iab-8 cis-regulatory DNA, including the Fab-8 insulator. We discuss the significance of these findings with regard to Abd-B transvection and long-range enhancer-promoter interactions in mammalian globin loci.
The role of tolloid/mini fin in dorsoventral pattern formation of the zebrafish embryo
Authors: Connors Stephanie A. , Trout Jamie , Ekker Marc , Mullins Mary C.
Published: April 2021

A highly conserved TGF-β signaling pathway is involved in the establishment of the dorsoventral axis of the vertebrate embryo. Specifically, Bone Morphogenetic Proteins (Bmps) pattern ventral tissues of the embryo while inhibitors of Bmps, such as Chordin, Noggin and Follistatin, are implicated in dorsal mesodermal and neural development. We investigated the role of Tolloid, a metalloprotease that can cleave Chordin and increase Bmp activity, in patterning the dorsoventral axis of the zebrafish embryo. Injection of tolloid mRNA into six dorsalized mutants rescued only one of these mutants, mini fin. Through chromosomal mapping, linkage and cDNA sequence analysis of several mini fin alleles, we demonstrate that mini fin encodes the tolloid gene. Characterization of the mini fin mutant phenotype reveals that Mini fin/Tolloid activity is required for patterning ventral tissues of the tail: the ventral fin, and the ventroposterior somites and vasculature. Gene expression studies show that mfn mutants exhibit reduced expression of ventrally restricted markers at the end of gastrulation, suggesting that the loss of ventral tail tissues is caused by a dorsalization occurring at the end of gastrulation. Based on the mini fin mutant phenotype and the expression of tolloid, we propose that Mini fin/Tolloid modifes the Bmp activity gradient at the end of gastrulation, when the ventralmost marginal cells of the embryo are in close proximity to the dorsal Chordin-expressing cells. At this time, unimpeded Chordin may diffuse to the most ventral marginal regions and inhibit high Bmp activity levels. In the presence of Mini fin/Tolloid, however, Chordin activity would be negatively modulated through proteolytic cleavage, thereby increasing Bmp signaling activity. This extracellular mechanism is amplified by an autoregulatory loop for bmp gene expression.
SIAMESE, a gene controlling the endoreduplication cell cycle in Arabidopsis thaliana trichomes
Authors: Walker Jason D. , Oppenheimer David G. , and Joshua Concienne , Larkin John C.
Published: April 2021

Cell differentiation is generally tightly coordinated with the cell cycle, typically resulting in a nondividing cell with a unique differentiated morphology. The unicellular trichomes of Arabidopsis are a well-established model for the study of plant cell differentiation. Here, we describe a new genetic locus, SIAMESE (SIM), required for coordinating cell division and cell differentiation during the development of Arabidopsis trichomes (epidermal hairs). A recessive mutation in the sim locus on chromosome 5 results in clusters of adjacent trichomes that appeared to be morphologically identical ‘twins’. Upon closer inspection, the sim mutant was found to produce multicellular trichomes in contrast to the unicellular trichomes produced by wild-type (WT) plants. Mutant trichomes consisting of up to 15 cells have been observed. Scanning electron microscopy of developing sim trichomes suggests that the cell divisions occur very early in the development of mutant trichomes. WT trichome nuclei continue to replicate their DNA after mitosis and cytokinesis have ceased, and as a consequence have a DNA content much greater than 2C. This phenomenon is known as endoreduplication. Individual nuclei of sim trichomes have a reduced level of endoreduplication relative to WT trichome nuclei. Endoreduplication is also reduced in dark-grown sim hypocotyls relative to WT, but not in light-grown hypocotyls. Double mutants of sim with either of two other mutants affecting endoreduplication, triptychon (try) and glabra3 (gl3) are consistent with a function for SIM in endoreduplication. SIM may function as a repressor of mitosis in the endoreduplication cell cycle. Additionally, the relatively normal morphology of multicellular sim trichomes indicates that trichome morphogenesis can occur relatively normally even when the trichome precursor cell continues to divide. The sim mutant phenotype also has implications for the evolution of multicellular trichomes.
Cap ‘n’ collar B cooperates with a small Maf subunit to specify pharyngeal development and suppress Deformed homeotic function in the Drosophila head
Authors: Veraksa Alexey , McGinnis Nadine , Li Xuelin , Mohler Jym , McGinnis William
Published: April 2021

The basic-leucine zipper protein Cap ‘n’ collar B (CncB) suppresses the segmental identity function of the Hox gene Deformed (Dfd) in the mandibular segment of Drosophila embryos. CncB is also required for proper development of intercalary, labral and mandibular structures. In this study, we provide evidence that the CncB-mediated suppression of Dfd requires the Drosophila homolog of the mammalian small Maf proteins, Maf-S, and that the suppression occurs even in the presence of high amounts of Dfd protein. Interestingly, the CncB/Maf-S suppressive effect can be partially reversed by overexpression of Homothorax (Hth), suggesting that Hth and Extradenticle proteins antagonize the effects of CncB/Maf-S on Dfd function in the mandibular segment. In embryos, multimers of simple CncB/Maf-S heterodimer sites are transcriptionally activated in response to CncB, and in tissue culture cells the amino-terminal domain of CncB acts as a strong transcriptional activation domain. There are no good matches to CncB/Maf binding consensus sites in the known elements that are activated in response to Dfd and repressed in a CncB-dependent fashion. This suggests that some of the suppressive effect of CncB/Maf-S proteins on Dfd protein function might be exerted indirectly, while some may be exerted by direct binding to as yet uncharacterized Dfd response elements. We also show that ectopic CncB is sufficient to transform ventral epidermis in the trunk into repetitive arrays of ventral pharynx. We compare the functions of CncB to those of its vertebrate and invertebrate homologs, p45 NF-E2, Nrf and Skn-1 proteins, and suggest that the pharynx selector function of CncB is highly conserved on some branches of the evolutionary tree.
Mesenchymal cells engulf and clear apoptotic footplate cells in macrophageless PU.1 null mouse embryos
Authors: Wood William , Turmaine Mark , Weber Roberta , Camp Victoria , Maki Richard A. , McKercher Scott R. , Martin Paul
Published: April 2021

Apoptosis is one of the key tools used by an embryo to regulate cell numbers and sculpt body shape. Although massive numbers of cells die during development, they are so rapidly phagocytosed that very few corpses are ever seen in most embryonic tissues. In this paper, we focus on the catastrophic cell death that occurs as the developing footplate is remodelled to transform webbed regions into free interdigital spaces. In the wild-type embryo, these dead cells are rapidly engulfed and cleared by macrophages. We show that in a macrophageless mouse embryo, null for the haemopoetic-lineage-specific transcription factor, PU.1, the task of phagocytosis is taken over by ‘stand-in’ mesenchymal neighbours in a clear example of cell redundancy. However, it takes three times as many of these mesenchymal phagocytes to complete the task and, at each stage of the clearance process – in the recognition of apoptotic debris, its engulfment and finally its digestion – they appear to be less efficient than macrophages. A molecular explanation for this may be that several of the engulfment genes expressed by macrophages, including the ABC1 transporter (believed to be part of the phagocytic machinery conserved from Caenorhabditis elegans to mouse), are not upregulated by these ‘stand-in’ phagocytes.
Transcription of meiotic cell cycle and terminal differentiation genes depends on a conserved chromatin associated protein, whose nuclear localisation is regulated
Authors: White-Cooper Helen , Leroy Dorothée , MacQueen Amy , Fuller Margaret T.
Published: April 2021

The Drosophila always early (aly) gene coordinately regulates meiotic cell cycle progression and terminal differentiation during male gametogenesis. aly is required for transcription of key G2-M cell cycle control genes and of spermatid differentiation genes, and for maintenance of normal chromatin structure in primary spermatocytes. We show that aly encodes a homologue of the Caenorhabditis elegans gene lin-9, a negative regulator of vulval development that acts in the same SynMuvB genetic pathway as the LIN-35 Rb-like protein. The aly gene family is conserved from plants to humans. Aly protein is both cytoplasmic and nuclear in early primary spermatocytes, then resolves to a chromatin-associated pattern. It remains cytoplasmic in a loss-of-function missense allele, suggesting that nuclear localisation is critical for Aly function, and that other factors may alter Aly activity by controlling its subcellular localisation. MAPK activation occurs normally in aly mutant testes. Therefore aly, and by inference lin-9, act in parallel to, or downstream of, activation of MAPK by the RTK-Ras signalling pathway. We favour a model where aly may regulate cell cycle progression and terminal differentiation during male gametogenesis by regulating chromatin conformation in primary spermatocytes.
Windbeutel is required for function and correct subcellular localization of the Drosophila patterning protein Pipe
Authors: Sen Jonaki , Goltz Jason S. , Konsolaki Mary , Schüpbach Trudi , Stein David
Published: April 2021

Drosophila embryonic dorsal-ventral polarity originates in the ovarian follicle through the restriction of pipe gene expression to a ventral subpopulation of follicle cells. Pipe, a homolog of vertebrate glycosaminoglycan-modifying enzymes, directs the ventral activation of an extracellular serine proteolytic cascade which defines the ventral side of the embryo. When pipe is expressed uniformly in the follicle cell layer, a strong ventralization of the resulting embryos is observed. Here, we show that this ventralization is dependent on the other members of the dorsal group of genes controlling dorsal-ventral polarity, but not on the state of the Epidermal Growth Factor Receptor signal transduction pathway which defines egg chamber polarity. Pipe protein expressed in vertebrate tissue culture cells localizes to the endoplasmic reticulum. Strikingly, coexpression of the dorsal group gene windbeutel in those cells directs Pipe to the Golgi. Similarly, Pipe protein exhibits an altered subcellular localization in the follicle cells of females mutant for windbeutel. Thus, Windbeutel protein enables the correct subcellular distribution of Pipe to facilitate its pattern-forming activity.
Sperm-induced calcium oscillations at fertilisation in ascidians are controlled by cyclin B1-dependent kinase activity
Authors: Levasseur Mark , McDougall Alex
Published: April 2021

The generation of calcium oscillations at fertilisation and during mitosis appears to be controlled by the cell cycle machinery. For example, the calcium oscillations in oocytes and embryos occur during metaphase and terminate upon entry into interphase. Here we report the manipulation of sperm-triggered calcium oscillations by cyclin-dependent kinase (CDK) activity, the major component of maturation/ M phase promoting factor (MPF). To control the CDK activity we microinjected mRNAs encoding full-length GFP-tagged cyclin B1 or a truncated and therefore stabilised form of cyclin B1 (Δ90) into unfertilised oocytes. In the presence of full-length cyclin B1, the calcium oscillations terminate when cyclin B1 levels fall along with the concomitant fall in the associated CDK activity. In addition, when the CDK activity is elevated indefinitely with Δ90 cyclin B1, the calcium oscillations also continue indefinitely. Finally, in oocytes that contain low mitogen- activated protein (MAP) kinase activity and elevated CDK activity, the sperm-triggered calcium oscillations are again prolonged. We conclude that the CDK activity of the ascidian oocyte can be regarded as a positive regulator of sperm-triggered calcium oscillations, a finding that may apply to other oocytes that display sperm-triggered calcium oscillations at fertilisation. Furthermore, these findings may have a bearing upon the mitotic calcium signals of early embryos.
Organ shape in the Drosophila salivary gland is controlled by regulated, sequential internalization of the primordia
Authors: Myat Monn Monn , Andrew Deborah J.
Published: April 2021

During Drosophila development, the salivary primordia are internalized to form the salivary gland tubes. By analyzing immuno-stained histological sections and scanning electron micrographs of multiple stages of salivary gland development, we show that internalization occurs in a defined series of steps, involves coordinated cell shape changes and begins with the dorsal-posterior cells of the primordia. The ordered pattern of internalization is critical for the final shape of the salivary gland. In embryos mutant for hückebein (hkb), which encodes a transcription factor, or faint sausage (fas), which encodes a cell adhesion molecule, internalization begins in the center of the primordia, and completely aberrant tubes are formed. The sequential expression of hkb in selected cells of the primordia presages the sequence of cell movements. We propose that hkb dictates the initial site of internalization, the order in which invagination progresses and, consequently, the final shape of the organ. We propose that fas is required for hkb-dependent signaling events that coordinate internalization.
Redundant regulation of meristem identity and plant architecture by FRUITFULL, APETALA1 and CAULIFLOWER
Authors: Ferrándiz Cristina , Gu Qing , Martienssen Robert , Yanofsky Martin F.
Published: April 2021

The transition from vegetative to reproductive phases during Arabidopsis development is the result of a complex interaction of environmental and endogenous factors. One of the key regulators of this transition is LEAFY (LFY), whose threshold levels of activity are proposed to mediate the initiation of flowers. The closely related APETALA1 (AP1) and CAULIFLOWER (CAL) meristem identity genes are also important for flower initiation, in part because of their roles in upregulating LFY expression. We have found that mutations in the FRUITFULL (FUL) MADS-box gene, when combined with mutations in AP1 and CAL, lead to a dramatic non-flowering phenotype in which plants continuously elaborate leafy shoots in place of flowers. We demonstrate that this phenotype is caused both by the lack of LFY upregulation and by the ectopic expression of the TERMINAL FLOWER1 (TFL1) gene. Our results suggest that the FUL, AP1 and CAL genes act redundantly to control inflorescence architecture by affecting the domains of LFY and TFL1 expression as well as the relative levels of their activities.
Formation and specification of distal leg segments in Drosophila by dual Bar homeobox genes, BarH1 and BarH2
Authors: Kojima Tetsuya , Sato Makoto , Saigo Kaoru
Published: April 2021

Here, we show that BarH1 and BarH2, a pair of Bar homeobox genes, play essential roles in the formation and specification of the distal leg segments of Drosophila. In early third instar, juxtaposition of Bar-positive and Bar-negative tissues causes central folding that may separate future tarsal segments 2 from 3, while juxtaposition of tissues differentially expressing Bar homeobox genes at later stages gives rise to segmental boundaries of distal tarsi including the tarsus/pretarsus boundary. Tarsus/pretarsus boundary formation requires at least two different Bar functions, early antagonistic interactions with a pretarsus-specific homeobox gene, aristaless, and the subsequent induction of Fas II expression in pretarsus cells abutting tarsal segment 5. Bar homeobox genes are also required for specification of distal tarsi. Bar expression requires Distal-less but not dachshund, while early circular dachshund expression is delimited interiorly by BarH1 and BarH2.
Endoderm patterning by the notochord: development of the hypochord in Xenopus
Authors: Cleaver Ondine , Seufert Daniel W. , Krieg Paul A.
Published: April 2021

The patterning and differentiation of the vertebrate endoderm requires signaling from adjacent tissues. In this report, we demonstrate that signals from the notochord are critical for the development of the hypochord, which is a transient, endodermally derived structure that lies immediately ventral to the notochord in the amphibian and fish embryo. It appears likely that the hypochord is required for the formation of the dorsal aorta in these organisms. We show that removal of the notochord during early neurulation leads to the complete failure of hypochord development and to the elimination of expression of the hypochord marker, VEGF. Removal of the notochord during late neurulation, however, does not interfere with hypochord formation. These results suggest that signals arising in the notochord instruct cells in the underlying endoderm to take on a hypochord fate during early neural stages, and that the hypochord does not depend on further notochord signals for maintenance. In reciprocal experiments, when the endoderm receives excess notochord signaling, a significantly enlarged hypochord develops. Overall, these results demonstrate that, in addition to patterning neural and mesodermal tissues, the notochord plays an important role in patterning of the endoderm.
BMP signaling is essential for development of skeletogenic and neurogenic cranial neural crest
Authors: Kanzler Benoît , Foreman Ruth K. , Labosky Patricia A. , Mallo Moisés
Published: April 2021

BMP signaling is essential for a wide variety of developmental processes. To evaluate the role of Bmp2/4 in cranial neural crest (CNC) formation or differentiation after its migration into the branchial arches, we used Xnoggin to block their activities in specific areas of the CNC in transgenic mice. This resulted in depletion of CNC cells from the targeted areas. As a consequence, the branchial arches normally populated by the affected neural crest cells were hypomorphic and their skeletal and neural derivatives failed to develop. In further analyses, we have identified Bmp2 as the factor required for production of migratory cranial neural crest. Its spatial and temporal expression patterns mirror CNC emergence and Bmp2 mutant embryos lack both branchial arches and detectable migratory CNC cells. Our results provide functional evidence for an essential role of BMP signaling in CNC development.
A novel neuropeptide, Hym-355, positively regulates neuron differentiation in Hydra
Authors: Takahashi Toshio , Koizumi Osamu , Ariura Yuki , Romanovitch Anna , Bosch Thomas C. G. , Kobayakawa Yoshitaka , Mohri Shiro , Bode Hans R. , Yum Seungshic , Hatta Masayuki , Fujisawa Toshitaka
Published: April 2021

During the course of a systematic screening of peptide signaling molecules in Hydra a novel peptide, Hym-355 (FPQSFLPRG-NH2), was identified. A cDNA encoding the peptide was isolated and characterized. Using both in situ hybridization and immunohistochemistry, Hym-355 was shown to be expressed in neurons and hence is a neuropeptide. The peptide was shown to specifically enhance neuron differentiation throughout the animal by inducing interstitial cells to enter the neuron pathway. Further, co-treatment with a PW peptide, which inhibits neuron differentiation, nullified the effects of both peptides, suggesting that they act in an antagonistic manner. This effect is discussed in terms of a feedback mechanism for maintaining the steady state neuron population in Hydra.
Autoregulation and multiple enhancers control Math1 expression in the developing nervous system
Authors: Helms Amy W. , Abney Andrew L. , Ben-Arie Nissim , Zoghbi Huda Y. , Johnson Jane E.
Published: April 2021

Development of the vertebrate nervous system requires the actions of transcription factors that establish regional domains of gene expression, which results in the generation of diverse neuronal cell types. MATH1, a transcription factor of the bHLH class, is expressed during development of the nervous system in multiple neuronal domains, including the dorsal neural tube, the EGL of the cerebellum and the hair cells of the vestibular and auditory systems. MATH1 is essential for proper development of the granular layer of the cerebellum and the hair cells of the cochlear and vestibular systems, as shown in mice carrying a targeted disruption of Math1. Previously, we showed that 21 kb of sequence flanking the Math1-coding region is sufficient for Math1 expression in transgenic mice. Here we identify two discrete sequences within the 21 kb region that are conserved between mouse and human, and are sufficient for driving a lacZ reporter gene in these domains of Math1 expression in transgenic mice. The two identified enhancers, while dissimilar in sequence, appear to have redundant activities in the different Math1 expression domains except the spinal neural tube. The regulatory mechanisms for each of the diverse Math1 expression domains are tightly linked, as separable regulatory elements for any given domain of Math1 expression were not found, suggesting that a common regulatory mechanism controls these apparently unrelated domains of expression. In addition, we demonstrate a role for autoregulation in controlling the activity of the Math1 enhancer, through an essential E-box consensus binding site.
Different clonal dispersion in the rostral and caudal mouse central nervous system
Authors: Mathis Luc , François Nicolas Jean
Published: April 2021

We have performed a systematic clonal analysis to describe the modes of growth, dispersion and production of cells during the development of the mouse neural system. We have used mice expressing a LaacZ reporter gene under the control of the neuron specific enolase promoter to randomly generate LacZ clones in the central nervous system (CNS). We present evidence for (1) a pool of CNS founder cells that is not regionalized, i.e. give descendants dispersed along the entire A-P axis, (2) an early separation between pools of precursors for the anterior and posterior CNS and (3) distinct modes of production of progenitors in these two domains. More specifically, cell growth and dispersion of the progenitors follow a relatively coherent pattern throughout the anterior CNS, a mode that leads to a progressive regionalization of cell fates. In contrast, cell growth of progenitors of the SC appears to involve self-renewing stem cells that progress caudally during regression of the mode. Therefore, at least part of the area surrounding the node is composed of precursors with self-renewing properties and the development of the trunk is dependent on pools of stem cells regressing from A to P. Taken together with our analysis of the cell growth changes associated with neuromere formation (Mathis, L., Sieur, J., Voiculescu, O., Charnay, P. and Nicolas, J. F. (1999) Development 126, 4095-4106), our results suggest that major transitions in CNS development correspond to changes in cell behavior and may provide a link between morphogenesis and genetic patterning mechanisms (i.e. formation of the body plan).
Netrin 1 acts as an attractive or as a repulsive cue for distinct migrating neurons during the development of the cerebellar system
Authors: Alcántara S. , Ruiz M. , Castro F. De , Soriano E. , Sotelo C.
Published: April 2021

Netrin 1 is a long-range diffusible factor that exerts chemoattractive or chemorepulsive effects on developing axons growing to or away from the neural midline. Here we used tissue explants to study the action of netrin 1 in the migration of several cerebellar and precerebellar cell progenitors. We show that netrin 1 exerts a strong chemoattractive effect on migrating neurons from the embryonic lower rhombic lip at E12-E14, which give rise to precerebellar nuclei. Netrin 1 promotes the exit of postmitotic migrating neurons from the embryonic lower rhombic lip and upregulates the expression of TAG-1 in these neurons. In addition, in the presence of netrin 1, the migrating neurons are not isolated but are associated with thick fascicles of neurites, typical of the neurophilic way of migration. In contrast, the embryonic upper rhombic lip, which contains tangentially migrating granule cell progenitors, did not respond to netrin 1. Finally, in the postnatal cerebellum, netrin 1 repels both the parallel fibres and migrating granule cells growing out from explants taken from the external germinal layer. The developmental patterns of expression in vivo of netrin 1 and its receptors are consistent with the notion that netrin 1 secreted in the midline acts as chemoattractive cue for precerebellar neurons migrating circumferentially along the extramural stream. Similarly, the pattern of expression in the postnatal cerebellum suggests that netrin 1 could regulate the tangential migration of postmitotic premigratory granule cells. Thus, molecular mechanisms considered as primarily involved in axonal guidance appear also to steer neuronal cell migration.
Expression of achaete-scute homologues in discrete proneural clusters on the developing notum of the medfly Ceratitis capitata, suggests a common origin for the stereotyped bristle patterns of higher Diptera
Authors: Wülbeck Corinna , Simpson Pat
Published: April 2021

The stereotyped positioning of sensory bristles in Drosophila has been shown to result from complex spatiotemporal regulation of the proneural achaete-scute genes, that relies on an array of cis-regulatory elements and spatially restricted transcriptional activators such as Pannier. Other species of derived schizophoran Diptera have equally stereotyped, but different, bristle patterns. Divergence of bristle patterns could arise from changes in the expression pattern of proneural genes, resulting from evolution of the cis-regulatory sequences and/or altered expression patterns of transcriptional regulators. Here we describe the isolation of achaete-scute homologues in Ceratitis capitata, a species of acalyptrate Schizophora whose bristle pattern differs slightly from that of Drosophila. At least three genes, scute, lethal of scute and asense have been conserved, thus demonstrating that gene duplication within the achaete-scute complex preceded the separation of the families Drosophilidae and Tephritidae, whose common ancestor goes back more than 100 million years. The expression patterns of these genes provide evidence for conservation of many cis-regulatory elements as well as a common origin for the stereotyped patterns seen on the scutum of many Schizophora. Some aspects of the transcriptional regulation have changed, however, and correlate in the notum with differences in the bristle pattern. The Ceratitis pannier gene was isolated and displays a conserved expression domain in the notum.
A new function of BMP4: dual role for BMP4 in regulation ofSonic hedgehogexpression in the mouse tooth germ
Authors: Zhang Yanding , Zhang Zunyi , Zhao Xiang , Yu Xueyan , Hu Yueping , Geronimo Benedicto , Fromm Sigurd H. , Chen YiPing
Published: April 2021

The murine tooth development is governed by sequential and reciprocal epithelial-mesenchymal interactions. Multiple signaling molecules are expressed in the developing tooth germ and interact each other to mediate the inductive tissue interactions. Among them are Sonic hedgehog (SHH), Bone Morphogenetic Protein-2 (BMP2) and Bone Morphogenetic Protein-4 (BMP4). We have investigated the interactions between these signaling molecules during early tooth development. We found that the expression of Shh and Bmp2 is downregulated at E12.5 and E13.5 in the dental epithelium of the Msx1 mutant tooth germ where Bmp4 expression is significantly reduced in the dental mesenchyme. Inhibition of BMP4 activity by noggin resulted in repression of Shh and Bmp2 in wild-type dental epithelium. When implanted into the dental mesenchyme of Msx1 mutants, beads soaked with BMP4 protein were able to restore the expression of both Shh and Bmp2 in the Msx1 mutant epithelium. These results demonstrated that mesenchymal BMP4 represents one component of the signal acting on the epithelium to maintain Shh and Bmp2 expression. In contrast, BMP4-soaked beads repressed Shh and Bmp2 expression in the wild-type dental epithelium. TUNEL assay indicated that this suppression of gene expression by exogenous BMP4 was not the result of an increase in programmed cell death in the tooth germ. Ectopic expression of human Bmp4 to the dental mesenchyme driven by the mouse Msx1 promoter restored Shh expression in the Msx1 mutant dental epithelium but repressed Shh in the wild-type tooth germ in vivo. We further demonstrated that this regulation of Shh expression by BMP4 is conserved in the mouse developing limb bud. In addition, Shh expression was unaffected in the developing limb buds of the transgenic mice in which a constitutively active Bmpr-IB is ectopically expressed in the forelimb posterior mesenchyme and throughout the hindlimb mesenchyme, suggesting that the repression of Shh expression by BMP4 may not be mediated by BMP receptor-IB. These results provide evidence for a new function of BMP4. BMP4 can act upstream to Shh by regulating Shh expression in mouse developing tooth germ and limb bud. Taken together, our data provide insight into a new regulatory mechanism for Shh expression, and suggest that this BMP4-mediated pathway in Shh regulation may have a general implication in vertebrate organogenesis.
The C. elegans par-4 gene encodes a putative serine-threonine kinase required for establishing embryonic asymmetry
Authors: Watts Jennifer L. , Morton Diane G. , Bestman Jennifer , Kemphues Kenneth J.
Published: April 2021

During the first cell cycle of Caenorhabditis elegans embryogenesis, asymmetries are established that are essential for determining the subsequent developmental fates of the daughter cells. The maternally expressed par genes are required for establishing this polarity. The products of several of the par genes have been found to be themselves asymmetrically distributed in the first cell cycle. We have identified the par-4 gene of C. elegans, and find that it encodes a putative serine-threonine kinase with similarity to a human kinase associated with Peutz-Jeghers Syndrome, LKB1 (STK11), and a Xenopus egg and embryo kinase, XEEK1. Several strong par-4 mutant alleles are missense mutations that alter conserved residues within the kinase domain, suggesting that kinase activity is essential for PAR-4 function. We find that the PAR-4 protein is present in the gonads, oocytes and early embryos of C. elegans, and is both cytoplasmically and cortically distributed. The cortical distribution begins at the late 1- cell stage, is more pronounced at the 2- and 4-cell stages and is reduced at late stages of embryonic development. We find no asymmetry in the distribution of PAR-4 protein in C. elegans embryos. The distribution of PAR-4 protein in early embryos is unaffected by mutations in the other par genes.
Depolarisation causes reciprocal changes in GFRα-1 and GFRα-2 receptor expression and shifts responsiveness to GDNF and neurturin in developing neurons
Authors: Doxakis Epaminondas , Wyatt Sean , Davies Alun M.
Published: April 2021

GDNF and neurturin are structurally related neurotrophic factors that promote the survival of many different kinds of neurons and influence axonal and dendritic growth and synaptic function. These diverse effects are mediated via multicomponent receptors consisting of the Ret receptor tyrosine kinase plus one of two structurally related GPI- linked receptors, GFRα-1 and GFRα-2. To ascertain how the expression of these receptors is regulated during development, we cultured embryonic neurons under different experimental conditions and used competitive RT/PCR to measure the levels of the mRNAs encoding these receptors. We found that depolarising levels of KCl caused a marked increase in GFRα-1 mRNA and a marked decrease in GFRα-2 mRNA in sympathetic, parasympathetic and sensory neurons. These changes were accompanied by increased responsiveness to GDNF and decreased responsiveness to neurturin, and were inhibited by L-type Ca2+ channel antagonists, suggesting that they were due to elevated intracellular free-Ca2+. There was no consistent effect of depolarising levels of KCl on ret mRNA expression, and neither GDNF nor neurturin significantly affected receptor expression. These results show that depolarisation has marked and opposing actions on the expression of GFRα-1 and GFRα-2, which are translated into corresponding changes in neuronal responsiveness to GDNF and neurturin. This provides evidence for a mechanism of regulating the neurotrophic factor responses of neurons by neural activity that has important implications for structural and functional plasticity in the developing nervous system.
Local, possibly contact-mediated signalling restricted to homotypic neurons controls the regular spacing of cells within the cholinergic arrays in the developing rodent retina
Authors: Galli-Resta Lucia
Published: April 2021

In the vertebrate retina neurons of the same type commonly form non-random arrays, assembled by unknown positional mechanisms during development. Computational models in which no two cells are closer than a minimal distance, simulate many retinal arrays. These findings have important biological implications, since they suggest that cells are determined as neurons of specific types before entering their arrays, and that local, possibly contact-mediated interactions acting exclusively among the elements of an array account for its assembly. This is here verified by combining experimental manipulations in normal and transgenic models with computational analysis for the cholinergic mosaics, the only arrays so far for which the development of spatial ordering is known quantitatively. When generalised, these findings suggest a plan for vertebrate retinal patterning, where homotypic interactions organise retinal arrays first, then local interactions between synaptic partners suffice to establish the topographical connections that support retinal processing.
Persistence of Hunchback in the terminal region of the Drosophila blastoderm embryo impairs anterior development
Authors: Janody Florence , Reischl Joachim , Dostatni Nathalie
Published: April 2021

Anterior terminal development is controlled by several zygotic genes that are positively regulated at the anterior pole of Drosophila blastoderm embryos by the anterior (bicoid) and the terminal (torso) maternal determinants. Most Bicoid target genes, however, are first expressed at syncitial blastoderm as anterior caps, which retract from the anterior pole upon activation of Torso. To better understand the interaction between Bicoid and Torso, a derivative of the Gal4/UAS system was used to selectively express the best characterised Bicoid target gene, hunchback, at the anterior pole when its expression should be repressed by Torso. Persistence of hunchback at the pole mimics most of the torso phenotype and leads to repression at early stages of a labral (cap’n’collar) and two foregut (wingless and hedgehog) determinants that are positively controlled by bicoid and torso. These results uncovered an antagonism between hunchback and bicoid at the anterior pole, whereas the two genes are known to act in concert for most anterior segmented development. They suggest that the repression of hunchback by torso is required to prevent this antagonism and to promote anterior terminal development, depending mostly on bicoid activity.
Fate of the mammalian cardiac neural crest
Authors: Jiang Xiaobing , Rowitch David H. , Soriano Philippe , McMahon Andrew P. , Sucov Henry M.
Published: April 2021

A subpopulation of neural crest termed the cardiac neural crest is required in avian embryos to initiate reorganization of the outflow tract of the developing cardiovascular system. In mammalian embryos, it has not been previously experimentally possible to study the long-term fate of this population, although there is strong inference that a similar population exists and is perturbed in a number of genetic and teratogenic contexts. We have employed a two-component genetic system based on Cre/lox recombination to label indelibly the entire mouse neural crest population at the time of its formation, and to detect it at any time thereafter. Labeled cells are detected throughout gestation and in postnatal stages in major tissues that are known or predicted to be derived from neural crest. Labeling is highly specific and highly efficient. In the region of the heart, neural-crest-derived cells surround the pharyngeal arch arteries from the time of their formation and undergo an altered distribution coincident with the reorganization of these vessels. Labeled cells populate the aorticopulmonary septum and conotruncal cushions prior to and during overt septation of the outflow tract, and surround the thymus and thyroid as these organs form. Neural-crest-derived mesenchymal cells are abundantly distributed in midgestation (E9.5-12.5), and adult derivatives of the third, fourth and sixth pharyngeal arch arteries retain a substantial contribution of labeled cells. However, the population of neural-crest-derived cells that infiltrates the conotruncus and which surrounds the noncardiac pharyngeal organs is either overgrown or selectively eliminated as development proceeds, resulting for these tissues in a modest to marginal contribution in late fetal and postnatal life.