The beta 3 tubulin gene is a direct target of bagpipe and biniou in the visceral mesoderm of Drosophila

Previous studies have identified the NK homeobox gene bagpipe and the FoxF fork head domain gene biniou as essential regulators of visceral mesoderm development in Drosophila. Here we present additional genetic and molecular information on the functions of these two genes during visceral mesoderm morphogenesis and differentiation. We show that both genes are required for the activation of beta 3Tub60D in the visceral mesoderm, which encodes beta 3 tubulin. We demonstrate that a 254 bp derivative of a previously defined visceral mesoderm-specific enhancer element, vm1, from beta 3Tub60D contains one specific in vitro binding site for Bagpipe and two such sites for Biniou. While the wild-type version of the 254 bp enhancer is able to drive significant levels of reporter gene expression within the entire trunk visceral mesoderm, mutation of either the Bagpipe or the Biniou binding sites within this element results in a severe decrease of enhancer activity. Moreover, mutation of all three binding sites for Bagpipe and Biniou, respectively, results in the complete loss of enhancer activity. Together, these observations suggest that Bagpipe and Biniou serve as direct, partially redundant, and tissue-specific activators of the terminal differentiation gene beta 3Tub60D in the visceral mesoderm.     

Functions for Drosophila brachyenteron and forkhead in mesoderm specification and cell signalling.

The visceral musculature of the larval midgut of Drosophila has a lattice-type structure and consists of an inner stratum of circular fibers and an outer stratum of longitudinal fibers. The longitudinal fibers originate from the posterior tip of the mesoderm anlage, which has been termed the caudal visceral mesoderm (CVM). In this study, we investigate the specification of the CVM and particularly the role of the Drosophila Brachyury-homologue brachyenteron. Supported by fork head, brachyenteron mediates the early specification of the CVM along with zinc-finger homeodomain protein-1. This is the first function described for brachyenteron or fork head in the mesoderm of Drosophila. The mode of cooperation resembles the interaction of the Xenopus homologues Xbra and Pintallavis. Another function of brachyenteron is to establish the surface properties of the CVM cells, which are essential for their orderly migration along the trunk-derived visceral mesoderm. During this movement, the CVM cells, under the control of brachyenteron, induce the formation of one muscle/pericardial precursor cell in each parasegment. We propose that the functions of brachyenteron in mesodermal development of Drosophila are comparable to the roles of the vertebrate Brachyury genes during gastrulation.     

Expression patterns of fork head and goosecoid homologues in the mollusc Patella vulgata supports the ancestry of the anterior mesendoderm across Bilateria

We have characterised orthologues of the genes fork head and goosecoid in the gastropod Patella vulgata. In this species, the anterior-posterior (AP) axis is determined just before gastrulation, and leads to the specification of two mesodermal components on each side of the presumptive endoderm, one anterior (ectomesoderm), and one posterior (endomesoderm). Both fork head and goosecoid are expressed from the time the AP axis is specified, up to the end of gastrulation. fork head mRNA is detected in the whole endoderm, as well as in the anterior mesoderm, whereas goosecoid is only expressed anteriorly, in the three germ layers. The two genes are thus coexpressed in the anterior mesoderm, suggesting the latter's homology with vertebrate prechordal mesoderm. In addition, since prechordal plate is known to belong to an anterior, so called "head organiser", and since its inductive role is dependent on the function of the vertebrate fork head and goosecoid orthologues, we further suggest that the anterior mesoderm may also have a role in anterior inductive patterning in Spiralia. Finally, we propose that a mode of axial development involving two organisers, one anterior and one posterior, is ancestral to the Bilateria, and that both organisers evolved from the single head organiser of a putative hydra-like ancestor.     

Spatially distinct head and heart inducers within the Xenopus organizer region.

BACKGROUND: The mouse anterior visceral endoderm, an extraembryonic tissue, expresses several genes essential for normal development of structures rostral to the anterior limit of the notochord and has been termed the head organizer. This tissue also has heart-inducing activity and expresses mCer1 which, like its Xenopus homolog cerberus, can induce markers of cardiac specification and anterior neural tissue when ectopically expressed. We investigated the relationship between head and heart induction in Xenopus embryos, which lack extraembryonic tissues. RESULTS: We found three regions of gene expression in the Xenopus organizer: deep endoderm, which expressed cerberus; prechordal mesoderm, which showed overlapping but non-identical expression of genes characteristic of the murine head organizer, such as XHex and XANF-1; and leading-edge dorsoanterior endoderm, which expressed both cerberus and a subset of the genes expressed by the prechordal mesoderm. Microsurgical ablation of the cerberus-expressing endoderm decreased the incidence of heart, but not head, formation. Removal of prechordal mesoderm, in contrast, caused deficits of anterior head structures. Finally, although misexpression of cerberus induced ectopic heads, it was unable to induce genes thought to participate in head induction. CONCLUSIONS: In Xenopus, the cerberus-expressing endoderm is required for heart, but not head, inducing activity. Therefore, this tissue is not the topological equivalent of the murine anterior visceral endoderm. We propose that, in Xenopus, cerberus is redundant to other bone morphogenetic protein (BMP) and Wnt antagonists located in prechordal mesoderm for head induction, but may be necessary for heart induction.     

Functional subdivision of trunk visceral mesoderm parasegments in Drosophila is required for gut and trachea development

In Drosophila, trunk visceral mesoderm, a derivative of dorsal mesoderm, gives rise to circular visceral muscles. It has been demonstrated that the trunk visceral mesoderm parasegment is subdivided into at least two domains by connectin expression, which is regulated by Hedgehog and Wingless emanating from the ectoderm. We now extend these findings by examining a greater number of visceral mesodermal genes, including hedgehog and branchless. Each visceral mesodermal parasegment appears to be divided into five or six regions, based on differences in expression patterns of these genes. Ectodermal Hedgehog and Wingless differentially regulate the expression of these metameric targets in trunk visceral mesoderm. hedgehog expression in trunk visceral mesoderm is responsible for maintaining its own expression and con expression. hedgehog expressed in visceral mesoderm parasegment 3 may also be required for normal decapentaplegic expression in this region and normal gastric caecum development. branchless expressed in each trunk visceral mesodermal parasegment serves as a guide for the initial budding of tracheal visceral branches. The metameric pattern of trunk visceral mesoderm, organized in response to ectodermal instructive signals, is thus maintained at a later time via autoregulation, is required for midgut morphogenesis and exerts feedback effect on trachea, ectodermal derivatives.     

Role of the chicken homeobox-containing genes GHox-4.6 and GHox-8 in the specification of positional identities during the development of normal and polydactylous chick limb buds.

During early stages of normal chick limb development, the homeobox-containing (HOX) gene GHox-4.6 is expressed throughout the posterior mesoderm of the wing bud from which most of the skeletal elements including the digits will develop, whereas GHox-8 is expressed in the anterior limb bud mesoderm which will not give rise to skeletal elements. In the present study, we have examined the expression of GHox-4.6 and GHox-8 in the wing buds of two polydactylous mutant chick embryos, diplopodia-5 and talpid2, from which supernumerary digits develop from anterior limb mesoderm, and have also examined the expression of these genes in response to polarizing zone grafts and retinoic acid-coated bead implants which induce the formation of supernumerary digits from anterior limb mesoderm. We have found that the formation of supernumerary digits from the anterior mesoderm in mutant and experimentally induced polydactylous limb buds is preceded by the ectopic expression of GHox-4.6 in the anterior mesoderm and the coincident suppression of GHox-8 expression in the anterior mesoderm. These observations suggest that the anterior mesoderm of the polydactylous limb buds is "posteriorized" and support the suggestion that GHox-8 and GHox-4.6, respectively, are involved in specifying the anterior non-skeletal and posterior digit-forming regions of the limb bud. Although the anterior mesodermal domain of GHox-8 expression is severely impaired in the mutant and experimentally induced polydactylous limb buds, this gene is expressed by the prolonged, thickened apical ectodermal ridges of the polydactylous limb buds that extend along the distal anterior as well as the distal posterior mesoderm.(ABSTRACT TRUNCATED AT 250 WORDS)     

The immunoglobulin-like protein Hibris functions as a dose-dependent regulator of myoblast fusion and is differentially controlled by Ras and Notch signaling.

Hibris (Hbs) is a transmembrane immunoglobulin-like protein that shows extensive homology to Drosophila Sticks and stones (Sns) and human kidney protein Nephrin. Hbs is expressed in embryonic visceral, somatic and pharyngeal mesoderm among other tissues. In the somatic mesoderm, Hbs is restricted to fusion competent myoblasts and is regulated by Notch and Ras signaling pathways. Embryos that lack or overexpress hbs show a partial block of myoblast fusion, followed by abnormal muscle morphogenesis. Abnormalities in visceral mesoderm are also observed. In vivo mapping of functional domains suggests that the intracellular domain mediates Hbs activity. Hbs and its paralog, Sns, co-localize at the cell membrane of fusion-competent myoblasts. The two proteins act antagonistically: loss of sns dominantly suppresses the hbs myoblast fusion and visceral mesoderm phenotypes, and enhances Hbs overexpression phenotypes. Data from a P-homed enhancer reporter into hbs and co-localization studies with Sns suggest that hbs is not continuously expressed in all fusion-competent myoblasts during the fusion process. We propose that the temporal pattern of hbs expression within fusion-competent myoblasts may reflect previously undescribed functional differences within this myoblast population.     

Development of the head and trunk mesoderm in the dogfish,Scyliorhinus torazame: II. Comparison of gene expression between the head mesoderm and somites with reference to the origin of the vertebrate head

The vertebrate mesoderm differs distinctly between the head and trunk, and the evolutionary origin of the head mesoderm remains enigmatic. Although the presence of somite‐like segmentation in the head mesoderm of model animals is generally denied at molecular developmental levels, the appearance of head cavities in elasmobranch embryos has not been explained, and the possibility that they may represent vestigial head somites once present in an amphioxus‐like ancestor has not been ruled out entirely. To examine whether the head cavities in the shark embryo exhibit any molecular signatures reminiscent of trunk somites, we isolated several developmentally key genes, including Pax1, Pax3, Pax7, Pax9, Myf5, Sonic hedgehog, and Patched2, which are involved in myogenic and chondrogenic differentiation in somites, and Pitx2, Tbx1, and Engrailed2, which are related to the patterning of the head mesoderm, from an elasmobranch species, Scyliorhinus torazame. Observation of the expression patterns of these genes revealed that most were expressed in patterns that resembled those found in amniote embryos. In addition, the head cavities did not exhibit an overt similarity to somites; that is, the similarity was no greater than that of the unsegmented head mesoderm in other vertebrates. Moreover, the shark head mesoderm showed an amniote‐like somatic/visceral distinction according to the expression of Pitx2, Tbx1, and Engrailed2. We conclude that the head cavities do not represent a manifestation of ancestral head somites; rather, they are more likely to represent a derived trait obtained in the lineage of gnathostomes.     

Neural crest can form cartilages normally derived from mesoderm during development of the avian head skeleton

The lateral wall of the avian braincase, which is indicative of the primitive amniote condition, is formed from mesoderm. In contrast, mammals have replaced this portion of their head skeleton with a nonhomologous bone of neural crest origin. Features that characterize the local developmental environment may have enabled a neural crest-derived skeletal element to be integrated into a mesodermal region of the braincase during the course of evolution. The lateral wall of the braincase lies along a boundary in the head that separates neural crest from mesoderm, and also, neural crest cells migrate through this region on their way to the first visceral arch. Differences in the availability of one skeletogenic population versus the other may determine the final composition of the lateral wall of the braincase. Using the quail-chick chimeric system, this investigation tests if populations of neural crest, when augmented and expanded within populations of mesoderm, will give rise to the lateral wall of the braincase. Results demonstrate that neural crest can produce cartilages that are morphologically indistinguishable from elements normally generated by mesoderm. These findings (1) indicate that neural crest can respond to the same cues that both promote skeletogenesis and enable proper patterning in mesoderm, (2) challenge hypotheses on the nature of the boundary between neural crest and mesoderm in the head, and (3) suggest that changes in the allocation of migrating cells could have enabled a neural crest-derived skeletal element to replace a mesodermal portion of the braincase during evolution.     

Expression and function of the homoeotic genes Antennapedia and Sex combs reduced in the embryonic midgut of Drosophila

Drosophila homoeotic genes control the formation of external morphological features of the embryo and adult, and in addition affect differentiation of the nervous system. Here we describe the morphogenetic events in the midgut that are controlled by the homoeotic genes Sex combs reduced (Scr) and Antennapedia (Antp). The midgut is composed of two cell layers, an inner endoderm and an outer visceral mesoderm that surround the yolk. Scr and Antp are expressed in the visceral mesoderm but not in the endoderm. The two genes are required for different aspects of the midgut morphogenesis. In Scr null mutant embryos the gastric caeca fail to form. Scr is expressed in the visceral mesoderm cells posterior to the primordia of the gastric caeca and appears to be indirectly required for the formation of the caeca. Antp is expressed in visceral mesoderm cells that overlie a part of the midgut where a constriction will form, and Antp null mutant embryos fail to form this constriction. An ultrastructural analysis of the midgut reveals that the visceral mesoderm imposes the constriction on the endoderm and the yolk. The mesodermal tissue contracts within the constriction and thereby penetrates the layer of the midgut endoderm. Microtubules participate in the morphological changes of the visceral mesoderm cells. The analysis of the expression of Scr in Antp mutant embryos revealed a case of tissue-specific regulation of Scr expression by Antp. In the epidermis, Antp has been shown to negatively regulate Scr, but it positively regulates Scr in the visceral mesoderm.     

Spatial expression of Drosophila glutathione S-transferase-D1 in the alimentary canal is regulated by the overlying visceral mesoderm

Regional gene expression within Drosophila gut epithelium is regulated by the homeotic genes expressed in the overlying visceral mesoderm. Here it is reported that Glutathione S-transferase-D1 (Gst-D1) had three distinctive expression domains in the gut epithelia: the inner epithelium of the proventriculus, the anterior border of the hindgut epithelium, and the midgut epithelium. Gst-D1 expression in the midgut epithelium became restricted to the region that later formed the third midgut constriction. This spatial restriction within the midgut epithelium required abdominal-A activity in the overlying visceral mesoderm, suggesting that Gst-D1 will be a useful marker to analyze the mechanism of gene regulation across the mesoderm and endoderm.     

Homeotic gene expression in the visceral mesoderm of Drosophila embryos.

The visceral mesoderm adhering to the midgut constitutes an internal germ layer of the Drosophila embryo that stretches along most of the anteroposterior axis (parasegment 2-13). Most cells of the midgut visceral mesoderm express exclusively one of five homeotic genes. Three of these genes, Antennapedia, Ultrabithorax and abdominal-A are active in parasegmental domains characteristic for this germ layer as they are nonoverlapping and adjacent. The common boundaries between these domains depend on mutual regulatory interactions between the three genes. The same genes function to control gut morphogenesis. Two further homeotic genes Sex combs reduced and Abdominal-B are expressed at both ends of the midgut visceral mesoderm, although absence of their expression does not appear to affect gut morphogenesis. There are no regulatory interactions between these two and the other homeotic genes. As a rule, the anterior limit of each homeotic gene domain in the visceral mesoderm is shifted posteriorly by one parasegment compared to the ectoderm. The domains result from a set of regulatory processes that are distinct from the ones ruling in other germ layers.