bluuebunny/crossref_metadata_2025 huggingface dataset:
Temporally-regulated retinoic acid depletion produces specific neural crest, ocular and nervous system defects
Authors: Dickman Eileen D. , Thaller Christina , Smith Susan M.
Published: April 2021

Both retinoid receptor null mutants and classic nutritional deficiency studies have demonstrated that retinoids are essential for the normal development of diverse embryonic structures (e.g. eye, heart, nervous system, urogenital tract). Detailed analysis of retinoid-modulated events is hampered by several limitations of these models, including that deficiency or null mutation is present throughout gestation, making it difficult to isolate primary effects, and preventing analysis beyond embryolethality. We developed a mammalian model in which retinoid-dependent events are documented during distinct targeted windows of embryogenesis. This was accomplished through the production of vitamin A-depleted (VAD) female rats maintained on sufficient oral retinoic acid (RA) for growth and fertility. After mating to normal males, these RA-sufficient/VAD females were given oral RA doses which allowed for gestation in an RA-sufficient state; embryogenesis proceeded normally until retinoids were withdrawn dietarily to produce a sudden, acute retinoid deficiency during a selected gestational window. In this trial, final RA doses were administered on E11.5, vehicle at E12.5, and embryos analyzed on E13.5; during this 48 hour window, the last RA dose was metabolized and embryos progressed in a retinoid-deficient state. RA-sufficient embryos were normal. Retinoiddepleted embryos exhibited specific malformations of the face, neural crest, eyes, heart, and nervous system. Some defects were phenocopies of those seen in null mutant mice for RXRα−/−, RXRα−/−/RARα−/−, and RARα−/−/RARγ−/−, confirming that RA transactivation of its nuclear receptors is essential for normal embryogenesis. Other defects were unique to this deficiency model, showing that complete ligand ‘knock-out’ is required to see those retinoiddependent events previously concealed by receptor functional redundancy, and reinforcing that retinoid receptors have separate yet overlapping contributions in the embryo. This model allows for precise targeting of retinoid form and deficiency to specific developmental windows, and will facilitate studies of distinct temporal events.
HOXD4 and regulation of the group 4 paralog genes
Authors: Morrison Alastair , Ariza-McNaughton Linda , Gould Alex , Featherstone Mark , Krumlauf Robb
Published: April 2021

From an evolutionary perspective, it is important to understand the degree of conservation of cis-regulatory mechanisms between paralogous Hox genes. In this study, we have used transgenic analysis of the human HOXD4 locus to identify one neural and two mesodermal 3′ enhancers that are capable of mediating the proper anterior limits of expression in the hindbrain and paraxial mesoderm (somites), respectively. In addition to directing expression in the central nervous system (CNS) up to the correct rhombomere 6/7 boundary in the hindbrain, the neural enhancer also mediates a three rhombomere anterior shift from this boundary in response to retinoic acid (RA), mimicking the endogenous Hoxd4 response. We have extended the transgenic analysis to Hoxa4 identifying mesodermal, neural and retinoid responsive components in the 3′ flanking region of that gene, which reflect aspects of endogenous Hoxa4 expression. Comparative analysis of the retinoid responses of Hoxd4, Hoxa4 and Hoxb4 reveals that, while they can be rapidly induced by RA, there is a window of competence for this response, which is different to that of more 3′ Hox genes. Mesodermal regulation involves multiple regions with overlapping or related activity and is complex, but with respect to neural regulation and response to RA, Hoxb4 and Hoxd4 appear to be more closely related to each other than Hoxa4. These results illustrate that much of the general positioning of 5′ and 3′ flanking regulatory regions has been conserved between three of the group 4 paralogs during vertebrate evolution, which most likely reflects the original positioning of regulatory regions in the ancestral Hox complex.
Medaka spalt acts as a target gene of hedgehog signaling
Authors: Köster Reinhard , Stick Reimer , Loosli Felix , Wittbrodt Joachim
Published: April 2021

In vertebrates, pattern formation in the eye, central nervous system, somites, and limb depends on hedgehog activity, but a general target gene controlled by hedgehog in all these signaling centers has remained largely elusive. The medaka fish gene spalt encodes a zinc-finger transcription factor, which is expressed in all known hedgehog signaling centers of the embryo and in the organizer region at the midbrain-hindbrain boundary. We show that the spalt expression domains expand in response to ectopic hedgehog activity and narrow in the presence of protein kinase A activity, an antagonist of hedgehog signaling, indicating that spalt is a hedgehog target gene. Our results also suggest a signaling mechanism for anterior-posterior patterning of the vertebrate brain that controls spalt expression at the midbrain-hindbrain boundary in a protein kinase A dependent manner likely to involve an unknown member of the hedgehog family.
Zebrafish vasa homologue RNA is localized to the cleavage planes of 2- and 4-cell-stage embryos and is expressed in the primordial germ cells
Authors: Yoon Christina , Kawakami Koichi , Hopkins Nancy
Published: April 2021

Identification and manipulation of the germ line are important to the study of model organisms. Although zebrafish has recently emerged as a model for vertebrate development, the primordial germ cells (PGCs) in this organism have not been previously described. To identify a molecular marker for the zebrafish PGCs, we cloned the zebrafish homologue of the Drosophila vasa gene, which, in the fly, encodes a germ-cell-specific protein. Northern blotting revealed that zebrafish vasa homologue (vas) transcript is present in embryos just after fertilization, and hence it is probably maternally supplied. Using wholemount in situ hybridization, we investigated the expression pattern of vas RNA in zebrafish embryos from the 1-cell stage to 10 days of development. Here we present evidence that vas RNA is a germ-cell-specific marker, allowing a description of the zebrafish PGCs for the first time. Furthermore, vas transcript was detected in a novel pattern, localized to the cleavage planes in 2- and 4-cell-stage embryos. During subsequent cleavages, the RNA is segregated as subcellular clumps to a small number of cells that may be the future germ cells. These results suggest new ways in which one might develop techniques for the genetic manipulation of zebrafish. Furthermore, they provide the basis for further studies on this novel RNA localization pattern and on germ-line development in general.
mago nashi mediates the posterior follicle cell-to-oocyte signal to organize axis formation in Drosophila
Authors: Newmark Phillip A. , Mohr Stephanie E. , Gong Lei , Boswell Robert E.
Published: April 2021

Establishment of the anteroposterior and dorsoventral axes in the Drosophila egg chamber requires reciprocal signaling between the germ line and soma. Upon activation of the Drosophila EGF receptor in the posterior follicle cells, these cells signal back to the oocyte, resulting in a reorganization of the oocyte cytoplasm and anterodorsal migration of the oocyte nucleus. We demonstrate that the gene mago nashi (mago) encodes an evolutionarily conserved protein that must be localized within the posterior pole plasm for germ-plasm assembly and Caenorhabditis elegans mago is a functional homologue of Drosophila mago. In the absence of mago+ function during oogenesis, the anteroposterior and dorsoventral coordi-nates of the oocyte are not specified and the germ plasm fails to assemble.
The Drosophila gene morula inhibits mitotic functions in the endo cell cycle and the mitotic cell cycle
Authors: Reed Bruce H. , Orr-Weaver Terry L.
Published: April 2021

In the endo cell cycle, rounds of DNA replication occur in the absence of mitosis, giving rise to polyploid or polytene cells. We show that the Drosophila morula gene is essential to maintain the absence of mitosis during the endo cycle. During oogenesis in wild-type Drosophila, nurse cells become polyploid and do not contain cyclin B protein. Nurse cells in female-sterile alleles of morula begin to become polyploid but revert to a mitotic-like state, condensing the chromosomes and forming spindles. In strong, larval lethal alleles of morula, the polytene ring gland cells also inappropriately regress into mitosis and form spindles. In addition to its role in the endo cycle, morula function is necessary for dividing cells to exit mitosis. Embryonic S-M cycles and the archetypal (G1-S-G2-M) cell cycle are both arrested in metaphase in different morula mutants. These phenotypes suggest that morula acts to block mitosis-promoting activity in both the endo cycle and at the metaphase/anaphase transition of the mitotic cycle. Consistent with this, we found cyclin B protein to be inappropriately present in morula mutant nurse cells. Thus morula serves a dual function as a cell cycle regulator that promotes exit from mitosis and maintains the absence of mitosis during the endo cycle, possibly by activating the cyclin destruction machinery.
Differential regulation of gastrulation and neuroectodermal gene expression by Snail in the Drosophila embryo
Authors: Hemavathy Kirugaval , Meng Xiangjun , Ip Y. Tony
Published: April 2021

The initiation of mesoderm differentiation in the Drosophila embryo requires the gene products of twist and snail. In either mutant, the ventral cell invagination during gastrulation is blocked and no mesoderm-derived tissue is formed. One of the functions of Snail is to repress neuroectodermal genes and restrict their expressions to the lateral regions. The derepression of the neuroectodermal genes into the ventral region in snail mutant is a possible cause of defects in gastrulation and in mesoderm differentiation. To investigate such possibility, we analysed a series of snail mutant alleles. We found that different neuroectodermal genes respond differently in various snail mutant background. Due to the differential response of target genes, one of the mutant alleles, V2, that has reduced Snail function showed an intermediate phenotype. In V2 embryos, neuroectodermal genes, such as single-minded and rhomboid, are derepressed while ventral invagination proceeds normally. However, the differentiation of these invaginated cells into mesodermal lineage is disrupted. The results suggest that the establishment of mesodermal cell fate requires the proper restriction of neuroectodermal genes, while the ventral cell movement is independent of the expression patterns of these genes. Together with the data showing that the expression of some ventral genes disappear in snail mutants, we propose that Snail may repress or activate another set of target genes that are required specifically for gastrulation.
Wingless signaling generates pattern through two distinct mechanisms
Authors: Hays Rebecca , Gibori Gil B. , Bejsovec Amy
Published: April 2021

wingless (wg) and its vertebrate homologues, the Wnt genes, play critical roles in the generation of embryonic pattern. In the developing Drosophila epidermis, wg is expressed in a single row of cells in each segment, but it influences cell identities in all rows of epidermal cells in the 10- to 12-cell-wide segment. Wg signaling promotes specification of two distinct aspects of the wild-type intrasegmental pattern: the diversity of denticle types present in the anterior denticle belt and the smooth or naked cuticle constituting the posterior surface of the segment. We have manipulated the expression of wild-type and mutant wg transgenes to explore the mechanism by which a single secreted signaling molecule can promote these distinctly different cell fates.

We present evidence consistent with the idea that naked cuticle cell fate is specified by a cellular pathway distinct from the denticle diversity-generating pathway. Since these pathways are differentially activated by mutant Wg ligands, we propose that at least two discrete classes of receptor for Wg may exist, each transducing a different cellular response. We also find that broad Wg protein distribution across many cell diameters is required for the generation of denticle diversity, suggesting that intercellular transport of the Wg protein is an essential feature of pattern formation within the epidermal epithelium. Finally, we demonstrate that an 85 amino acid region not conserved in vertebrate Wnts is dispensable for Wg function and we discuss structural features of the Wingless protein required for its distinct biological activities.
Integration of the head and trunk segmentation systems controls cephalic furrow formation in Drosophila
Authors: Vincent Alain , Blankenship J. Todd , Wieschaus Eric
Published: April 2021

Genetic and molecular analyses of patterning of the Drosophila embryo have shown that the process of segmentation of the head is fundamentally different from the process of segmentation of the trunk. The cephalic furrow (CF), one of the first morphological manifestations of the patterning process, forms at the juxtaposition of these two patterning systems. We report here that the initial step in CF formation is a change in shape and apical positioning of a single row of cells. The anteroposterior position of these initiator cells may be defined by the overlapping expression of the head gap gene buttonhead (btd) and the primary pairrule gene even-skipped (eve). Re-examination of the btd and eve phenotypes in live embryos indicated that both genes are required for CF formation. Further, Eve expression in initiator cells was found to be dependent upon btd activity. The control of eve expression by btd in these cells is the first indication of a new level of integrated regulation that interfaces the head and trunk segmentation systems. In conjunction with previous data on the btd and eve embryonic phenotypes, our results suggest that interaction between these two genes both controls initiation of a specific morphogenetic movement that separates two morphogenetic fields and contributes to patterning the hinge region that demarcates the procephalon from the segmented germ band.
Pax6-dependent regulation of adhesive patterning, R-cadherin expression and boundary formation in developing forebrain
Authors: Stoykova A. , Götz M. , Gruss P. , Price J.
Published: April 2021

Mutations in the gene for the transcription factor, Pax6, induce marked developmental abnormalities in the CNS and the eye, but the cellular mechanisms that underlie the phenotype are unknown. We have examined the adhesive properties of cells from the developing forebrain in Small eye, the Pax6 mutant mouse. We have found that the seg-regation normally observed in aggregates of cortical and striatal cells in an in vitro assay is lost in Small eye. This correlates with an alteration of in vivo expression of the homophilic adhesion molecule, R-cadherin. Moreover, the boundary between cortical and striatal regions of the telencephalon is dramatically altered in Small eye: radial glial fascicles do not form at the border, and the normal expression of R-cadherin and tenascin-C at the border is lost. These data suggest a link between the transcription factor, Pax6, R-cadherin expression, cellular adhesion and boundary formation between developing forebrain regions.
Cell morphogenesis of trichomes in Arabidopsis: differential control of primary and secondary branching by branch initiation regulators and cell growth
Authors: Folkers Ulrike , Berger Jürgen , Hülskamp Martin
Published: April 2021

Cell morphogenesis, i.e. the acquisition of a particular cell shape, can be examined genetically in the three-branched trichomes that differentiate from single epidermal cells on the leaves of Arabidopsis thaliana. In normal development, the growing trichome cell undergoes two successive branching events, resulting in a proximal side stem and a distal main stem which subsequently splits in two branches. Using new and previously described trichome mutants, we have analyzed the branching pattern in single and double mutants affecting branch number or cell size in order to determine underlying mechanisms. Our results suggest that primary branching is genetically distinct from subsequent branching events and that the latter, secondary events are initiated in response to positive and negative regulators of branching as well as subject to control by cell growth. We propose a model of how trichome cell morphogenesis is regulated during normal development.
Suppression of GATA factor activity causes axis duplication in Xenopus
Authors: Sykes T. G. , Rodaway A. R. F. , Walmsley M. E. , Patient R. K.
Published: April 2021

In Xenopus, the dorsoventral axis is patterned by the interplay between active signalling in ventral territories, and secreted antagonists from Spemann’s organiser. Two signals are important in ventral cells, bone morphogenetic protein-4 (BMP-4) and Wnt-8. BMP-4 plays a conserved role in patterning the vertebrate dorsoventral axis, whilst the precise role of Wnt-8 and its relationship with BMP-4, are still unclear. Here we have investigated the role played by the GATA family of transcription factors, which are expressed in ventral mesendoderm during gastrulation and are required for the differentiation of blood and endodermal tissues. Injection ventrally of a dominantinterfering GATA factor (called G2en) induced the formation of secondary axes that phenocopy those induced by the dominant-negative BMP receptor. However, unlike inhibiting BMP signalling, inhibiting GATA activity in the ectoderm does not lead to neuralisation. In addition,analysis of gene expression in G2en injected embryos reveals that at least one known target gene for BMP-4, the homeobox gene Vent-2, is unaffected. In contrast, the expression of Wnt-8 and the homeobox gene Vent-1 is suppressed by G2en, whilst the organiser-secreted BMP antagonist chordin becomes ectopically expressed. These data therefore suggest that GATA activity is essential for ventral cell fate and that subsets of ventralising and dorsalising genes require GATA activity for their expression and suppression, respectively. Finally, using G2en, we show that suppression of Wnt-8 expression, in conjunction with blocked BMP signalling, does not lead to head formation, suggesting that the head-suppressing Wnt signal may not be Wnt-8.
Early development of the zebrafish pronephros and analysis of mutations affecting pronephric function
Authors: Drummond Iain A. , Majumdar Arindam , Hentschel Hartmut , Elger Marlies , Solnica-Krezel Lila , Schier Alexander F. , Neuhauss Stephan C. F. , Stemple Derek L. , Zwartkruis Fried , Rangini Zehava , Driever Wolfgang , Fishman Mark C.
Published: April 2021

The zebrafish pronephric kidney provides a simplified model of nephron development and epithelial cell differentiation which is amenable to genetic analysis. The pronephros consists of two nephrons with fused glomeruli and paired pronephric tubules and ducts. Nephron formation occurs after the differentiation of the pronephric duct with both the glomeruli and tubules being derived from a nephron primordium. Fluorescent dextran injection experiments demonstrate that vascularization of the zebrafish pronephros and the onset of glomerular filtration occurs between 40 and 48 hpf. We isolated fifteen recessive mutations that affect development of the pronephros. All have visible cysts in place of the pronephric tubule at 2-2.5 days of development. Mutants were grouped in three classes: (1) a group of twelve mutants with defects in body axis curvature and manifesting the most rapid and severe cyst formation involving the glomerulus, tubule and duct, (2) the fleer mutation with distended glomerular capillary loops and cystic tubules, and (3) the mutation pao pao tang with a normal glomerulus and cysts limited to the pronephric tubules. double bubble was analyzed as a representative of mutations that perturb the entire length of the pronephros and body axis curvature. Cyst formation begins in the glomerulus at 40 hpf at the time when glomerular filtration is established suggesting a defect associated with the onset of pronephric function. Basolateral membrane protein targeting in the pronephric duct epithelial cells is also severely affected, suggesting a failure in terminal epithelial cell differentiation and alterations in electrolyte transport. These studies reveal the similarity of normal pronephric development to kidney organogenesis in all vertebrates and allow for a genetic dissection of genes needed to establish the earliest renal function.
Differentiation: the selective potentiation of chromatin domains
Authors: Kramer Jeffrey A. , McCarrey John R. , Djakiew Daniel , Krawetz Stephen A.
Published: April 2021

Potentiation is requisite for the expression of our genome. It is the mechanism of opening chromatin domains to render genes accessible to tissue-specific and ubiquitous transacting-factors that enables transcription. The results presented in this study demonstrate that modulation of stage- and cell-type-specific gene expression during mammalian spermatogenesis involves selective potentiation of testis-expressed genes that reverses their repressive state when present in the spermatogonial stem cell. This directly contrasts hematopoiesis, which acts to selectively restrict lineage potential during differentiation from its permissive stem cell. These results are key to understanding how differentiative pathways are controlled and cellular phenotypes determined. A window to their modulation is presented.
Math5 encodes a murine basic helix-loop-helix transcription factor expressed during early stages of retinal neurogenesis
Authors: Brown Nadean L. , Kanekar Shami , Vetter Monica L. , Tucker Priscilla K. , Gemza Debra L. , Glaser Tom
Published: April 2021

We have identified Math5, a mouse basic helix-loop-helix (bHLH) gene that is closely related to Drosophila atonal and Xenopus Xath5 and is largely restricted to the developing eye. Math5 retinal expression precedes differentiation of the first neurons and persists within progenitor cells until after birth. To position Math5 in a hierarchy of retinal development, we compared Math5 and Hes1 expression in wild-type and Pax6-deficient (Sey) embryos. Math5 expression is downregulated in Sey/+ eyes and abolished in Sey/Sey eye rudiments, whereas the bHLH gene Hes1 is upregulated in a similar dose-dependent manner. These results link Pax6 to the process of retinal neurogenesis and provide the first molecular correlate for the dosage-sensitivity of the Pax6 phenotype. During retinogenesis, Math5 is expressed significantly before NeuroD, Ngn2 or Mash1. To test whether these bHLH genes influence the fates of distinct classes of retinal neurons, we ectopically expressed Math5 and Mash1 in Xenopus retinal progenitors. Unexpectedly, lipofection of either mouse gene into the frog retina caused an increase in differentiated bipolar cells. Directed expression of Math5, but not Xath5, in Xenopus blastomeres produced an expanded retinal phenotype. We propose that Math5 acts as a proneural gene, but has properties different from its most closely related vertebrate family member, Xath5.
XBF-2 is a transcriptional repressor that converts ectoderm into neural tissue
Authors: Mariani Francesca V. , Harland Richard M.
Published: April 2021

We have identified Xenopus Brain Factor 2 (XBF-2) as a potent neuralizing activity in an expression cloning screen. In ectodermal explants, XBF-2 converts cells from an epidermal to a neural fate. Such explants contain neurons with distinct axonal profiles and express both anterior and posterior central nervous system (CNS) markers. In striking contrast to X-ngnR-1a or X-NeuroD, ectopic expression of XBF-2 in Xenopus embryos results in an expansion of the neural plate to the ventral midline. The enlarged neural plate consists predominantly of undifferentiated neurons. XBF-2 lies downstream of the BMP antagonists noggin, cerberus, and gremlin since ectodermal explants expressing these molecules exhibit strong expression of XBF-2. While XBF-2 does not upregulate the expression of secreted neural inducers, it downregulates the transcription of BMP-4, an epidermal inducer. We show that XBF-2 acts as a transcriptional repressor and that its effects can be phenocopied with either the engrailed or hairy repressor domain fused to the XBF-2 DNA-binding domain. A fusion of the DNA-binding domain to the activator domain of VP16 blocks the effects of XBF-2 and prevents neural plate development in the embryo. This provides evidence that a transcriptional repressor can affect both regional neural development and neurogenesis in vertebrates.
Hyperactivation of the folded gastrulation pathway induces specific cell shape changes
Authors: Morize Pierre , Christiansen Audrey E. , Costa Mike , Parks Suki , Wieschaus Eric
Published: April 2021

During Drosophila gastrulation, mesodermal precursors are brought into the interior of the embryo by formation of the ventral furrow. The first steps of ventral furrow formation involve a flattening of the apical surface of the presumptive mesodermal cells and a constriction of their apical diameters. In embryos mutant for folded gastrulation (fog), these cell shape changes occur but the timing and synchrony of the constrictions are abnormal. A similar phenotype is seen in a maternal effect mutant, concertina (cta). fog encodes a putative secreted protein whereas cta encodes an α-subunit of a heterotrimeric G protein. We have proposed that localized expression of the fog signaling protein induces apical constriction by interacting with a receptor whose downstream cellular effects are mediated by the cta Gα protein.

In order to test this model, we have ectopically expressed fog at the blastoderm stage using an inducible promoter. In addition, we have examined the constitutive activation of cta protein by blocking GTP hydrolysis using both in vitro synthesized mutant alleles and cholera toxin treatment. Activation of the fog/cta pathway by any of these procedures results in ectopic cell shape changes in the gastrula. Uniform fog expression rescues the gastrulation defects of fog null embryos but not cta mutant embryos, arguing that cta functions downstream of fog expression. The normal location of the ventral furrow in embryos with uniformly expressed fog suggests the existence of a fog-independent pathway determining mesoderm-specific cell behaviors and invagination. Epistasis experiments indicate that this pathway requires snail but not twist expression.
Sensory axons are guided by local cues in the developing dorsal spinal cord
Authors: Sharma Kamal , Frank Eric
Published: April 2021

During development, different classes of sensory neurons establish distinctive central projections within the spinal cord. Muscle spindle afferents (Ia fibers) grow ventrally through the dorsal horn to the ventral cord, whereas cutaneous sensory collaterals remain confined to the dorsal horn. We have studied the nature of the cues used by Ia fibers in establishing their characteristic projections within the dorsal horn. An organotypic culture preparation of embryonic chicken spinal cord and sensory ganglia was used to test the influence of ventral spinal cord and local cues within the dorsal spinal cord on the growing Ia afferents.

When the ventral half of the spinal cord was replaced with an inverted duplicate dorsal half, Ia fibers entering through the dorsal columns still grew ventrally within the host dorsal horn. After the fibers entered the duplicate dorsal half, they continued growing in the same direction. With respect to the duplicate dorsal tissue, this was in an opposite, ventral-to-dorsal, direction. In both cases, however, Ia collaterals remained confined to the medial dorsal laminae. Restriction to these laminae was maintained even when the fibers had to change their direction of growth to stay within them. These results show that cues from the ventral cord are not required for the development of correct Ia projections within the dorsal horn. Local, rather than long-range directional, cues appear to determine the pattern of these projections. When the ventral half of the spinal cord was left intact but sensory axons were forced to enter the dorsal gray matter growing rostrally or caudally, their collateral axons grew in random directions, further showing the absence of directional cues even when the ventral cord was present. Taken together, these observations suggest that Ia fibers are guided by local positional cues that keep them confined to the medial gray matter within the dorsal horn, but their direction of growth is determined primarily by their orientation and position as they enter the dorsal gray matter.
Two regulatory genes, cNkx5-1 and cPax2, show different responses to local signals during otic placode and vesicle formation in the chick embryo
Authors: Herbrand Heike , Guthrie Sarah , Hadrys Thorsten , Hoffmann Sonja , Arnold Hans-Henning , Rinkwitz-Brandt Silke , Bober Eva
Published: April 2021

The early stages of otic placode development depend on signals from neighbouring tissues including the hindbrain. The identity of these signals and of the responding placodal genes, however, is not known. We have identified a chick homeobox gene cNkx5-1, which is expressed in the otic placode beginning at stage 10 and exhibits a dynamic expression pattern during formation and further differentiation of the otic vesicle. In a series of heterotopic transplantation experiments, we demonstrate that cNkx5-1 can be activated in ectopic positions. However, significant differences in otic development and cNkx5-1 gene activity were observed when placodes were transplanted into the more rostral positions within the head mesenchyme or into the wing buds of older hosts. These results indicate that only the rostral tissues were able to induce and/or maintain ear development. Ectopically induced cNkx5-1 expression always reproduced the endogenous pattern within the lateral wall of the otocyst that is destined to form vestibular structures. In contrast, cPax2 which is expressed in the medial wall of the early otic vesicle later forming the cochlea never resumed its correct expression pattern after transplantation. Our experiments illustrate that only some aspects of gene expression and presumably pattern formation during inner ear development can be established and maintained ectopically. In particular, the dorsal vestibular structures seem to be programmed earlier and differently from the ventral cochlear part.
The genetic control of the distinction between fat body and gonadal mesoderm in Drosophila
Authors: Riechmann Veit , Rehorn Klaus-Peter , Reuter Rolf , Leptin Maria
Published: April 2021

The somatic muscles, the heart, the fat body, the somatic part of the gonad and most of the visceral muscles are derived from a series of segmentally repeated primordia in the Drosophila mesoderm. This work describes the early development of the fat body and its relationship to the gonadal mesoderm, as well as the genetic control of the development of these tissues.

Segmentation and dorsoventral patterning genes define three regions in each parasegment in which fat body precursors can develop. Fat body progenitors in these regions are specified by different genetic pathways. Two regions require engrailed and hedgehog for their development while the third is controlled by wingless. decapentaplegic and one or more unknown genes determine the dorsoventral extent of these regions. In each of parasegments 10-12 one of these regions generates somatic gonadal precursors instead of fat body. The balance between fat body and somatic gonadal fate in these serially homologous cell clusters is controlled by at least five genes. We suggest a model in which tinman, engrailed and wingless are necessary to permit somatic gonadal develoment, while serpent counteracts the effects of these genes and promotes fat body development. The homeotic gene abdominalA limits the region of serpent activity by interfering in a mutually repressive feed back loop between gonadal and fat body development.
Genetic control of branching pattern and floral identity during Petunia inflorescence development
Authors: Souer Erik , Krol Alexander van der , Kloos Daisy , Spelt Cornelis , Bliek Mattijs , Mol Jos , Koes Ronald
Published: April 2021

A main determinant of inflorescence architecture is the site where floral meristems are initiated. We show that in wild-type Petunia bifurcation of the inflorescence meristem yields two meristems of approximately equal size. One terminates into a floral meristem and the other maintains its inflorescence identity. By random transposon mutagenesis we have generated two mutants in which the architecture of the inflorescence is altered. In the extra petals− (exp) mutant the inflorescence terminates with the formation of a single terminal flower. Phenotypic analysis showed that exp is required for the bifurcation of inflorescence meristems. In contrast, the aberrant leaf and flower− (alf) mutant is affected in the specification of floral meristem identity while the branching pattern of the inflorescence remains unaltered. A weak alf allele was identified that, after bifurcation of the inflorescence meristem, yields a ‘floral’ meristem with partial inflorescence characteristics. By analysing independent transposon dTph1 insertion alleles we show that the alf locus encodes the Petunia FLORICAULA/LEAFY homolog. In situ hybridisation shows that alf is expressed in the floral meristem and also in the vegetative meristem. Differences and similarities between these Petunia mutants and mutations affecting inflorescence architecture in other species will be discussed.
The hsp70 protein is involved in the acquisition of gamete self-sterility in the ascidian Ciona intestinalis
Authors: Marino Rita , Pinto Maria Rosaria , Cotelli Franco , Lamia Carla Lora , De Santis Rosaria
Published: April 2021

In the hermaphrodite ascidian Ciona intestinalis, gamete self-incompatibility is a mechanism that prevents selffertilization and is based on the ability of the oocyte vitelline coat to distinguish and accept only heterologous spermatozoa. The onset of self-sterility occurs during oogenesis and involves or is controlled by the follicle cells. Gamete self-nonself discrimination, a process that can be likened to an immune recognition event, represents a useful model with which to study the evolution of self-nonself recognition. Hsp70 genes, which belong to the major histocompatibility complex (MHC) class III, are supposedly ancestors of the MHC class I and II genes, and chaperonins are known to be involved in antigen processing and presentation. We have isolated and characterized an hsp70gene (Cihsp70) that is constitutively expressed during oogenesis in the follicle cells of previtellogenic and vitellogenic oocytes. Using a polyclonal antibody against Cihsp70 protein, we demonstrate that the expression of Cihsp70 is required for the switch from self-fertility to self-sterility. The functional involvement of Cihsp70 in gamete self-nonself recognition provides evidence for an ancestral MHC-like system in protochordates.
Progenitors of dorsal commissural interneurons are defined by MATH1 expression
Authors: Helms Amy W. , Johnson Jane E.
Published: April 2021

MATH1 is a neural-specific basic helix-loop-helix transcription factor. Members of this family of transcription factors are involved in the development of specific subsets of neurons in the developing vertebrate nervous system. Here we examine the cells expressing MATH1 with respect to their proliferative state and coexpression of cell-type-specific differentiation markers. We localize the MATH1 protein to the nucleus of cells in the dorsal neural tube and the external germinal layer (EGL) of the developing cerebellum. Using double-label immunofluorescence, we demonstrate that MATH1- expressing cells span both the proliferating and the differentiating zones within the dorsal neural tube, but within the EGL of the cerebellum are restricted to the proliferating zone. The early differentiating MATH1-expressing cells in the dorsal neural tube co-express TAG-1, DCC-1 and LH2, markers of dorsal commissural interneurons. In addition, transgenic mice with lacZ under the transcriptional control of MATHl-flanking DNA sequences express β -galactosidase specifically in the developing nervous system, in a manner that mimics subsets of the MATH 1-expression pattern, including the dorsal spinal neural tube. Expression of the MATHl/lacZ transgene persists in differentiated dorsal commissural interneurons. Taken together, we demonstrate MATH1 expression in a differentiating population of neuronal precursors in the dorsal neural tube that appear to give rise specifically to dorsal commissural interneurons.
Integrated FGF and BMP signaling controls the progression of progenitor cell differentiation and the emergence of pattern in the embryonic anterior pituitary
Authors: Ericson Johan , Norlin Stefan , Jessell Thomas M. , Edlund Thomas
Published: April 2021

The mechanisms by which inductive signals control the identity, proliferation and timing of differentiation of progenitor cells in establishing spatial pattern in developing vertebrate tissues remain poorly understood. We have addressed this issue in the embryonic anterior pituitary, an organ in which distinct hormone cell types are generated in a precise temporal and spatial order from an apparently homogenous ectodermal primordium. We provide evidence that in this tissue the coordinate control of progenitor cell identity, proliferation and differentiation is imposed by spatial and temporal restrictions in FGF- and BMP-mediated signals. These signals derive from adjacent neural and mesenchymal signaling centers: the infundibulum and ventral juxtapituitary mesenchyme. The infundibulum appears to have a dual signaling function, serving initially as a source of BMP4 and subsequently of FGF8. The ventral juxtapituitary mesenchyme appears to serve as a later source of BMP2 and BMP7. In vitro, FGFs promote the proliferation of progenitor cells, prevent their exit from the cell cycle and contribute to the specification of progenitor cell identity. BMPs, in contrast, have no apparent effect on cell proliferation but instead appear to act with FGFs to control the initial selection of thyrotroph and corticotroph progenitor identity.
Genetic interactions between Hoxa1 and Hoxb1 reveal new roles in regulation of early hindbrain patterning
Authors: Studer Michèle , Gavalas Anthony , Marshall Heather , Ariza-McNaughton Linda , Rijli Filippo M. , Chambon Pierre , Krumlauf Robb
Published: April 2021

In the developing vertebrate hindbrain Hoxa1 and Hoxb1 play important roles in patterning segmental units (rhombomeres). In this study, genetic analysis of double mutants demonstrates that both Hoxa1 and Hoxb1 participate in the establishment and maintenance of Hoxb1 expression in rhombomere 4 through auto- and para-regulatory interactions. The generation of a targeted mutation in a Hoxb1 3′ retinoic acid response element (RARE) shows that it is required for establishing early high levels of Hoxb1 expression in neural ectoderm. Double mutant analysis with this Hoxb13′RARE allele and other targeted loss-of-function alleles from both Hoxa1 and Hoxb1 reveals synergy between these genes. In the absence of both genes, a territory appears in the region of r4, but the earliest r4 marker, the Eph tyrosine kinase receptor EphA2, fails to be activated. This suggests a failure to initiate rather than maintain the specification of r4 identity and defines new roles for both Hoxb1 and Hoxa1 in early patterning events in r4. Our genetic analysis shows that individual members of the vertebrate labial-related genes have multiple roles in different steps governing segmental processes in the developing hindbrain.
dorsotonals/homothorax, the Drosophila homologue of meis1, interacts with extradenticle in patterning of the embryonic PNS
Authors: Kurant Estee , Pai Chi-yun , Sharf Rakefet , Halachmi Naomi , Sun Y. Henry , Salzberg Adi
Published: April 2021

The homeotic genes of the bithorax complex are required, among other things, for establishing the patterns of sensory organs in the embryonic peripheral nervous system (PNS). However, the molecular mechanisms by which these genes affect pattern formation in the PNS are not understood and other genes that function in this pathway are not characterized. Here we report the phenotypic and molecular analysis of one such gene, homothorax (hth; also named dorsotonals). Mutations in the hth gene seem to alter the identity of the abdominal chordotonal neurons, which depend on Abd-A for their normal development. However, these mutations do not alter the expression of the abd-A gene, suggesting that hth may be involved in modulating abd-A activity. We have generated multiple mutations in the hth locus and cloned the hth gene. hth encodes a homeodomain-containing protein that is most similar to the murine proto-oncogene meis1. The hth gene is expressed throughout embryonic development in a spatially restricted pattern, which is modulated in abdominal segments by abd-A and Ubx.

The spatial distribution of the HTH protein during embryonic development is very similar to the distribution of the Extradenticle (EXD) protein, a known modulator of homeotic gene activity. Here we show that the PNS phenotype of exd mutant embryos is virtually indistinguishable from that of hth mutant embryos and does not simply follow the homeotic transformations observed in the epidermis. We also show that the HTH protein is present in extremely low levels in embryos lacking exd activity as compared to wild-type embryos. In contrast, the EXD protein is present in fairly normal levels in hth mutant embryos, but fails to accumulate in nuclei and remains cytoplasmic. Ectopic expression of hth can drive ectopic nuclear localization of EXD. Based on our observations we propose that the genetic interactions between hth and exd serve as a novel mechanism for regulating homeotic protein activity in embryonic PNS development.
EGL-17(FGF) expression coordinates the attraction of the migrating sex myoblasts with vulval induction in C. elegans
Authors: Burdine Rebecca D. , Branda Catherine S. , Stern Michael J.
Published: April 2021

During the development of the egg-laying system in Caenorhabditis elegans hermaphrodites, central gonadal cells organize the alignment of the vulva with the sex myoblasts, the progenitors of the egg-laying muscles. A fibroblast growth factor [EGL-17(FGF)] and an FGF receptor [EGL-15(FGFR)] are involved in the gonadal signals that guide the migrations of the sex myoblasts. Here we show that EGL-17(FGF) can act as an instructive guidance cue to direct the sex myoblasts to their final destinations. We find that egl-17 reporter constructs are expressed in the primary vulval cell and that EGL-17(FGF) expression in this cell correlates with the precise positioning of the sex myoblasts. We postulate that EGL-17(FGF) helps to coordinate the development of a functional egg-laying system, linking vulval induction with proper sex myoblast migration.
Regulation in the heart field of zebrafish
Authors: Serbedzija George N. , Chen Jau-Nian , Fishman Mark C.
Published: April 2021

In many vertebrates, removal of early embryonic heart precursors can be repaired, leaving the heart and embryo without visible deficit. One possibility is that this ‘regulation’ involves a cell fate switch whereby cells, perhaps in regions surrounding normal progenitors, are redirected to the heart cell fate. However, the lineage and spatial relationships between cells that are normal heart progenitors and those that can assume that role after injury are not known, nor are their molecular distinctions.

We have adapted a laser-activated technique to label single or small patches of cells in the lateral plate mesoderm of the zebrafish and to track their subsequent lineage. We find that the heart precursor cells are clustered in a region adjacent to the prechordal plate, just anterior to the notochord tip. Complete unilateral ablation of all heart precursors with a laser does not disrupt heart development, if performed before the 18-somite stage. By combining extirpation of the heart precursors with cell labeling, we find that cells anterior to the normal cardiogenic compartments constitute the source of regulatory cells that compensate for the loss of the progenitors.

One of the earliest embryonic markers of the premyocardial cells is the divergent homeodomain gene, Nkx2.5. Interestingly, normal cardiogenic progenitors derive from only the anterior half of the Nkx2.5-expressing region in the lateral plate mesoderm. The posterior half, adjacent to the notochord, does not include cardiac progenitors and the posterior Nkx2.5-expressing cells do not contribute to the heart, even after ablation of the normal cardiogenic region.

The cells that can acquire a cardiac cell fate after injury to the normal progenitors also reside near the prechordal plate, but anterior to the Nkx2.5-expressing domain. Normally they give rise to head mesenchyme. They share with cardiac progenitors early expression of GATA 4. The location of the different elements of the cardiac field, and their response to injury, suggests that the prechordal plate supports and/or the notochord suppresses the cardiac fate.
Tripartite organization of the ancestral chordate brain and the antiquity of placodes: insights from ascidian Pax-2/5/8, Hox and Otx genes
Authors: Wada Hiroshi , Saiga Hidetoshi , Satoh Noriyuki , Holland Peter W. H.
Published: April 2021

Ascidians and vertebrates belong to the Phylum Chordata and both have dorsal tubular central nervous systems. The structure of the ascidian neural tube is extremely simple, containing less than 400 cells, among which less than 100 cells are neurons. Recent studies suggest that, despite its simple organization, the mechanisms patterning the ascidian neural tube are similar to those of the more complex vertebrate brain. Identification of homologous regions between vertebrate and ascidian nervous systems, however, remains to be resolved. Here we report the expression of HrPax-258 gene: an ascidian homologue of vertebrate Pax-2, Pax-5 and Pax-8 genes. Molecular phylogenetic analyses indicate that HrPax-258 is descendant from a single precursor gene that gave rise to the three vertebrate genes. The expression pattern of HrPax-258 suggests that this subfamily of Pax genes has conserved roles in regional specification of the brain. Comparison with expression of ascidian Otx (Hroth) and a Hox gene (HrHox1) by double-staining in situ hybridizations indicate that the ascidian brain region can be subdivided into three regions; the anterior region marked by Hroth probably homologous to the vertebrate forebrain and midbrain, the middle region marked by HrPax-258 probably homologous to the vertebrate anterior hindbrain (and maybe also midbrain) and the posterior region marked by Hox genes which is homologous to the vertebrate hindbrain and spinal cord. Later expression of HrPax-258 in atrial primordia implies that basal chordates such as ascidians have already acquired a sensory organ that develops from epidermal thickenings (placodes) and expresses HrPax-258; we suggest it is homologous to the vertebrate ear. Therefore, placodes are not likely to be a newly acquired feature in vertebrates, but may have already been possessed by the earliest chordates.