P53 Protocols

ISBN-13: 9781588291066 / Angielski / Twarda / 2003 / 280 str.

Sumitra Deb; Swati Palit Deb; Sumitra Deb

Opis

Since the discovery of p53 as a tumor suppressor, numerous methods have evolved to reveal the unique structural features and biochemical functions of this protein. Several unique properties of p53 posed a challenge to understa- ing its normal function in the initial phase of its research. The low levels of p53 in normal cells, its stabilization under situations of genotoxic stress, induction of growth arrest, and apoptosis with stabilization of the protein, obstructed the visibility of its normal, unmutated function. The property of p53 that can sense a promoter and transactivate or inhibit is still not well understood. It is still not known whether it is the absence of the protein that causes tumorigenesis, or if its mutants have a dominant role in inducing cancer. p53 Protocols comprises eighteen chapters for the study of the diverse properties of p53 and related proteins. The methods included are invaluable for delineating the function of other proteins that may function as tumor suppr- sors or growth suppressors. The chapters are not presented in any schematic order, for the importance and diversity of the functions of p53 make it imp- sible to organize them suitably. We have made a sincere effort to collect the methods most useful to those investigators working on tumor suppressors or growth suppressors. The purpose of p53 Protocols is not only to provide investigators with methods to analyze similar biochemical functions, but also to familiarize them with the associated problems that arose during the course of investigations.

Spis treści

Adenovirus Expressing p53 Matthew Holmes, Elizabeth Rosenberg, and Kristoffer Valerie Purification of Recombinant p53 from Sf9 Insect Cells Xiu Zhu Sun, John Nguyen, and Jamil Momand Generation and Characterization of p53 Mutant Mice Melissa L. Dumble, Lawrence A. Donehower, and Xiongbin Lu Identification of p53 Target Genes by Fluorescent Differential Display Susanne Stein, Yong-Jig Cho, Roger S. Jackson II, and Peng Liang p53-Induced Gene Expression Analysis Joanna K. Sax and Wafik S. El-Deiry Real-Time Polymerase Chain Reaction Quantitation of Relative Expression of Genes Modulated by p53 Using SYBR® Green I Katherine E. Roth Stagliano, Evie Carchman, and Sumitra Deb Transactivation and Transrepression Studies with p53 Mariano J. Scian, Rebecca Frum, Swati Palit Deb, and Sumitra Deb Methods to Study p53-Repressed Promoters Patrick Dumont, Anthony Della Pietra, and Maureen E. Murphy Interaction of p53 with Cellular Proteins David M. Keller, Shelya X. Zeng, and Hua Lu Interaction of p53 with the Adenovirus E1B-55 kDa Protein Judith Roth and Matthias Dobbelstein In Vivo Footprinting and DNA Affinity Chromatography for Analysis of p53 DNA Binding Ability Maria Patricia Molina, Christine Cain, and Jill Bargonetti Signaling to p53: The Use of Phospho-Specific Antibodies to Probe for In Vivo Kinase Activation Ashley L. Craig, Susan E. Bray, Lee E. Finlan, Neil M. Kernohan, and Ted R. Hupp Detection of Mitochondrial Localization of p53 Motohiro Mihara and Ute M. Moll Analysis of Nuclear and Cytoplasmic Degradation of p53 in Cells after Stress Troy W. Joseph and Ute M. Moll Mutational Analysis of p53 in Human Tumors: Direct DNA Sequencing and SSCP Susan Erster, Neda Slade, and Ute M. Moll Mutational Analysis of p53 in Human Tumors:Immunocytochemistry Neda Slade and Ute M. Moll Flow Cytometric Analysis of p53-Induced Apoptosis Michael Berger and Ygal Haupt Flow Cytometric Analysis of MDM2-Mediated Growth Arrest Rebecca Frum and Swati Palit Deb Index

Blurb

Since the discovery of p53 as a tumor suppressor, numerous methods have evolved to reveal its unique structural features and biochemical functions. In p53 Protocols, Sumitra and Swati Palit Deb have assembled an indispensable collection of novel techniques that have proven most useful for studying the physiological properties of p53 both in vitro and in vivo. Described by leading basic and clinical researchers who have successfully used the methods, the techniques provide proven solutions to problems in studying the purification, target identification, gene expression, quantitation, interaction, signaling, transactivation, and transrepression of p53. The methods are also useful for delineating the functions of other proteins that may act as tumor or growth suppressors. Each technique includes step-by-step instructions, troubleshooting notes, a theoretical review, and discussion of associated problems that might arise during the course of investigation. Special effort has been made to discuss problems and their solutions to help ease the learning curve when standardizing a new method.
Diverse and highly practical, p53 Protocols offers both beginning and experienced researchers in cancer biology a gold-standard compendium of readily reproducible techniques for studying p53 and related proteins.