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Glycosyltransferase encoding gene EXTL3 is differentially expressed in the developing and adult mouse cerebral cortex.

Exostosin tumor-like 3 (EXTL3) is a glycosyltransferase involved in heparan sulfate (HS) biosynthesis. HS proteoglycans are critically involved in different steps during brain development. The present in situ hybridization in mice revealed wide EXTL3 expression at different grades in the central and peripheral nervous system components including the neural retina and neural crest-derived structures at embryonic days (E) 11.5, E12.5, E14.5, and E16.5. In the neopallial cortex, an intense EXTL3 expression was observed in the neuroepithelial cells lining the ventricular zone at E11.5 and E12.5. The signal decreased at E14.5 and was further downregulated at E16.5 in the ventricular zone. The pioneer neurons of the preplate at E12.5 differentially expressed the gene. Heavily stained among weakly or negatively stained neurons were observed. At E14.5, the cortical plate cells were moderately and homogeneously stained. In contrast, at E16.5, an upregulated and differential expression pattern was detected. The labeling pattern at E16.5 subdivided the cortical plate cells into a large number of heavily, a moderate number of less intensely, and some negatively stained cell populations. Interestingly, the distinct expression pattern displayed by the three main cell types of the adult cerebral cortex was similar to that of the late corticogenesis stage (E16.5). In the adult, the strongest expression was observed in the pyramidal neurons. The granule-type neurons showed less intense staining while the glia cells were devoid of signals. Our data revealed that EXTL3 expression is developmentally regulated in the mouse nervous system and suggested that it differentially contributes to brain development and corticogenesis.

Animals↗

Effects of male and female sex steroids on the development of normal and the transient Froriep's dorsal root ganglia of the chick embryo.

Sex steroids can influence developmental processes and support the survival of neurons in the embryonic central nervous system. Recent studies have shown that estrogen receptors are also expressed in the peripheral nervous system, in the dorsal root ganglia (DRG) of chick embryos. However, no studies have examined the effects of sex steroids on development of embryonic DRG. In the present study, 0.2 microg, 1.0 microg, 5.0 microg 10 microg, 20 microg, 25 microg, and 40 microg doses of testosterone or estradiol were delivered to chick embryos at Hamburger and Hamilton stage 18 (E3). The actions of these doses of sex steroids on the development of the C5DRG (fifth cervical ganglion, a "normal" DRG) and C2DRG (a transient ganglion known as a "Froriep's DRG") were then evaluated by quantifying ganglionic volumes, cell number, proliferation, and apoptosis after 1 day of growth to stage 23. We found that both testosterone and estradiol promoted proliferation of cells in both normal DRG and the Froriep's ganglia. By contrast, estradiol significantly increased the number of apoptotic cells, while testosterone strongly inhibited apoptosis. These actions of sex steroids on DRG development were dose-dependent, and C5DRG and C2DRG showed different sensitivities to the applied sex steroids. In addition, the present results demonstrated that specific ER and AR inhibitors (tamoxifen and flutamide) did not influence the effects of 5 microg E2 and 5 microg T on C2 and C5DRG significantly. These results demonstrate that male and female sex steroids can modulate DRG development through an epigenetic mechanism, as had been shown for the central nervous system.

Androgen Antagonists↗

Control of cell proliferation in the Drosophila eye by Notch signaling.

Cell proliferation in animals must be precisely controlled, but the signaling mechanisms that regulate the cell cycle are not well characterized. A regulated terminal mitosis, called the second mitotic wave (SMW), occurs during Drosophila eye development, providing a model for the genetic analysis of proliferation control. We report a cell cycle checkpoint at the G1-S transition that initiates the SMW, and we demonstrate that Notch signaling is required for cells to overcome this checkpoint. Notch triggers the onset of proliferation by multiple pathways, including the activation of dE2F1, a member of the E2F transcription factor family. Delta to Notch signaling derepresses the inhibition of dE2F1 by RBF, and Delta expression depends on the secreted proteins Hedgehog and Dpp. Notch is also required for the expression of Cyclin A in the SMW.

Animals↗

Echinoid is a component of adherens junctions that cooperates with DE-Cadherin to mediate cell adhesion.

Echinoid is an immunoglobulin domain-containing transmembrane protein that modulates cell-cell signaling by Notch and the EGF receptors. We show that, in the Drosophila wing disc epithelium, Echinoid is a component of adherens junctions that cooperates with DE-Cadherin in cell adhesion. Echinoid and beta-catenin (a DE-Cadherin interacting protein) each possess a C-terminal PDZ domain binding motif that binds to Bazooka/PAR-3; these motifs redundantly position Bazooka to adherens junctions. Echinoid also links to actin filaments by binding to Canoe/AF-6/afadin. Moreover, interfaces between Echinoid- and Echinoid+ cells, like those between DE-Cadherin- and DE-Cadherin+ cells, are deficient in adherens junctions and form actin cables. These characteristics probably facilitate the strong sorting behavior of cells that lack either of these cell-adhesion molecules. Finally, cells lacking either Echinoid or DE-Cadherin accumulate a high density of the reciprocal protein, further suggesting that Echinoid and DE-Cadherin play similar and complementary roles in cell adhesion.

Actins↗

Drosophila E2F1 has context-specific pro- and antiapoptotic properties during development.

E2F transcription factors are generally believed to be positive regulators of apoptosis. In this study, we show that dE2F1 and dDP are important for the normal pattern of DNA damage-induced apoptosis in Drosophila wing discs. Unexpectedly, the role that E2F plays varies depending on the position of the cells within the disc. In irradiated wild-type discs, intervein cells show a high level of DNA damage-induced apoptosis, while cells within the D/V boundary are protected. In irradiated discs lacking E2F regulation, intervein cells are largely protected, but apoptotic cells are found at the D/V boundary. The protective effect of E2F at the D/V boundary is due to a spatially restricted role in the repression of hid. These loss-of-function experiments demonstrate that E2F cannot be classified simply as a pro- or antiapoptotic factor. Instead, the overall role of E2F in the damage response varies greatly and depends on the cellular context.

Animals↗

Heparan sulfate in trans potentiates VEGFR-mediated angiogenesis.

Several receptor tyrosine kinases require heparan sulfate proteoglycans (HSPGs) as coreceptors for efficient signal transduction. We have studied the role of HSPGs in the development of blood capillary structures from embryonic stem cells, a process strictly dependent on signaling via vascular endothelial growth factor receptor-2 (VEGFR-2). We show, by using chimeric cultures of embryonic stem cells defective in either HS production or VEGFR-2 synthesis, that VEGF signaling in endothelial cells is fully supported by HS expressed in trans by adjacent perivascular smooth muscle cells. Transactivation of VEGFR-2 leads to prolonged and enhanced signal transduction due to HS-dependent trapping of the active VEGFR-2 signaling complex. Our data imply that direct signaling via HSPG core proteins is dispensable for a functional VEGF response in endothelial cells. We propose that transactivation of tyrosine kinase receptors by HSPGs constitutes a mechanism for crosstalk between adjacent cells.

Amidohydrolases↗

Eml5, a novel WD40 domain protein expressed in rat brain.

We have isolated a novel transcript with homology to the major microtubule-associated protein in dividing sea urchin embryos, EMAP. The protein has a predicted MW of approximately 180 kDa and we have named it Eml5 (EMAP-like protein 5). Eml5 contains 11 putative WD40 domains and 3 hydrophobic stretches of 43 aa, HELP domains, which have been suggested to be involved in microtubule binding. Eml5 appears to consist of two tandem repeats of the complete EMAP protein separated by a putative dimerization domain. Eml5 mRNA and protein is expressed at high levels in the hippocampus, cerebellum and olfactory bulb, as determined by in situ hybridization and immunocytochemistry. Eml5 transcripts can be detected in fore- and hindbrain structures from embryonic day 13 onwards. Because other EMAP-like proteins are involved in regulating microtubule dynamics, it is likely that Eml5 plays a role in the regulation of cytoskeletal rearrangements during neuronal development and in adult brain

Alternative Splicing↗

The mouse Trm1-like gene is expressed in neural tissues and plays a role in motor coordination and exploratory behaviour.

Using a gene trap approach in ES cells, the novel mouse gene Trm1-like with substantial sequence homology to human C1orf25 mRNA (GenBank accession no. ) was identified. Murine Trm1-like encodes a putative protein with limited similarity to N2,N2-dimethylguanosine tRNA methyltransferase (Trm1) from other organisms, however its function is not known. The potential role of Trm1-like was investigated in a mouse mutant lacking intact Trm1-like transcripts due to integration of the gene trap vector in the first intron. Trm1-like deficient mice are viable and show no apparent anatomical defects. Behavioural tests, however, revealed significantly altered motor coordination and aberrant exploratory behaviour. LacZ activity of the trapped mouse Trm1-like gene reflects expression in various neuronal structures during embryonic development, including spinal ganglia, trigeminal nerve and ganglion, olfactory and nasopharyngeal epithelium, and nuclei of the metencephalon, thalamus and medulla oblongata. The gene is also expressed in lung, oesophagus, epiglottis, ependyma, vertebral column, spinal cord, and brown adipose tissue. Trm1-like expression persists in the adult brain with dynamically changing patterns in cortex and cerebellum. Although Trm1-like is not essential for embryonic mouse development, it may have a role in modulating postnatal neuronal functions.

Animals↗

Holoprosencephaly--report of two cases.

Holoprosencephaly (HPE) is a developmental defect resulting from incomplete cleavage of the embryonic forebrain structures during early embryogenesis. It includes a series of rare complex disorders. Chromosomal abnormalities, single gene disorders and teratogenic agents are responsible for holoprosencephaly. We report two rare cases of alobar HPE with a rudimental nasal structure (proboscis) located under the eyes. One of the patients survived for 9 months and the other one survived 40 days.

Brain↗

Localization of Brachyury (T) in embryonic and extraembryonic tissues during mouse gastrulation.

T-box gene family members have important roles during murine embryogenesis, gastrulation, and organogenesis. Although relatively little is known about how T-box genes are regulated, published gene expression studies have revealed dynamic and specific patterns in both embryonic and extraembryonic tissues of the mouse conceptus. Mutant alleles of the T-box gene Brachyury (T) have identified roles in formation of mesoderm and its derivatives, such as somites and the allantois. However, given the cell autonomous nature of T gene activity and conflicting results of gene expression studies, it has been difficult to attribute a primary function to T in normal allantoic development. We report localization of T protein by sectional immunohistochemistry in both embryonic and extraembryonic tissues during mouse gastrulation, emphasizing T localization within the allantois. T was detected in all previously reported sites within the conceptus, including the primitive streak and its derivatives, nascent embryonic mesoderm, the node and notochord, as well as notochord-associated endoderm and posterior neurectoderm. In addition, we have clarified T within the allantois, where it was first detected in the proximal midline of the late allantoic bud (approximately 7.5 days postcoitum, dpc) and persisted within an expanded midline domain until 6-somite pairs (s; approximately 8.5 dpc). Lastly, we have discovered several novel T sites, including the developing heart, visceral endoderm, extraembryonic ectoderm, and its derivative, chorionic ectoderm. Together, these data provide a unified picture of T in the mammalian conceptus, and demonstrate T's presence in unrelated cell types and tissues in highly dynamic spatiotemporal patterns in both embryonic and extraembryonic tissues.

Allantois↗

Hypoxic incubation creates differential morphological effects during specific developmental critical windows in the embryo of the chicken (Gallus gallus).

Hypoxia inhibits vertebrate development, but the magnitude and timing of organ-specific effects are poorly understood. Chick embryos were exposed continuously to hypoxia (15% O2) throughout Days 1-6, 6-12, 12-18 or Days 1-18 of development, followed by morphometric measurements of major organ systems. Early hypoxic exposure reduced eye mass and beak length when measured in middle development. Liver, brain, heart, kidneys, stomach, intestines and skeletal long bones were not affected by hypoxia at any developmental stage. The chorioallantoic membrane (CAM) mass was unchanged by hypoxic exposure in early or mid-development, but CAM mass on Day 18 increased strikingly (40 and 60% in late and continuous populations, respectively) in response to hypoxic exposure. The increase in CAM mass presumably enhances oxygen delivery, thus minimizing the detrimental effects of hypoxia on development and growth. Hypoxic exposure at key critical windows in development thus results in differential effects on organ development, some of which can subsequently be repaired through additional incubation (yolk mass, eye mass, beak length).

Animals↗

Real-time ultrasound measure of the fetal eye (vitreous body) for prediction of parturition date in small ponies.

Ultrasonographic fetal eye measures have been used to estimate gestational age of the fetus in light horse mares. However, fetal eye measures have not been published for smaller pony breeds. This study was conducted to develop reference ranges for ultrasonographic measures of fetal eyes in small ponies for the purpose of predicting days before parturition (DBP) when breeding or ovulation dates are unknown. Twenty-three Shetland-type pony mares were studied across one (n = 10) or two (n = 13) gestations in 2004 (18 pregnancies) and 2005 (18 pregnancies). Measurements of fetal eyes were obtained during transrectal ultrasound examination. Examinations were conducted once monthly in a field situation beginning in December (2003) or August (2004) until mares foaled (March through July). The length (from sclera to sclera) and width (from retina to cornea) of the vitreous body were measured. For the 273 examinations in which gestation age was greater than 2 months, eye measures were obtainable in 248 (91%). Mixed-effects linear regression modeling was used to account for serial growth measures within pregnancy, repeated measurements across mares, and unbalanced study design. Independent variables evaluated included vitreous body length, vitreous body width, the ratio of length to width, parity, and mare height at the withers at parturition. Eye length was the best single predictor of days before parturition, with almost no additional predictive value of the other variables considered. Our resulting regression equation is: days before parturition=265.16-0.21*(vitreous body length in mm)(2). This study suggests that measure of the fetal eye is a practical on-farm procedure for estimating days before parturition in small ponies.

Animals↗

Rapid replacement of somatic linker histones with the oocyte-specific linker histone H1foo in nuclear transfer.

The most distinctive feature of oocyte-specific linker histones is the specific timing of their expression during embryonic development. In Xenopus nuclear transfer, somatic linker histones in the donor nucleus are replaced with oocyte-specific linker histone B4, leading to the involvement of oocyte-specific linker histones in nuclear reprogramming. We recently have discovered a mouse oocyte-specific linker histone, named H1foo, and demonstrated its expression pattern in normal preimplantation embryos. The present study was undertaken to determine whether the replacement of somatic linker histones with H1foo occurs during the process of mouse nuclear transfer. H1foo was detected in the donor nucleus soon after transplantation. Thereafter, H1foo was restricted to the chromatin in up to two-cell stage embryos. After fusion of an oocyte with a cell expressing GFP (green fluorescent protein)-tagged somatic linker histone H1c, immediate release of H1c in the donor nucleus was observed. In addition, we used fluorescence recovery after photobleaching (FRAP), and found that H1foo is more mobile than H1c in living cells. The greater mobility of H1foo may contribute to its rapid replacement and decreased stability of the embryonic chromatin structure. These results suggest that rapid replacement of H1c with H1foo may play an important role in nuclear remodeling.

Animals↗

Follistatin complexes Myostatin and antagonises Myostatin-mediated inhibition of myogenesis.

Follistatin is known to antagonise the function of several members of the TGF-beta family of secreted signalling factors, including Myostatin, the most powerful inhibitor of muscle growth characterised to date. In this study, we compare the expression of Myostatin and Follistatin during chick development and show that they are expressed in the vicinity or in overlapping domains to suggest possible interaction during muscle development. We performed yeast and mammalian two-hybrid studies and show that Myostatin and Follistatin interact directly. We further show that single modules of the Follistatin protein cannot associate with Myostatin suggesting that the entire protein is required for the interaction. We analysed the interaction kinetics of the two proteins and found that Follistatin binds Myostatin with a high affinity of 5.84 x 10(-10) M. We next tested whether Follistatin suppresses Myostatin activity during muscle development. We confirmed our previous observation that treatment of chick limb buds with Myostatin results in a severe decrease in the expression of two key myogenic regulatory genes Pax-3 and MyoD. However, in the presence of Follistatin, the Myostatin-mediated inhibition of Pax-3 and MyoD expression is blocked. We additionally show that Myostatin inhibits terminal differentiation of muscle cells in high-density cell cultures of limb mesenchyme (micromass) and that Follistatin rescues muscle differentiation in a concentration-dependent manner. In summary, our data suggest that Follistatin antagonises Myostatin by direct protein interaction, which prevents Myostatin from executing its inhibitory effect on muscle development.

Animals↗

EphA4, RhoB and the molecular development of feather buds are maintained by the integrity of the actin cytoskeleton.

The development of feather buds is a highly ordered process involving epithelial-mesenchymal signalling. Cellular morphology is determined by the actin cytoskeleton, which is controlled by networks of regulators such as the GTPases. EphA4 belongs to a receptor tyrosine kinase family that has been consistently shown to regulate the cytoskeleton via Rho family GTPases in neural development and is expressed in early stages of feather bud development though its role has not been defined. We therefore used an in vitro skin culture system to interfere with EphA4 levels in feather buds using anti-sense oligonucleotides, demonstrating a severe effect on both their number and morphological form. Analysis of the Rho family of GTPases revealed that this effect was mediated by the GTPase RhoB, the expression of which was altered in response to altered levels of EphA4. In addition, the inhibition of RhoB mimicked the effects of reduced EphA4 levels on feather development. Significantly, manipulation of cytoskeletal dynamics revealed that those cells undergoing morphogenetic change regulate the patterning signals responsible for initiating feather development. We propose that this molecular maintenance mechanism between EphA4-RhoB and the actin cytoskeleton converges or coordinates with other morphogenic signalling systems to control feather bud development.

Actins↗

Molecular and genetic regulation of testis descent and external genitalia development.

Testicular descent as a prerequisite for the production of mature spermatozoa and normal external genitalia morphogenesis, and therefore facilitating copulation and internal fertilization, are essential developmental steps in reproduction of vertebrate species. Cryptorchidism, the failure of testis descent, and feminization of external genitalia in the male, usually in the form of hypospadias, in which the opening of the urethra occurs along the ventral aspect of the penis, are the most frequent pediatric complications. Thus, elucidating the molecular mechanisms involved in the regulation of testis descent and the formation of external genitalia merits a special focus. Natural and transgenic rodent models have demonstrated both morphogenic processes to be under the control of a plethora of genetic factors with complex time-, space-, and dose-restricted expression pattern. The review elucidates the molecular mechanisms involved in the regulation of testis descent and the formation of external genitalia and, wherever possible, assesses the differences between these rodent animal models and other mammalian species, including human.

Androgens↗

Discovery of genes implicated in placode formation.

The peripheral nervous system of the head is derived from cranial ectodermal placodes and neural crest cells. Placodes arise from thickenings in the cranial ectoderm that invaginate or ingress to form sensory ganglia and the paired sense organs. We have combined embryological techniques with array technology to identify genes that are expressed as a consequence of placode induction. As a secondary screen, we used whole mount in situ hybridization to determine the expression of candidate genes in various placodal domains. The results reveal 52 genes that are found in one or more placodes, including the olfactory, trigeminal, and otic placodes. Expression of some of these genes is retained in placodal derivatives. Furthermore, several genes are common to both neural crest and ectodermal placodes. This study presents the first array of candidate genes implicated in placode development, providing numerous new molecular markers for various stages of placode formation. Importantly, the results uncover previously unknown commonalities in genes expressed by multiple placodes and shared properties between placodes and other migratory cells, like neural crest cells.

Animals↗

Abnormalities of caudal pharyngeal pouch development in Pbx1 knockout mice mimic loss of Hox3 paralogs.

Pbx1 is a TALE-class homeodomain protein that functions in part as a cofactor for Hox class homeodomain proteins. Previous analysis of the in vivo functions of Pbx1 by targeted mutagenesis in mice has revealed roles for this gene in skeletal patterning and development and in the organogenesis of multiple systems. Both RNA expression and protein localization studies have suggested a possible role for Pbx1 in pharyngeal region development. As several Hox mutants have distinct phenotypes in this region, we investigated the potential requirement for Pbx1 in the development of the pharyngeal arches and pouches and their organ derivatives. Pbx1 homozygous mutants exhibited delayed or absent formation of the caudal pharyngeal pouches, and disorganized patterning of the third pharyngeal pouch. Formation of the third pouch-derived thymus/parathyroid primordia was also affected, with absent or hypoplastic primordia, delayed expression of organ-specific differentiation markers, and reduced proliferation of thymic epithelium. The fourth pouch and the fourth pouch-derived ultimobranchial bodies were usually absent. These phenotypes are similar to those previously reported in Hoxa3(-/-) single mutants and Hoxa1(-/-);Hoxb1(-/-) or Hoxa3(+/-);Hoxb3(-/-);Hoxd3(-/-) compound mutants, suggesting that Pbx1 acts together with multiple Hox proteins in the development of the caudal pharyngeal region. However, some aspects of the Pbx1 mutant phenotype included specific defects that were less severe than those found in known Hox mutant mice, suggesting that some functions of Hox proteins in this region are Pbx1-independent.

Animals↗