TGF-ß Signaling Methods and Protocols

This volume details up-to-date information and attempts to take the reader into the exciting realm of TGF-ß from the basic principles to the practical applications. Chapters provide basic introduction of TGF-ß signaling from the cell surface to the nucleus, methods and techniques for the investigation of TGF-ß signaling mechanism including receptors, Smads, intracellular kinases, microRNA, epigenetic regulation, post-translational regulations, non-Smad pathway; the physiological implications including those in cell cycle arrest, epithelial-mesenchymal transition, endothelial cells, adipogenesis, Th differentiation, stem cell, bone remodeling, ovary, zebrofish development, and frog animal capping; and the methodologies including metastasis imaging, 3D morphogenesis, membrane receptor quantification, conditional knockout, bone remodeling, kinase and phosphatase assays, BiFC interaction assays, and genome-wide siRNA screen. Written in the highly successful Methods in Molecular Biology series format, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and key tips on troubleshooting and avoiding known pitfalls.

Authoritative and practical, TGF-Beta Signaling: Methods and Protocols aims to ensure successful results in the further study of this vital field.

1.                  Regulation of TGF-b Receptors

Erine Budi, Jian Xu and Rik Derynck

2.                  Determining TGF-ß Receptor Levels in the Cell Membrane

Long Zhang, Fangfang Zhou, Maarten van Dinther, and Peter ten Dijke

3.                  Post-translational Modifications of TGF-b Receptors

Xiaohua Yan and Ye-Guang Chen     

4.                  Production, Isolation,and Structural Analysis of Ligands and Receptors of the TGF-b Superfamily

5.                  Phosphorylation of Smads by Intracellular Kinases

Fang Liu and Isao Matsuura

6.         Analysis of Smad Phosphatase Activity in vitro

Tao Shen, Lan Qin and Xia Lin

7.                  Three-dimensional Mammary Epithelial Cell Morphogenesis Model for Analysis of TGF-b Signaling

Juliet Rashidian and Kunxin Luo

8.                  TGF-ß Signaling in Stem Cell Regulation

Wenlin Li, Wanguo Wei, Sheng Ding

9.                  Analysis of Epithelial-Mesenchymal Transition Induced by Transforming Growth Factor ß

Ulrich Valcourt, Jonathon Carthy, Yukari Okita, Lindsay Alcaraz, Mitsuyasu Kato, Sylvie Thuault, Laurent Bartholin and Aristidis Moustakas

10.              In vitro Th differentiation protocol

Takashi Sekiya and Akihiko Yoshimura

11.              Interrogating TGF-b Function and Regulation in Endothelial Cells

J.A. Maring, L.A. van Meeteren, M.J. Goumans and P. ten Dijke

12.       Isolation and Manipulation of Adipogenic Cells to Assess TGF-b Superfamily Functions

Maria Namwanje, Juan .C. Bournat and Chester .W. Brown

13.              Imaging TGFb Signaling in Mouse Models of Cancer Metastasis

Yibin Kang

14.              Generation and Characterization of Smad7 Conditional Knockout Mice

Yi Pan and Yan Chen

15.              Monitoring Smad Activity in vivo Using the Xenopus Model System

Marco Montagner, Graziano Martello and Stefano Piccolo

16.              Animal Cap Assay for TGF-b Signaling

Chenbei Chang

17.              Detection of Smad Signaling in Zebrafish Embryos

Xingfeng Liu, Qiang Wang and Anming Meng

18.              Role of TGF-ß signaling in Coupling Bone Remodeling

Janet L. Crane, Lingling Xian and Xu Cao

19.              Studying the Functions of TGF-ß Signaling in the Ovary

Chao Yu, Jian-Jie Zhou and Heng-Yu Fan

20.              Quantitative Real-Time PCR Analysis of MicroRNAs and Their Precursors Regulated by TGF-b Signaling

Hara Kang and Akiko Hata

21.              TGF-ß Regulated microRNAs and Their Function in Cancer Biology

Pengyuan Yang, Yun Zhang, Geoffrey J. Markowitz, Xing Guo and Xiao-Fan Wang

22.              Epigenomic Regulation Of Smad1 Signaling During Cellular Senescence Induced by Ras Activation

Atsushi Kaneda, AyaNonaka, Takanori Fujita, Ryota Yamanaka, Mai Fujimoto, Kohei Miyazono and Hiroyuki Aburatani

23.              The Role of Ubiquitination to Determine Non-Smad Signaling Responses

Shyam Kumar Gudey and Marene Landström

24.              Genome-wide RNAi Screening to Dissect the TGF-b Signal Transduction Pathway

Xiaochu Chen and LanXu

Zipei Feng and Xuedong Liu