Rulang Jiang

The Mouse Snail Gene Encodes a Key Regulator of the Epithelial-Mesenchymal Transition

ABSTRACT Snail family genes encode DNA binding zinc finger proteins that act as transcriptional repressors. Mouse embryos deficient for the Snail (Sna) gene exhibit defects in the formation of the mesoderm germ layer. In Sna −/− mutant embryos, a mesoderm layer forms and mesodermal marker genes are induced but the mutant mesoderm is morphologically abnormal. Lacunae form within the mesoderm layer of the mutant embryos, and cells lining these lacunae retain epithelial characteristics. These cells resemble a columnar epithelium and have apical-basal polarity, with microvilli along the apical surface and intercellular electron-dense adhesive junctions that resemble adherens junctions. E-cadherin expression is retained in the mesoderm of the Sna −/− embryos. These defects are strikingly similar to the gastrulation defects observed insnail-deficient Drosophila embryos, suggesting that the mechanism of repression of E-cadherin transcription by Snail family proteins may have been present in the metazoan ancestor of the arthropod and mammalian lineages.

Notch signalling pathway mediates hair cell development in mammalian cochlea

The mammalian cochlea contains an invariant mosaic of sensory hair cells and non-sensory supporting cells reminiscent of invertebrate structures such as the compound eye in Drosophila melanogaster. The sensory epithelium in the mammalian cochlea (the organ of Corti) contains four rows of mechanosensory hair cells: a single row of inner hair cells and three rows of outer hair cells. Each hair cell is separated from the next by an interceding supporting cell, forming an invariant and alternating mosaic that extends the length of the cochlear duct. Previous results suggest that determination of cell fates in the cochlear mosaic occurs via inhibitory interactions between adjacent progenitor cells (lateral inhibition). Cells populating the cochlear epithelium appear to constitute a developmental equivalence group in which developing hair cells suppress differentiation in their immediate neighbours through lateral inhibition. These interactions may be mediated through the Notch signalling pathway, a molecular mechanism that is involved in the determination of a variety of cell fates. Here we show that genes encoding the receptor protein Notch1 and its ligand, Jagged 2, are expressed in alternating cell types in the developing sensory epithelium. In addition, genetic deletion of Jag2 results in a significant increase in sensory hair cells, presumably as a result of a decrease in Notch activation. These results provide direct evidence for Notch-mediated lateral inhibition in a mammalian system and support a role for Notch in the development of the cochlear mosaic.

The slug gene is not essential for mesoderm or neural crest development in mice

The Slug gene encodes a zinc finger protein, homologous to the product of the Drosophila Snail gene, that is implicated in the generation and migration of both mesoderm and neural crest cells in several vertebrate species. We describe here the cloning and genetic analysis of the mouse Slug (Slugh) gene. Slugh encodes a 269-amino-acid protein the shares 92% amino acid identity with the product of the chicken Slug gene. We have characterized Slugh gene expression during early mouse embryogenesis by whole mount in situ hybridization of Slugh mRNA and through detection of beta-galactosidase expression from an in-frame SlughIacZ allele generated through homologous recombination. Slugh expression is first detected in extraembryonic mesoderm and is later detected in many mesodermal subsets, although it is not detected in the primitive streak. In contrast to many other vertebrates, the mouse Slug gene is not expressed in premigratory neural crest cells but is expressed in migratory neural crest cells. Analysis of a targeted null mutation that deleted all Slugh coding sequences revealed that Slugh is not required for mesoderm formation or for neural crest generation, migration, or development in mice. These results indicate that neither the expression pattern nor the biological function of the Slug gene is conserved among all vertebrates. These data also raise interesting questions about the regulation of neural crest generation, which is one of the distinguishing characteristics of the vertebrate subphylum.

Development of the upper lip: Morphogenetic and molecular mechanisms

The vertebrate upper lip forms from initially freely projecting maxillary, medial nasal, and lateral nasal prominences at the rostral and lateral boundaries of the primitive oral cavity. These facial prominences arise during early embryogenesis from ventrally migrating neural crest cells in combination with the head ectoderm and mesoderm and undergo directed growth and expansion around the nasal pits to actively fuse with each other. Initial fusion is between lateral and medial nasal processes and is followed by fusion between maxillary and medial nasal processes. Fusion between these prominences involves active epithelial filopodial and adhering interactions as well as programmed cell death. Slight defects in growth and patterning of the facial mesenchyme or epithelial fusion result in cleft lip with or without cleft palate, the most common and disfiguring craniofacial birth defect. Recent studies of craniofacial development in animal models have identified components of several major signaling pathways, including Bmp, Fgf, Shh, and Wnt signaling, that are critical for proper midfacial morphogenesis and/or lip fusion. There is also accumulating evidence that these signaling pathways cross‐regulate genetically as well as crosstalk intracellularly to control cell proliferation and tissue patterning. This review will summarize the current understanding of the basic morphogenetic processes and molecular mechanisms underlying upper lip development and discuss the complex interactions of the various signaling pathways and challenges for understanding cleft lip pathogenesis. Developmental Dynamics 235:1152–1166, 2006.

Palatogenesis: morphogenetic and molecular mechanisms of secondary palate development

Mammalian palatogenesis is a highly regulated morphogenetic process during which the embryonic primary and secondary palatal shelves develop as outgrowths from the medial nasal and maxillary prominences, respectively, remodel and fuse to form the intact roof of the oral cavity. The complexity of control of palatogenesis is reflected by the common occurrence of cleft palate in humans. Although the embryology of the palate has long been studied, the past decade has brought substantial new knowledge of the genetic control of secondary palate development. Here, we review major advances in the understanding of the morphogenetic and molecular mechanisms controlling palatal shelf growth, elevation, adhesion and fusion, and palatal bone formation.

Odd-skipped related 1 is required for development of the metanephric kidney and regulates formation and differentiation of kidney precursor cells

Formation of kidney tissue requires the generation of kidney precursor cells and their subsequent differentiation into nephrons, the functional filtration unit of the kidney. Here we report that the gene odd-skipped related 1 (Odd1) plays an important role in both these processes. Odd1 is the earliest known marker of the intermediate mesoderm, the precursor to all kidney tissue. It is localized to mesenchymal precursors within the mesonephric and metanephric kidney and is subsequently downregulated upon tubule differentiation. Mice lacking Odd1 do not form metanephric mesenchyme, and do not express several other factors required for metanephric kidney formation, including Eya1, Six2, Pax2, Sall1 and Gdnf. In transient ectopic expression experiments in the chick embryo, Odd1 can promote expression of the mesonephric precursor markers Pax2 and Lim1. Finally, persistent expression of Odd1 in chick mesonephric precursor cells inhibits differentiation of these precursors into kidney tubules. These data indicate that Odd1 plays an important role in establishing kidney precursor cells, and in regulating their differentiation into kidney tubular tissue.

Wnt/beta-catenin signaling plays an essential role in activation of odontogenic mesenchyme during early tooth development.

Classical tissue recombination studies demonstrated that initiation of tooth development depends on activation of odontogenic potential in the mesenchyme by signals from the presumptive dental epithelium. Although several members of the Wnt family of signaling molecules are expressed in the presumptive dental epithelium at the beginning of tooth initiation, whether Wnt signaling is directly involved in the activation of the odontogenic mesenchyme has not been characterized. In this report, we show that tissue-specific inactivation of beta-catenin, a central component of the canonical Wnt signaling pathway, in the developing tooth mesenchyme caused tooth developmental arrest at the bud stage in mice. We show that mesenchymal beta-catenin function is required for expression of Lef1 and Fgf3 in the developing tooth mesenchyme and for induction of primary enamel knot in the developing tooth epithelium. Expression of Msx1 and Pax9, two essential tooth mesenchyme transcription factors downstream of Bmp and Fgf signaling, respectively, were not altered in the absence of beta-catenin in the tooth mesenchyme. Moreover, we found that constitutive stabilization of beta-catenin in the developing palatal mesenchyme induced aberrant palatal epithelial invaginations that resembled early tooth buds both morphologically and in epithelial molecular marker expression, but without activating expression of Msx1 and Pax9 in the mesenchyme. Together, these results indicate that activation of the mesenchymal odontogenic program during early tooth development requires concerted actions of Bmp, Fgf and Wnt signaling from the presumptive dental epithelium to the mesenchyme.

Odd-skipped related 2 (Osr2) encodes a key intrinsic regulator of secondary palate growth and morphogenesis

Development of the mammalian secondary palate involves multiple steps of highly regulated morphogenetic processes that are frequently disturbed during human development, resulting in the common birth defect of cleft palate. Neither the molecular processes governing normal palatogenesis nor the causes of cleft palate is well understood. In an expression screen to identify new transcription factors regulating palate development, we previously isolated the odd-skipped related 2 (Osr2) gene, encoding a zinc-finger protein homologous to the Drosophila odd-skipped gene product, and showed that Osr2 mRNA expression is specifically activated in the nascent palatal mesenchyme at the onset of palatal outgrowth. We report that a targeted null mutation in Osr2 impairs palatal shelf growth and causes delay in palatal shelf elevation, resulting in cleft palate. Whereas palatal outgrowth initiates normally in the Osr2 mutant embryos, a significant reduction in palatal mesenchyme proliferation occurs specifically in the medial halves of the downward growing palatal shelves at E13.5, which results in retarded, mediolaterally symmetric palatal shelves before palatal shelf elevation. The developmental timing of palatal growth retardation correlates exactly with the spatiotemporal pattern of Osr1 gene expression during palate development. Furthermore, we show that the Osr2 mutants exhibit altered gene expression patterns, including those of Osr1, Pax9 and Tgfb3, during palate development. These data identify Osr2 as a key intrinsic regulator of palatal growth and patterning.

Antagonistic Actions of Msx1 and Osr2 Pattern Mammalian Teeth into a Single Row

Mammals have single-rowed dentitions, whereas many nonmammalian vertebrates have teeth in multiple rows. Neither the molecular mechanism regulating iterative tooth initiation nor that restricting mammalian tooth development in one row is known. We found that mice lacking the transcription factor odd-skipped related-2 (Osr2) develop supernumerary teeth lingual to their molars because of expansion of the odontogenic field. Osr2 was expressed in a lingual-to-buccal gradient and restricted expression of bone morphogenetic protein 4 (Bmp4), an essential odontogenic signal, in the developing tooth mesenchyme. Expansion of odontogenic field in Osr2-deficient mice required Msx1, a feedback activator of Bmp4 expression. These findings suggest that the Bmp4-Msx1 pathway propagates mesenchymal activation for sequential tooth induction and that spatial modulation of this pathway provides a mechanism for patterning vertebrate dentition.

Expression of Wnt9b and activation of canonical Wnt signaling during midfacial morphogenesis in mice.

Cleft lip with or without cleft palate (CLP) is the most common craniofacial birth defect in humans. Recently, mutations in the WNT3 and Wnt9b genes, encoding two members of the Wnt family of signaling molecules, were found associated with CLP in human and mice, respectively. To investigate whether Wnt3 and Wnt9b directly regulate facial development, we analyzed their developmental expression patterns and found that both Wnt3 and Wnt9b are expressed in the facial ectoderm at critical stages of midfacial morphogenesis during mouse embryogenesis. Whereas Wnt3 mRNA is mainly expressed in the maxillary and medial nasal ectoderm, Wnt9b mRNA is expressed in maxillary, medial nasal, and lateral nasal ectoderm. During lip fusion, Wnt9b, but not Wnt3, is expressed in the epithelial seam between the fusing medial and lateral nasal processes. Furthermore, we found that expression of TOPGAL, a transgenic reporter of activation of canonical Wnt signaling pathway, is specifically activated in the distal regions of the medial nasal, lateral nasal, and maxillary processes prior to lip fusion. During lip fusion, the epithelial seam between the medial and lateral nasal processes as well as the facial mesenchyme directly beneath the fusing epithelia strongly expresses TOPGAL. These data, together with the CLP lip phenotype in WNT3−/− humans and Wnt9b−/− mutant mice, indicate that Wnt3 and Wnt9b signal through the canonical Wnt signaling pathway to regulate midfacial development and lip fusion. Developmental Dynamics, 2006.