Non-cell autonomous control of apoptosis by ligand-independent Hedgehog signaling in Drosophila

Hedgehog (Hh) signaling is important for development and homeostasis in vertebrates and invertebrates. Ligand-independent, deregulated Hh signaling caused by loss of negative regulators such as Patched causes excessive cell proliferation, leading to overgrowth in Drosophila and tumors in humans, including basal-cell carcinoma and medulloblastoma. We show that in Drosophila deregulated Hh signaling also promotes cell survival by increasing the resistance to apoptosis. Surprisingly, cells with deregulated Hh activity do not protect themselves from apoptosis; instead, they promote cell survival of neighboring wild-type cells. This non-cell autonomous effect is mediated by Hh-induced Notch signaling, which elevates the protein levels of Drosophila inhibitor of apoptosis protein-1 (Diap-1), conferring resistance to apoptosis. In summary, we demonstrate that deregulated Hh signaling not only promotes proliferation but also cell survival of neighboring cells. This non-cell autonomous control of apoptosis highlights an underappreciated function of deregulated Hh signaling, which may help to generate a supportive micro-environment for tumor development.     

Notch signaling activates Yorkie non-cell autonomously in Drosophila

In Drosophila imaginal epithelia, cells mutant for the endocytic neoplastic tumor suppressor gene vps25 stimulate nearby untransformed cells to express Drosophila Inhibitor-of-Apoptosis-Protein-1 (DIAP-1), conferring resistance to apoptosis non-cell autonomously. Here, we show that the non-cell autonomous induction of DIAP-1 is mediated by Yorkie, the conserved downstream effector of Hippo signaling. The non-cell autonomous induction of Yorkie is due to Notch signaling from vps25 mutant cells. Moreover, activated Notch in normal cells is sufficient to induce non-cell autonomous Yorkie activity in wing imaginal discs. Our data identify a novel mechanism by which Notch promotes cell survival non-cell autonomously and by which neoplastic tumor cells generate a supportive microenvironment for tumor growth.     

Hedgehogs as Negative Regulators of the Cell Cycle

During the development of multicellular animals, cell proliferation must be precisely controlled, as deregulated proliferation can lead to overgrowth and cancer. In addition, proliferation must be tightly integrated with pattern formation and differentiation to generate the required number of cells in the right organs, and at the right time. All major signaling pathways employed during embryogenesis have been implicated in cell cycle regulation, indicating that no single pathway has been dedicated to this task. Also, the precise role of a particular signaling pathway in regulating proliferation is highly dependent on the cellular context, and may have opposite effects on cell-cycle progression in different cells and tissues. The Hedgehog (Hh) family of signaling proteins is known to control both differentiation and proliferation during development. So far, studies addressing the effect of Hh signaling on proliferation have shown it to have a stimulatory effect on cell-cycle progression. Here we review several recent studies indicating that Hh signaling can also have the opposite effect, directing cell-cycle exit in a number of cell types in vertebrate and in invertebrate embryos.     

The role of Wingless signaling in establishing the anteroposterior and dorsoventral axes of the eye disc

The posteriorly expressed signaling molecules Hedgehog and Decapentaplegic drive photoreceptor differentiation in the Drosophila eye disc, while at the anterior lateral margins Wingless expression blocks ectopic differentiation. We show here that mutations in axin prevent photoreceptor differentiation and lead to tissue overgrowth and that both these effects are due to ectopic activation of the Wingless pathway. In addition, ectopic Wingless signaling causes posterior cells to take on an anterior identity, reorienting the direction of morphogenetic furrow progression in neighboring wild-type cells. We also show that signaling by Decapentaplegic and Hedgehog normally blocks the posterior expression of anterior markers such as Eyeless. Wingless signaling is not required to maintain anterior Eyeless expression and in combination with Decapentaplegic signaling can promote its downregulation, suggesting that additional molecules contribute to anterior identity. Along the dorsoventral axis of the eye disc, Wingless signaling is sufficient to promote dorsal expression of the Iroquois gene mirror, even in the absence of the upstream factor pannier. However, Wingless signaling does not lead to ventral mirror expression, implying the existence of ventral repressors.     

Hedgehog信号通路与脂肪形成

Hedgehog(Hh)信号通路是从果蝇到人类都非常保守的信号通路,在脊椎动物和非脊椎动物胚胎期多种组织器官的发育中发挥着重要作用。Hh信号通路的异常会导致疾病(先天性缺陷和癌症)的发生。近年的研究发现,Hh信号通路在脂肪生长发育中发挥重要作用,激活Hh信号通路能特异性地抑制白色脂肪组织细胞的分化,而对棕色脂肪组织细胞分化没有作用。该文综述了Hh信号通路在脂肪细胞分化中的作用及其分子机制,并对今后的研究和应用作了展望。     

Distinct mechanisms of apoptosis-induced compensatory proliferation in proliferating and differentiating tissues in the Drosophila eye

In multicellular organisms, apoptotic cells induce compensatory proliferation of neighboring cells to maintain tissue homeostasis. In the Drosophila wing imaginal disc, dying cells trigger compensatory proliferation through secretion of the mitogens Decapentaplegic (Dpp) and Wingless (Wg). This process is under control of the initiator caspase Dronc, but not effector caspases. Here we show that a second mechanism of apoptosis-induced compensatory proliferation exists. This mechanism is dependent on effector caspases which trigger the activation of Hedgehog (Hh) signaling for compensatory proliferation. Furthermore, whereas Dpp and Wg signaling is preferentially employed in apoptotic proliferating tissues, Hh signaling is activated in differentiating eye tissues. Interestingly, effector caspases in photoreceptor neurons stimulate Hh signaling which triggers cell-cycle reentry of cells that had previously exited the cell cycle. In summary, dependent on the developmental potential of the affected tissue, different caspases trigger distinct forms of compensatory proliferation in an apparent nonapoptotic function.     

Suppression of Hedgehog signaling by Cul3 ligases in proliferation control of retinal precursors

Cullin-RING ubiquitin ligases ubiquitinate protein substrates and control their levels through degradation. Here we show that cullin3 (Cul3) suppresses Hedgehog (Hh) signaling through downregulating the level of the signaling pathway effector cubitus interruptus (Ci). High-level Hh signaling promotes Cul3-dependent Ci degradation, leading to the downregulation of Hh signaling. This process is manifested in controlling cell proliferation during Drosophila retinal development. In Cul3 mutants, the population of interommatidial cells is increased, which can be mimicked by overexpression of Ci and suppressed by depleting endogenous Ci. Hh also regulates the population of interommatidial cells in the pupal stage. Alterations in the interommatidial cell population correlate with alterations in precursor proliferation in the second mitotic wave of larval eye discs. Taken together, these results suggest that Cul3 downregulates Ci levels to modulate Hh signaling activity, thus ensuring proper cell proliferation during retinal development.     

Hedgehog signaling in the Drosophila eye and head: an analysis of the effects of different patched trans-heterozygotes

Characterization of different alleles of the Hedgehog receptor patched (ptc) indicates that they can be grouped into several classes. Most mutations result in complete loss of Ptc function. However, missense mutations located within the putative sterol-sensing domain (SSD) or C terminus of ptc encode antimorphic proteins that are unable to repress Smo activity and inhibit wild-type Ptc from doing so, but retain the ability to bind and sequester Hh. Analysis of the eye and head phenotypes of Drosophila melanogaster in various ptc/ptc(tuf1) heteroallelic combinations shows that these two classes of ptc allele can be easily distinguished by their eye phenotype, but not by their head phenotype. Adult eye size is inversely correlated with head vertex size, suggesting an alteration of cell fate within the eye-antennal disc. A balance between excess cell division and cell death in the mutant eye discs may also contribute to final eye size. In addition, contrary to results reported recently, the role of Hh signaling in the Drosophila head vertex appears to be primarily in patterning rather than in proliferation, with Ptc and Smo having opposing effects on formation of medial structures.     

Understanding human cancer using Drosophila: Tid47, a cytosolic product of the DnaJ-like tumor suppressor gene l2Tid,is a novel molecular partner of patched related to skin cancer

Recessive mutations of the Drosophila gene lethal(2)-tumorous imaginal discs (l(2)tid) cause neoplastic growth of the anlagen of the adult organs, the imaginal discs. Here we report that the three proteins encoded by this evolutionarily conserved gene, Tid50, Tid47, and Tid40, identified as members of the DnaJ cochaperone family, are destined for different cellular compartments, build complexes with many proteins in a developmental stage-specific manner, and are likely to be involved in different cellular processes. We show that the cytosolic Tid47 molecule is a novel component of the Hedgehog (Hh)-Patched (Ptc) signaling regulating cell/tissue polarity and spatial patterning during development and is associated with human tumors such as basal cell carcinoma (BCC) and medulloblastoma. We provide functional evidence for its direct in vivo interaction with the Hh-bound Ptc receptor during signal transmission. Because loss of l(2)tid causes neoplastic transformation of Hh-responsive cells, we suggest that Tid47 may at least act as a guardian of the Hh signaling gradient by regulating Ptc homeostasis in the tissue. Finally, we show that the expression of htid-1, the human counterpart of l(2)tid, is altered in human BCCs. We demonstrate that in BCCs loss of htid expression correlates with loss of differentiation capacity of the neoplastic cells similar to that found in the Drosophila tumor model.     

Cell-sorting at the A/P boundary in the Drosophila wing primordium: a computational model to consolidate observed non-local effects of Hh signaling

Non-intermingling, adjacent populations of cells define compartment boundaries; such boundaries are often essential for the positioning and the maintenance of tissue-organizers during growth. In the developing wing primordium of Drosophila melanogaster, signaling by the secreted protein Hedgehog (Hh) is required for compartment boundary maintenance. However, the precise mechanism of Hh input remains poorly understood. Here, we combine experimental observations of perturbed Hh signaling with computer simulations of cellular behavior, and connect physical properties of cells to their Hh signaling status. We find that experimental disruption of Hh signaling has observable effects on cell sorting surprisingly far from the compartment boundary, which is in contrast to a previous model that confines Hh influence to the compartment boundary itself. We have recapitulated our experimental observations by simulations of Hh diffusion and transduction coupled to mechanical tension along cell-to-cell contact surfaces. Intriguingly, the best results were obtained under the assumption that Hh signaling cannot alter the overall tension force of the cell, but will merely re-distribute it locally inside the cell, relative to the signaling status of neighboring cells. Our results suggest a scenario in which homotypic interactions of a putative Hh target molecule at the cell surface are converted into a mechanical force. Such a scenario could explain why the mechanical output of Hh signaling appears to be confined to the compartment boundary, despite the longer range of the Hh molecule itself. Our study is the first to couple a cellular vertex model describing mechanical properties of cells in a growing tissue, to an explicit model of an entire signaling pathway, including a freely diffusible component. We discuss potential applications and challenges of such an approach.     

Splitting the Hedgehog signal: sex and patterning in Drosophila

Sex-lethal (Sxl), the Drosophila sex-determination master switch, is on in females and controls sexual development as a splicing and translational regulator. Hedgehog (Hh) is a secreted protein that specifies cell fate during development. Previous work has demonstrated that Sxl protein is part of the Hh cytoplasmic signaling complex and that Hh promotes Sxl nuclear entry. In the wing disc anterior compartment, Patched (Ptc), the Hh receptor, acts positively in this process. Here, it is shown that the levels and rate of nuclear entry of full-length Cubitus interruptus (Ci), the Hh signaling target, are enhanced by Sxl. This effect requires the cholesterol but not palmitoyl modification on Hh, and expands the zone of full-length Ci expression. Expansion of Ci activation and its downstream targets, particularly decapentaplegic the Drosophila TGFbeta homolog, suggests a mechanism for generating different body sizes in the sexes; in Drosophila, females are larger and this difference is controlled by Sxl. Consistent with this proposal, discs expressing ectopic Sxl show an increase in growth. In keeping with the idea of the involvement of a signaling system, this growth effect by Sxl is not cell autonomous. These results have implications for all organisms that are sexually dimorphic and use Hh for patterning.     

Proliferation of Murine Midbrain Neural Stem Cells Depends upon an Endogenous Sonic Hedgehog (Shh) Source

The Sonic Hedgehog (Shh) pathway is responsible for critical patterning events early in development and for regulating the delicate balance between proliferation and differentiation in the developing and adult vertebrate brain. Currently, our knowledge of the potential role of Shh in regulating neural stem cells (NSC) is largely derived from analyses of the mammalian forebrain, but for dorsal midbrain development it is mostly unknown. For a detailed understanding of the role of Shh pathway for midbrain development in vivo, we took advantage of mouse embryos with cell autonomously activated Hedgehog (Hh) signaling in a conditional Patched 1 (Ptc1) mutant mouse model. This animal model shows an extensive embryonic tectal hypertrophy as a result of Hh pathway activation. In order to reveal the cellular and molecular origin of this in vivo phenotype, we established a novel culture system to evaluate neurospheres (nsps) viability, proliferation and differentiation. By recreating the three-dimensional (3-D) microenvironment we highlight the pivotal role of endogenous Shh in maintaining the stem cell potential of tectal radial glial cells (RGC) and progenitors by modulating their Ptc1 expression. We demonstrate that during late embryogenesis Shh enhances proliferation of NSC, whereas blockage of endogenous Shh signaling using cyclopamine, a potent Hh pathway inhibitor, produces the opposite effect. We propose that canonical Shh signaling plays a central role in the control of NSC behavior in the developing dorsal midbrain by acting as a niche factor by partially mediating the response of NSC to epidermal growth factor (EGF) and fibroblast growth factor (FGF) signaling. We conclude that endogenous Shh signaling is a critical mechanism regulating the proliferation of stem cell lineages in the embryonic dorsal tissue.