Segment polarity genes in neuroblast formation and identity specification during Drosophila neurogenesis

The relatively simple central nervous system (CNS) of the Drosophila embryo provides a useful model system for investigating the mechanisms that generate and pattern complex nervous systems. Central to the generation of different types of neurons by precursor neuroblasts is the initial specification of neuroblast identity and the Drosophila segment polarity genes, genes that specify regions within a segment or repeating unit of the Drosophila embryo, have emerged recently as significant players in this process. During neurogenesis the segment polarity genes are expressed in the neuroectodermal cells from which neuroblasts delaminate and they continue to be expressed in neuroblasts and their progeny. Loss-of-function mutations in these genes lead to a failure in the formation of neuroblasts and/or specification of neuroblast identity. Results from several recent studies suggest that regulatory interactions between segment polarity genes during neurogenesis lead to an increase in the number of neuroblasts and specification of different identities to neuroblasts within a population of cells. BioEssays 21:472–485, 1999. © 1999 John Wiley & Sons, Inc.     

The specification of sensory neuron identity in Drosophila

Different types of sense organs are present on the larva of Drosophila. Several genes that specify the type of sense organ that will form at a particular position have been recently identified. Here we review the functional and molecular analyses of these genes, and summarize the evidence which supports a role in the choice of which type of organ will be formed. Most or all of these genes are required for the appropriate specification of adult as well as larval sense organs, suggesting that the larval and adult systems share many gene requirements. Interestingly, the specifying genes identified so far in the peripheral nervous system are also expressed in subsets of cells in the central nervous system, where they might have similar roles.     

The ladybird homeobox genes are essential for the specification of a subpopulation of neural cells

In Drosophila, neurons and glial cells are produced by neural precursor cells called neuroblasts (NBs), which can be individually identified. Each NB generates a characteristic cell lineage specified by a precise spatiotemporal control of gene expression within the NB and its progeny. Here we show that the homeobox genes ladybird early and ladybird late are expressed in subsets of cells deriving from neuroblasts NB 5-3 and NB 5-6 and are essential for their correct development. Our analysis revealed that ladybird in Drosophila, like their vertebrate orthologous Lbx1 genes, play an important role in cell fate specification processes. Among those cells that express ladybird are NB 5-6-derived glial cells. In ladybird loss-of-function mutants, the NB 5-6-derived exit glial cells are absent while overexpression of these genes leads to supernumerary glial cells of this type. Furthermore, aberrant glial cell positioning and aberrant spacing of axonal fascicles in the nerve roots observed in embryos with altered ladybird function suggest that the ladybird genes might also control directed cell movements and cell-cell interactions within the developing Drosophila ventral nerve cord.     

Cross-regulatory interactions among tracheal genes support a co-operative model for the induction of tracheal fates in the Drosophila embryo

The Drosophila tracheal system arises from clusters of ectodermal cells that invaginate and migrate to originate a network of epithelial tubes. Genetic analyses have identified several genes that are specifically expressed in the tracheal cells and are required for tracheal development. Among them, trachealess (trh) is able to induce ectopic tracheal pits and therefore it has been suggested that it would act as an inducer of tracheal cell fates; however, this capacity appears to be spatially restricted. Here we analyze the expression of the tracheal specific genes in the early steps of tracheal development and their cross-interactions. We find that there is a set of primary genes including trh and ventral veinless (vvl) whose expression does not depend on any other tracheal gene and a set of downstream genes whose expression requires different combinations of the primary genes. We also find that the combined expression of primary genes is sufficient to induce some downstream genes but not others. These results indicate that there is not a single master gene responsible for the appropriate expression of the tracheal genes and support a model where tracheal cell fates are induced by the co-operation of several factors rather than by the activity of a single tracheal inducer.     

Proximodistal domain specification and interactions in developing Drosophila appendages

The morphological diversification of appendages represents a crucial aspect of animal body plan evolution. The arthropod antenna and leg are homologous appendages, thought to have arisen via duplication and divergence of an ancestral structure (Snodgrass, R. (1935) Book Principles of Insect Morphology. New York: McGraw-Hill). To gain insight into how variations between the antenna and the leg may have arisen, we have compared the epistatic relationships among three major proximodistal patterning genes, Distal-less, dachshund and homothorax, in the antenna and leg of the insect arthropod Drosophila melanogaster. We find that Drosophila appendages are subdivided into different proximodistal domains specified by specific genes, and that limb-specific interactions between genes and the functions of these genes are crucial for antenna-leg differences. In particular, in the leg, but not in the antenna, mutually antagonistic interactions exist between the proximal and medial domains, as well as between medial and distal domains. The lack of such antagonism in the antenna leads to extensive coexpression of Distal-less and homothorax, which in turn is essential for differentiation of antennal morphology. Furthermore, we report that a fundamental difference between the two appendages is the presence in the leg and absence in the antenna of a functional medial domain specified by dachshund. Our results lead us to propose that the acquisition of particular proximodistal subdomains and the evolution of their interactions has been essential for the diversification of limb morphology.     

Bmp and Fgf signaling are essential for liver specification in zebrafish

Based on data from in vitro tissue explant and ex vivo cell/bead implantation experiments, Bmp and Fgf signaling have been proposed to regulate hepatic specification. However, genetic evidence for this hypothesis has been lacking. Here, we provide in vivo genetic evidence that Bmp and Fgf signaling are essential for hepatic specification. We utilized transgenic zebrafish that overexpress dominant-negative forms of Bmp or Fgf receptors following heat-shock induction. These transgenes allow one to bypass the early embryonic requirements for Bmp and Fgf signaling, and also to completely block Bmp or Fgf signaling. We found that the expression of hhex and prox1, the earliest liver markers in zebrafish, was severely reduced in the liver region when Bmp or Fgf signaling was blocked just before hepatic specification. However, hhex and prox1 expression in adjacent endodermal and mesodermal tissues appeared unaffected by these manipulations. Additional genetic studies indicate that the endoderm maintains competence for Bmp-mediated hepatogenesis over an extended window of embryonic development. Altogether, these data provide the first genetic evidence that Bmp and Fgf signaling are essential for hepatic specification, and suggest that endodermal cells remain competent to differentiate into hepatocytes for longer than anticipated.     

The CNS midline cells and spitz class genes are required for proper patterning of Drosophila ventral neuroectoderm.

The Drosophila embryonic central nervous system (CNS) develops from sets of neuroblasts (NBs) which segregate from the ventral neuroectoderm during early embryogenesis. It is not well established how each individual NB in the neuroectoderm acquires its characteristic identity along the dorsal-ventral axis. Since it is known that CNS midline cells and spitz class genes (pointed, rhomboid, single-minded, spitz and Star) are required for the proper patterning of ventral CNS and epidermis originated from the ventral neuroectoderm, this study was carried out to determine the functional roles of the CNS midline cells and spitz class genes in the fate determination of ventral NBs and formation of mature neurons and their axon pathways. Several molecular markers for the identified NBs, neurons, and axon pathways were employed to examine marker gene expression profile, cell lineage and axon pathway formation in the spitz class mutants. This analysis showed that the CNS midline cells specified by single-minded gene as well as spitz class genes are required for identity determination of a subset of ventral NBs and for formation of mature neurons and their axon pathways. This study suggests that the CNS midline cells and spitz class genes are necessary for proper patterning of the ventral neuroectoderm along the dorsal-ventral axis.     

Another notch in stem cell biology: Drosophila intestinal stem cells and the specification of cell fates

Previous work has suggested that many stem cells can be found in microanatomic niches, where adjacent somatic cells of the niche control the differentiation and proliferation states of their resident stem cells. Recently published work examining intestinal stem cells (ISCs) in the adult Drosophila midgut suggests a new paradigm where some stem cells actively control the cell fate decisions of their daughters. Here, we review recent literature((1)) demonstrating that, in the absence of a detectable stem cell niche, multipotent Drosophila ISCs modulate the Notch signaling pathway in their adjacent daughter cells in order to specify the differentiated lineages of their descendants. These observations made in Drosophila are challenging and advancing our understanding of stem cell biology.     

Abdominal-B is essential for proper sexually dimorphic development of the Drosophila gonad

Sexual dimorphism requires the integration of positional information in the embryo with the sex determination pathway. Homeotic genes are a major source of positional information responsible for patterning along the anterior-posterior axis in embryonic development, and are likely to play a critical role in sexual dimorphism. Here, we investigate the role of homeotic genes in the sexually dimorphic development of the gonad in Drosophila. We have found that Abdominal-B (ABD-B) is expressed in a sexually dimorphic manner in the embryonic gonad. Furthermore, Abd-B is necessary and sufficient for specification of a sexually dimorphic cell type, the male-specific somatic gonadal precursors (msSGPs). In Abd-B mutants, the msSGPs are not specified and male gonads now resemble female gonads with respect to these cells. Ectopic expression of Abd-B is sufficient to induce formation of extra msSGPs in additional segments of the embryo. Abd-B works together with abdominal-A to pattern the non-sexually dimorphic somatic gonad in both sexes, while Abd-B alone specifies the msSGPs. Our results indicate that Abd-B acts at multiple levels to regulate gonad development and that Abd-B class homeotic genes are conserved factors in establishing gonad sexual dimorphism in diverse species.     

Coordinating proliferation and tissue specification to promote regional identity in the Drosophila head

The Decapentaplegic and Notch signaling pathways are thought to direct regional specification in the Drosophila eye-antennal epithelium by controlling the expression of selector genes for the eye (Eyeless/Pax6, Eyes absent) and/or antenna (Distal-less). Here, we investigate the function of these signaling pathways in this process. We find that organ primordia formation is indeed controlled at the level of Decapentaplegic expression but critical steps in regional specification occur earlier than previously proposed. Contrary to previous findings, Notch does not specify eye field identity by promoting Eyeless expression but it influences eye primordium formation through its control of proliferation. Our analysis of Notch function reveals an important connection between proliferation, field size, and regional specification. We propose that field size modulates the interaction between the Decapentaplegic and Wingless pathways, thereby linking proliferation and patterning in eye primordium development.     

Drosophila E-cadherin is essential for proper germ cell-soma interaction during gonad morphogenesis

In most animal species, germ cells require intimate contact with specialized somatic cells in the gonad for their proper development. We have analyzed the establishment of germ cell-soma interaction during embryonic gonad formation in Drosophila melanogaster, and find that somatic cells undergo dramatic changes in cell shape and individually ensheath germ cells as the gonad coalesces. Germ cell ensheathment is independent of other aspects of gonad formation, indicating that separate morphogenic processes are at work during gonadogenesis. The cell-cell adhesion molecule Drosophila E-cadherin is essential both for germ cell ensheathment and gonad compaction, and is upregulated in the somatic gonad at the time of gonad formation. Our data indicate that differential cell adhesion contributes to cell sorting and the formation of proper gonad architecture. In addition, we find that Fear of Intimacy, a novel transmembrane protein, is also required for both germ cell ensheathment and gonad compaction. E-cadherin expression in the gonad is dramatically decreased in fear of intimacy mutants, indicating that Fear of Intimacy may be a regulator of E-cadherin expression or function.     

Otx genes are required for tissue specification in the developing eye

Patterning of the vertebrate eye appears to be controlled by the mutual regulation and the progressive restriction of the expression domains of a number of genes initially co-expressed within the eye anlage. Previous data suggest that both Otx1 and Otx2 might contribute to the establishment of the different eye territories. Here, we have analysed the ocular phenotype of mice carrying different functional copies of Otx1 and Otx2 and we show that these genes are required in a dose-dependent manner for the normal development of the eye. Thus, all Otx1(-/-); Otx2(+/-) and 30% of Otx1(+/-); Otx2(+/-) genotypes presented consistent and profound ocular malformation, including lens, pigment epithelium, neural retina and optic stalk defects. During embryonic development, optic vesicle infolding was severely altered and the expression of pigment epithelium-specific genes, such as Mitf or tyrosinase, was lost. Lack of pigment epithelium specification was associated with an expansion of the prospective neural retina and optic stalk territories, as determined by the expression of Pax6, Six3 and Pax2. Later in development the presumptive pigment epithelium region acquired features of mature neural retina, including the generation of Islet1-positive neurones. Furthermore, in Otx1(-/-); Otx2(+/-) mice neural retina cell proliferation, cell differentiation and apoptotic cell death were also severely affected. Based on these findings we propose a model in which Otx gene products are required for the determination and differentiation of the pigment epithelium, co-operating with other eye patterning genes in the determination of the specialised tissues that will constitute the mature vertebrate eye.     

Expression of evolutionarily conserved eye specification genes during Drosophila embryogenesis

Eye specification in Drosophila is thought be controlled by a set of seven nuclear factors that includes the Pax6 homolog, Eyeless. This group of genes is conserved throughout evolution and has been repeatedly recruited for eye specification. Several of these genes are expressed within the developing eyes of vertebrates and mutations in several mouse and human orthologs are the underlying causes of retinal disease syndromes. Ectopic expression in Drosophila of any one of these genes is capable of inducing retinal development, while loss-of-function mutations delete the developing eye. These nuclear factors comprise a complex regulatory network and it is thought that their combined activities are required for the formation of the eye. We examined the expression patterns of four eye specification genes, eyeless (ey), sine oculis (so), eyes absent (eya), and dachshund (dac) throughout all time points of embryogenesis and show that only eyeless is expressed within the embryonic eye anlagen. This is consistent with a recently proposed model in which the eye primordium acquires its competence to become retinal tissue over several time points of development. We also compare the expression of Ey with that of a putative antennal specifying gene Distal-less (Dll). The expression patterns described here are quite intriguing and raise the possibility that these genes have even earlier and wide ranging roles in establishing the head and visual field.     

Delta-mediated specification of midline cell fates in zebrafish embryos

BACKGROUND: Fate mapping studies have shown that progenitor cells of three vertebrate embryonic midline structures - the floorplate in the ventral neural tube, the notochord and the dorsal endoderm - occupy a common region prior to gastrulation. This common region of origin raises the possibility that interactions between midline progenitor cells are important for their specification prior to germ layer formation. RESULTS: One of four known zebrafish homologues of the Drosophila melanogaster cell-cell signaling gene Delta, deltaA (dlA), is expressed in the developing midline, where progenitor cells of the ectodermal floorplate, mesodermal notochord and dorsal endoderm lie close together before they occupy different germ layers. We used a reverse genetic strategy to isolate a missense mutation of dlA, dlAdx2, which coordinately disrupts the development of floorplate, notochord and dorsal endoderm. The dlAdx2 mutant embryos had reduced numbers of floorplate and hypochord cells; these cells lie above and beneath the notochord, respectively. In addition, mutant embryos had excess notochord cells. Expression of a dominant-negative form of Delta protein driven by mRNA microinjection produced a similar effect. In contrast, overexpression of dlA had the opposite effect: fewer trunk notochord cells and excess floorplate and hypochord cells. CONCLUSION: Our results indicate that Delta signaling is important for the specification of midline cells. The results are most consistent with the hypothesis that developmentally equivalent midline progenitor cells require Delta-mediated signaling prior to germ layer formation in order to be specified as floorplate, notochord or hypochord.     

Cell-based therapies for coronavirus disease 2019: proper clinical investigations are essential

The serious consequences of the global coronavirus disease 2019 (COVID-19) pandemic have prompted a rapid global response to develop effective therapies that can lessen disease severity in infected patients. Cell-based approaches, primarily using mesenchymal stromal cells (MSCs), have demonstrated a strong safety profile and possible efficacy in patients with acute respiratory distress syndrome (ARDS), but whether these therapies are effective for treating respiratory virus-induced ARDS is unknown. According to the World Health Organization International Clinical Trials Registry Platform and the National Institutes of Health ClinicalTrials.gov databases, 27 clinical investigations of MSC-based cell therapy approaches have begun in China since the onset of the COVID-19 outbreak, with a growing number of academic and industry trials elsewhere as well. Several recent published reports have suggested potential efficacy; however, the available data presented are either anecdotal or from incomplete, poorly controlled investigations. Therefore, although there may be a potential role for MSCs and other cell-based therapies in treatment of COVID-19, these need to be investigated in a rationally designed, controlled approach if safety and efficacy are to be demonstrated accurately. The authors urge that the field proceed by finding a balance between swift experimentation and communication of results and scientifically coherent generation and analysis of clinical data.