Two-Color 3D STORM Reveals Ciliary Transition Zone Architecture and its Role in Ciliary Signaling
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The Hedgehog (Hh) signaling pathway differentially utilizes the primary cilium in mammals and fruit flies. Recent work, including a study in BMC Biology, demonstrates that Hh signals through the cilium in zebrafish, clarifying the evolution of Hh signal transduction. See research article: http://www.biomedcentral.com/1741-7007/8/65
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Indian hedgehog
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Cilia mediate Hedgehog (Hh) signaling in vertebrates and Hh deregulation results in several clinical manifestations, such as obesity, cognitive disabilities, developmental malformations, and various cancers. Drosophila cells are nonciliated during development, which has led to the assumption that cilia-mediated Hh signaling is restricted to vertebrates. Here, we identify and characterize a cilia-mediated Hh pathway in Drosophila olfactory sensory neurons. We demonstrate that several fundamental key aspects of the vertebrate cilia pathway, such as ciliary localization of Smoothened and the requirement of the intraflagellar transport system, are present in Drosophila. We show that Cos2 and Fused are required for the ciliary transport of Smoothened and that cilia mediate the expression of the Hh pathway target genes. Taken together, our data demonstrate that Hh signaling in Drosophila can be mediated by two pathways and that the ciliary Hh pathway is conserved from Drosophila to vertebrates.
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Hedgehog signaling is transduced at the primary cilium, but the precise mechanisms underlying this action are not clear. In this issue of Developmental Cell, Dorn et al., 2012Dorn K.V. Hughes C.E. Rohatgi R. Dev. Cell. 2012; 23 (this issue): 823-835Abstract Full Text Full Text PDF PubMed Scopus (127) Google Scholar describe a novel mechanism for control of Hedgehog signaling by Evc proteins within the primary cilium. Hedgehog signaling is transduced at the primary cilium, but the precise mechanisms underlying this action are not clear. In this issue of Developmental Cell, Dorn et al., 2012Dorn K.V. Hughes C.E. Rohatgi R. Dev. Cell. 2012; 23 (this issue): 823-835Abstract Full Text Full Text PDF PubMed Scopus (127) Google Scholar describe a novel mechanism for control of Hedgehog signaling by Evc proteins within the primary cilium. The Hedgehog (Hh) signaling pathway has received enormous attention over the years because it plays an essential role in many aspects of vertebrate embryonic development and tumorigenesis (reviewed in Ingham et al., 2011Ingham P.W. Nakano Y. Seger C. Nat. Rev. Genet. 2011; 12: 393-406Crossref PubMed Scopus (449) Google Scholar). Despite this focus, the biochemical steps leading from ligand binding to target gene activation are far from clear. Roughly a decade ago, genetic experiments revealed the requirement of the primary cilium, a microtubule-based, membrane-enclosed structure, for mammalian Hh signaling. Subsequent work indicated that this small yet nearly ubiquitous “organelle” represents a central processing center where Hh signaling information is transduced. Indeed, most of the core factors regulating the pathway, including a transmembrane activator lying upstream in the pathway, Smoothened (Smo), and the pathway effectors, the Gli transcription factors, localize to primary cilia in a manner gated by pathway activity. Current models posit that activated Smo localizes to ciliary membranes in response to ligands, where it then suppresses Protein Kinase A (PKA)-mediated phosphorylation of the Gli proteins and induces dissociation of Gli proteins from their inhibitor, Suppressor of Fused (SuFu). However, how each of these events occurs at the biochemical level within the cilium is unknown. To begin to tackle this problem, Dorn et al., 2012Dorn K.V. Hughes C.E. Rohatgi R. Dev. Cell. 2012; 23 (this issue): 823-835Abstract Full Text Full Text PDF PubMed Scopus (127) Google Scholar have focused their attention on the transmembrane protein Evc2, which is mutated in two human disorders, Ellis–van Creveld syndrome (EvC) and Weyers acrodental dystosis (Weyers) (Ruiz-Perez et al., 2000Ruiz-Perez V.L. Ide S.E. Strom T.M. Lorenz B. Wilson D. Woods K. King L. Francomano C. Freisinger P. Spranger S. et al.Nat. Genet. 2000; 24: 283-286Crossref PubMed Scopus (247) Google Scholar). EvC and Weyers are classified as “ciliopathies,” which encompass a spectrum of related disorders caused by defective ciliary structure or function. EvC is an autosomal-recessive disorder in which patients harbor loss-of-function mutations in Evc2, the neighboring Evc2-interacting protein Evc, or, in rare cases, both proteins. Weyers is an autosomal-dominant disorder with phenotypic characteristics similar to EvC, and patients with this disease harbor deletions in the C terminus of Evc2. The fact that at least some of the phenotypic features are linked to dysregulation of the Hh pathway led to previous studies showing that mammalian Hh signaling requires Evc proteins and that Evc and Evc2 localize near the base of primary cilia (Blair et al., 2011Blair H.J. Tompson S. Liu Y.N. Campbell J. MacArthur K. Ponting C.P. Ruiz-Perez V.L. Goodship J.A. BMC Biol. 2011; 9: 14Crossref PubMed Scopus (64) Google Scholar). Interestingly, the human and mouse mutant phenotypes and evc expression patterns suggest that this requirement is tissue specific. In the present work, Dorn et al. show that Evc2, in a complex with Evc, physically associates with Smo in the cilium in response to pathway activation. These findings are backed up by similar results from Jin Jiang and colleagues (Yang et al., 2012Yang C. Chen W. Chen Y. Jiang J. Cell Res. 2012; (Published online September 18, 2012)https://doi.org/10.1038/cr.2012.134Crossref Scopus (72) Google Scholar). While Evc/Evc2 are required for cilium-dependent Hh signaling, they do not appear to control ciliary assembly or structure. Epistasis analysis indicates that these proteins act in the Hh pathway at a step upstream of the Gli proteins and the negative regulators SuFu and PKA. Disruption of Evc2 function does not affect the ability of Smo to localize to the cilium upon ligand stimulation but does interfere with steps downstream of Smo, such as suppression of Gli proteolytic processing, recruitment of Gli proteins and SuFu to the tips of cilia, and Gli nuclear localization. These and other data raise the possibility that, through their association with Smo, the Evc proteins provide a link between active ciliary Smo and downstream events of the pathway. Both Evc2 and Evc localize to a novel membrane subdomain within the cilium lying just distal to the ciliary transition zone, which the authors name the “EvC zone.” These findings suggest that the EvC zone is the critical location in which Smo acts to control the Hh pathway in the cilium. Restricting Evc2 to the EvC zone requires the so-called W peptide, deleted in mutant Evc2 variants of Weyers patients. Consistent with the genetics, expression of Evc2 lacking this peptide (Evc2ΔW) leads to its mislocalization throughout the cilium and dominant suppression of Hh signaling. Evc2ΔW may inhibit the pathway by binding Smo without the ability of endogenous Evc2 to homomultimerize. Interestingly, the W peptide appears to be sufficient to restrict localization of a heterologous protein, Smo, to the EvC zone. In support of the hypothesis that the EvC zone is the site of Smo action, enriching Smo within this zone increases its activity in response to pathway stimulation. The work raises a number of new questions and ideas. The most important of these is the precise nature of Evc2 biochemical function. While the data provide a good argument for the importance of the Evc2-Smo interaction in mediating Hh signaling, they are consistent with either of two modes of regulation: Evc2 may transduce information from active Smo to factors controlling Gli function, or Evc2 may allow Smo to adopt a fully active state once it has localized to the cilium. Importantly, Evc2 does not appear to directly interact with Gli proteins, PKA, or SuFu, so if the first mode applies, the nature of Gli regulation is likely to be indirect and may require the identification of additional Evc2-interacting proteins that control Hh signaling. Because previous work has extensively characterized the changes in Smo conformation in response to Hh signaling (Zhao et al., 2007Zhao Y. Tong C. Jiang J. Nature. 2007; 450: 252-258Crossref PubMed Scopus (229) Google Scholar), this type of analysis may be used to determine whether Evc proteins are essential for a late step in the pathway leading to formation of active ciliary Smo. Another interesting result from this study is the suggestion that although Smo, Sufu, and Gli proteins primarily colocalize at the tip of the cilium, active Smo may only be functional in the EvC zone. If this idea is correct, Gli proteins may be subject to Smo regulation during their transport from the base to the tips of cilia. Another important question that remains is how the Hh pathway is regulated in tissues such as the nervous system, where Evc and Evc2 appear to be dispensable. This point raises the intriguing possibility that there is no single, canonical Hh pathway acting in mammals but, rather, that cells make use of both core and tissue-specific machinery to transduce Hh signaling information depending on context. A Smoothened-Evc2 Complex Transduces the Hedgehog Signal at Primary CiliaDorn et al.Developmental CellSeptember 13, 2012In BriefSmoothened transduces Hedgehog pathway signals at primary cilia. Dorn et al. show that this key signaling event requires a Smoothened-Evc2 signaling complex at the base of the cilium. Disruption of this complex blocks Hedgehog signaling and causes birth defects, suggesting a strategy for targeting this pathway in cancer and regeneration. Full-Text PDF Open Archive
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Abstract Recently, it has become clear that the developmental hedgehog pathway is activated in ischaemic adult tissue where it aids in salvaging damaged tissue. The exact driving force for the initial hedgehog response is unclear and as most physiological and cellular processes are disturbed in ischaemic tissue, hedgehog‐activating signals are hard to dissect. Here, we demonstrate that hypoxia per se is able to induce a rapid systemic hedgehog response in adult mice, as evident from expression of the pathway ligand, Sonic hedgehog, as well as the pathway activity marker Patched1 in various organs. Using in vitro models of hypoxia, we showed that the hedgehog response was transient and preceded by the accumulation of HIF‐1α, which we hypothesized to communicate between hypoxia and hedgehog expression. Indeed, pharmacological inhibition, knockdown or genetic ablation of HIF‐1α abolished hedgehog pathway activation. In conclusion, we have established that hypoxia is translated into a hedgehog response through HIF‐1α and this mechanism is likely to be responsible for the hedgehog response observed in various ischaemia models.
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The control and mediation of the cell cycle is influenced by cell signals. Different types of cell signaling molecules: Proteins (growth factors), peptide hormones, amino acids, steroids, retenoids, fatty acid derivatives, and small gases can all act as signaling molecules. The hedgehog signaling pathway is one of the key regulators of animal development conserved from flies (wing development in Drosophila) to humans (development of the brain, GI tract, fingers and toes in mammals). The pathway name is from its polypeptide ligand, an intercellular signaling molecule called Hedgehog (Hh) found in fruit flies of the genus Drosophila. Mutations or other sorts of regulatory errors in the hedgehog pathway are associated with a number of birth defects as well as some cancers. In this report, we will review the role of Hedgehog in cell signaling and impact of it in clinical medicine.
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The vertebrate hedgehog pathway is organized in primary cilia, and hedgehog components relocate into or out of cilia during signaling. Defects in intraflagellar transport (IFT) typically disrupt ciliary assembly and attenuate hedgehog signaling. Determining whether IFT drives the movement of hedgehog components is difficult due to the requirement of IFT for building cilia. Unlike most IFT proteins, IFT27 is dispensable for cilia formation but affects hedgehog signaling similarly to other IFTs, allowing us to examine its role in the dynamics of signaling. Activating signaling at points along the pathway in Ift27 mutant cells showed that IFT is extensively involved in the pathway. Similar analysis of Bbs mutant cells showed that BBS proteins participate at many levels of signaling but are not needed to concentrate Gli transcription factors at the ciliary tip. Our analysis showed that smoothened delivery to cilia does not require IFT27, but the role of other IFTs is not known. Using a rapamycin-induced dimerization system to sequester IFT-B proteins at the mitochondria in cells with fully formed cilia did not affect the delivery of Smo to cilia, suggesting that this membrane protein may not require IFT-B for delivery.
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Fiona Bangs and Kathryn V. Anderson Developmental Biology Program, Sloan Kettering Institute, Memorial Sloan Kettering Cancer Center, New York, New York 10065 Correspondence: k-anderson{at}ski.mskcc.org
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