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Hedgehog Signaling

Methods and Protocols

  • Book
  • © 2022

Overview

Editors:
  1. Xiaochun Li

    1. Department of Molecular Genetics and Biophysics, University of Texas Southwestern Medical Center, Dallas, USA

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  • Includes cutting-edge techniques
  • Provides step-by-step detail essential for reproducible results
  • Contains key implementation advice from the experts

Part of the book series: Methods in Molecular Biology (MIMB, volume 2374)

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Table of contents (20 protocols)

  1. Front Matter

    Pages i-xi
    Download chapter PDF

  2. Generation and Analysis of Mosaic Spinal Cord Organoids Derived from Mouse Embryonic Stem Cells

    • Carina Jägers, Henk Roelink
    Pages 1-11

  3. Evaluating Hedgehog Acyltransferase Activity and Inhibition Using the Acylation-coupled Lipophilic Induction of Polarization (Acyl-cLIP) Assay

    • Sebastian A. Andrei, Edward W. Tate, Thomas Lanyon-Hogg
    Pages 13-26

  4. Analysis of Protein Cholesterylation by Biorthogonal Labeling

    • Ao Hu, Meng Zhou, Bao-Liang Song
    Pages 27-36

  5. An In Situ Fluorescence Assay for Cholesterol Transporter Activity of the Patched

    • Wonhwa Cho, Arthur Ralko, Ashutosh Sharma
    Pages 37-47

  6. Imaging of the Ciliary Cholesterol Underlying the Sonic Hedgehog Signal Transduction

    • Tatsuo Miyamoto, Kosuke Hosoba, Silvia Natsuko Akutsu, Shinya Matsuura
    Pages 49-57

  7. Studying Hedgehog Signaling During Mouse Neural Tube Development

    • Sandii Constable, Kevin White, Bandarigoda Nipunika Somatilaka, Saikat Mukhopadhyay
    Pages 59-71

  8. Measuring and Manipulating Membrane Cholesterol for the Study of Hedgehog Signaling

    • Maia Kinnebrew, Kristen A. Johnson, Arun Radhakrishnan, Rajat Rohatgi
    Pages 73-87

  9. The Immunofluorescence-Based Detection of Hedgehog Pathway Components in Primary Cilia of Cultured Cells

    • Shu Yang, Xiaoyan Zheng
    Pages 89-94

  10. Analysis of Dispatched Protein Processing and Sonic Hedgehog Ligand Release

    • Elizabeth Cleverdon, Daniel P. Stewart, Stacey K. Ogden
    Pages 95-106

  11. Expression, Purification, and Structure Determination of Human PTCH1–HH-N Complexes

    • Xiaofeng Qi, Philip Schmiege, Leticia Esparza, Xiaochun Li
    Pages 107-120

  12. Characterization of Smoothened Phosphorylation and Activation

    • Jianhang Jia, Yuhong Han, Yong Suk Cho, Jin Jiang
    Pages 121-137

  13. Cell-Based Assays for Smoothened Ubiquitination and Sumoylation

    • Yuhong Han, Jin Jiang
    Pages 139-147

  14. Biochemical Assays to Directly Assess Smoothened Activation by a Conformationally Sensitive Nanobody

    • Ishan Deshpande, Aashish Manglik
    Pages 149-160

  15. Studies of SMOOTHENED Activation in Cell-Free and Reconstituted Systems

    • Isaac B. Nelson, Benjamin R. Myers
    Pages 161-174

  16. Rapid, Direct SMOOTHENED Activity Assays in Live Cells Using cAMP-Based Conformational Sensors

    • Madison F. Walker, Benjamin R. Myers
    Pages 175-184

  17. Light-Induced Differentiation of Forebrain Organoids by NVOC-SAG

    • Ryuji Misawa, Yoshiho Ikeuchi
    Pages 185-194

  18. NanoBRET and NanoBiT/BRET-Based Ligand Binding Assays Permit Quantitative Assessment of Small Molecule Ligand Binding to Smoothened

    • PaweÅ‚ Kozielewicz, Gunnar Schulte
    Pages 195-204

  19. Measuring Smoothened (SMO)-Mediated Activation of the Gi Protein

    • Heng Liu, Cheng Zhang
    Pages 205-212

  20. Ci/Gli Phosphorylation by the Fused/Ulk Family Kinases

    • Yuhong Han, Jin Jiang
    Pages 213-229
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Keywords

About this book

This detailed book explores the technical breakthroughs with biophysical and cell biological approaches that have advanced the study of molecular mechanisms underlying the generation and transduction of the hedgehog (HH) signal, indicating the important role of sterols in this pathway. Within its pages, the volume examines techniques involving several key protein components in this pathway, including HH, the signaling ligand; Dispatched, a HH secretion regulator; Patched, the receptor of HH; Smoothened, the signal transducer used to transduce the HH signal across the plasma membrane; and GLI, the transcription factor to turn on HH target genes. Written in the highly successful Methods in Molecular Biology series format, chapters feature introductions to their respective topics, lists of the necessary materials and reagents, step-by-step readily reproducible laboratory protocols, as well as tips on troubleshooting and avoiding known pitfalls. 

Authoritative and practical, Hedgehog Signaling: Methods and Protocols provides researchers in clinical and basic science with a fresh and reliable reference for analyzing HH signal transduction through cell biological, biochemical, and biophysical approaches.

Editors and Affiliations

  • Department of Molecular Genetics and Biophysics, University of Texas Southwestern Medical Center, Dallas, USA

    Xiaochun Li

Bibliographic Information

  • Book Title: Hedgehog Signaling

  • Book Subtitle: Methods and Protocols

  • Editors: Xiaochun Li

  • Series Title: Methods in Molecular Biology

  • DOI: https://doi.org/10.1007/978-1-0716-1701-4

  • Publisher: Humana New York, NY

  • eBook Packages: Springer Protocols

  • Copyright Information: The Editor(s) (if applicable) and The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature 2022

  • Hardcover ISBN: 978-1-0716-1700-7Published: 27 September 2021

  • Softcover ISBN: 978-1-0716-1703-8Published: 28 September 2022

  • eBook ISBN: 978-1-0716-1701-4Published: 25 September 2021

  • Series ISSN: 1064-3745

  • Series E-ISSN: 1940-6029

  • Edition Number: 1

  • Number of Pages: XI, 244

  • Number of Illustrations: 10 b/w illustrations, 27 illustrations in colour

  • Topics: Protein Science, Cancer Research

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