Structural basis for the recognition of Sonic Hedgehog by human Patched1

Gong, Xin; Qian, Hui‐Fen; Cao, Pingping; Zhao, Xin; Zhou, Qiang; Lei, Jianlin; Yan, Nieng

doi:10.1126/science.aas8935

Cited by 183 publications

(267 citation statements)

References 81 publications

(95 reference statements)

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“…The mechanism by which PTCH inhibits SMO was enigmatic for a long time because PTCH represses SMO without direct contact 3 . PTCH1 was shown to be able to transport cholesterol [4][5][6] which in turn will directly activate SMO 7 , a finding, which was supported by recent structural analyses 1,[8][9][10] . The structures suggest that Hh inhibits PTCH transporter function and hence plasma membrane cholesterol levels could increase.…”

Section: Introductionmentioning

confidence: 59%

“…Recent data suggested that mammalian PTCH1 acts as a cholesterol transporter 1,5,6,[8][9][10] . To investigate, whether PTC-3 shares the function of PTCH1 as cholesterol transporter, we first expressed PTC-3 in Saccharomyces cerevisiae, which does not contain any cholesterol 29 and measured cholesterol efflux from cells using TopFluor cholesterol in pulse-chase experiment ( Fig.…”

Section: Ptc-3 Is a Cholesterol Transportermentioning

confidence: 99%

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Patched regulates lipid homeostasis by controlling cellular cholesterol levels

Castillo

Hannich

Kaech

et al. 2019

Preprint

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words)Hedgehog (Hh) signaling is essential during development and in organ physiology. In the canonical pathway, Hh binding to Patched (PTCH) relieves the inhibition of Smoothened (SMO). Yet, PTCH may also perform SMO-independent functions. While the PTCH homolog PTC-3 is essential in C. elegans, worms lack SMO, providing an excellent model to probe non-canonical PTCH function. Here, we show that PTC-3 is a cholesterol transporter. ptc-3(RNAi) leads to accumulation of intracellular cholesterol and defects in ER structure and lipid droplet formation. These phenotypes were accompanied by a reduction in acyl chain (FA) length and desaturation. ptc-3(RNAi)induced lethality, fat storage and ER morphology defects were rescued by reducing dietary cholesterol. We provide evidence that cholesterol accumulation modulates the function of nuclear hormone receptors such as of the PPARa homolog NHR-49 and NHR-181, and affects FA composition. Our data uncover a novel role for PTCH in organelle structure maintenance and fat metabolism.No clear SMO homolog is encoded in the genome. In addition, some of the other downstream targets of the canonical Hh signaling pathway are also missing. In fact, it was proposed that SMO and those components were specifically lost during evolution in nematodes 15,21-23 . For example, SUFU is not conserved and the homolog of the transcription factor Gli, TRA-1, is involved in sex determination and gonad development in males and hermaphrodites 24 . Therefore, C. elegans provides an excellent model to study non-canonical, SMO-independent Hh signaling pathways, in particular in somatic tissues. To dissect SMO-independent PTCH functions, we concentrated on PTC-3, which is expressed in somatic tissues, in particular in the hypodermis, glia and gut 20 . We found that reduction of PTC-3 levels causes the accumulation of intracellular cholesterol and reduction in poly unsaturated fatty acids (PUFAs). Moreover, the endoplasmic reticulum lost most of its reticulate tubular form and developed elaborate sheet structures in the intestine. This effect in turn strongly impaired lipid droplet biogenesis, resulting in the inability of the animal to store fat.Reduction of dietary cholesterol rescued fat storage defects, the ER morphology defects, and improved development and survival in ptc-3(RNAi) animals. Cholesterol levels influence nuclear hormone receptor activity such as of the PPARa homolog NHR-49, which is involved in the regulation of FA synthesis. Thus, our data demonstrate that PTCH also controls intracellular cholesterol levels in C. elegans.Moreover, we show that PTCH thereby impinges on FA metabolism, organellar structure and fat storage capacity.

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Section: Introductionmentioning

confidence: 59%

Section: Ptc-3 Is a Cholesterol Transportermentioning

confidence: 99%

Patched regulates lipid homeostasis by controlling cellular cholesterol levels

Castillo

Hannich

Kaech

et al. 2019

Preprint

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“…Precedent for cholesterol passing through a related transporter comes from the structural work of Nieng Yan and Xiaochun Li and their colleagues who have recently reported structures of the related Patched (PTCH) protein that is important for Hedgehog signaling (Gong et al, 2018;Qi et al, 2018a, b). PTCH has 12 transmembrane domains instead of 13; it also lacks the cholesterol binding N-terminal domain of NPC1 and NPC1L1.…”

Section: Discussionmentioning

confidence: 99%

“…Transmembrane domains 2-6 comprise a sterol-sensing domain analogous to transmembrane domains 3-7 of NPC1. Yan and colleagues detected two sterol molecules in the PTCH structure: one in a cavity located between the two extracellular domains and the second, adjacent to the sterol-sensing domain (Gong et al, 2018). Their mutant analysis suggests that cholesterol binding causes significant conformational changes, with an untwisting of the interactions between the extracellular domains and reorientation of certain TMs.…”

Section: Discussionmentioning

confidence: 99%

Inter-domain dynamics drive cholesterol transport by NPC1 and NPC1L1 proteins

Saha

Shumate

Caldwell

et al. 2020

Preprint

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Transport of LDL-derived cholesterol from lysosomes into the cytoplasm requires NPC1 protein; NPC1L1 mediates uptake of dietary cholesterol. We introduced single disulfide bonds into NPC1 and NPC1L1 to explore the importance of inter-domain dynamics in cholesterol transport. Using a sensitive method to monitor lysosomal cholesterol efflux, we find that NPC1's N-terminal domain need not release from the rest of the protein for efficient cholesterol export. Either introducing single disulfide bonds to constrain lumenal/extracellular domains or shortening a cytoplasmic loop abolishes transport activity by both NPC1 and NPC1L1. The widely prescribed cholesterol uptake inhibitor, Ezetimibe, blocks NPC1L1; we show that interface residues that lie at the interface between NPC1L1's three extracellular domains comprise the drug's binding site. These data support a model in which cholesterol passes through the cores of NPC1/NPC1L1 proteins; concerted movement of various domains is needed for transfer and Ezetimibe blocks transport by binding to multiple domains simultaneously.

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“…For example, in the ternary complex of a single native Shh ligand bound to two human PTCH1 molecules 32, PTCH1 from chain a, the molecule whose sterol conduit is occluded by interaction with the N-terminal palmitoyl moiety of the SHH ligand, adopts pose 2, whereas PTCH1 from chain b adopts pose 1 32. Indeed, in all published structures of PTCH1 the switch helix adopts one or the other of these two poses15, [31][32][33][34][35]42, suggesting that they represent discrete alternative conformations preferentially populated within the PTCH1 activity cycle ( Fig. 4b).…”

Section: Structure Of the Ptch1::ti23 Complexmentioning

confidence: 99%

Hedgehog pathway activation through conformational blockade of the Patched sterol conduit

Zhang

Bulkley³

et al. 2019

Preprint

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Activation of the Hedgehog pathway may have therapeutic value for improved bone healing, taste receptor cell regeneration, and alleviation of colitis or other conditions. Systemic pathway activation, however, may be detrimental and therapeutic application has been difficult for lack of agents amenable to tissue targeting. We have developed a novel agonist, a conformation-specific nanobody against the Hedgehog receptor Patched1. This nanobody potently activates the Hedgehog pathway in vitro and in vivo by stabilizing an alternative conformation of a Patched1 "switch helix", as revealed by cryo-EM structure determination. Although this conformation likely constitutes part of the transport cycle, nanobody-trapping disrupts the cycle and prevents substrate movement through the Patched1 sterol conduit. Our conformation-selective nanobody approach provides a new route to the development of transporter-related pharmacologic agents and may be generally applicable to the study of other transporters.Therapeutic manipulation of the Hedgehog pathway has found its clearest application in patients with malignancies whose initiation and growth depend on pathway-activating mutations in the primary cells of the tumor, such as medulloblastoma and basal cell carcinoma 1,2. Pathway blockade, imposed by systemically administered small molecules, has provided clear benefits in such patients, albeit accompanied by unwanted but largely tolerable class effects such as muscle cramps, loss of hair, and loss of taste sensation 3. In contrast to promoting tumor growth, however, pathway activity recently has been found to suppress cancer growth and progression when it occurs in stromal cells rather than primary cells 4, particularly in cancers of endodermal

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Structural basis for the recognition of Sonic Hedgehog by human Patched1

Cited by 183 publications

References 81 publications

Patched regulates lipid homeostasis by controlling cellular cholesterol levels

Patched regulates lipid homeostasis by controlling cellular cholesterol levels

Inter-domain dynamics drive cholesterol transport by NPC1 and NPC1L1 proteins

Hedgehog pathway activation through conformational blockade of the Patched sterol conduit

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