Phosphodiesterase Methods and Protocols

The multigenic family of cyclic nucleotide phosphodiesterases (PDEs) is now known to offer many new therapeutic possibilities through their influence over intracellular signaling, though their precise cell mechanisms to modulating that process remain uncertain. In Phosphodiesterase Methods and Protocols, a panel of research leaders in the PDE field describes new concepts and techniques for investigating the role of PDEs in orchestrating normal and pathophysiological responses. Presented in step-by-step detail, these readily reproducible methods allow the measurement of cyclic nucleotide variations in living cells, as well as their visualization in a spatiotemporal manner, the localization and characterization of their activities in tissues and living cells, and the assessment of targeted PDEs in creating specific tools and drugs. Additional chapters discuss the generation of PDE4 knockout mice to demonstrate not only the potential role of targeted PDEs, but also their use in further studies of the central role of PDE regulation in intracellular signaling control. These techniques offer clinicians a way to find new therapies for numerous pathologies whose molecular origins are unknown and whose treatment is symptomatic. Alterations of intracellular signaling related to PDE deregulation in such pathologies as inflammation, neurodegeneration, and cancer may explain the difficulties observed in their prevention and treatment. The protocols follow the successful Methods in Molecular Biology™ series format, each offering step-by-step laboratory instructions, an introduction outlining the principle behind the technique, lists of the necessary equipment and reagents, and tips on troubleshooting and avoiding known pitfalls.

Path-breaking and highly practical, Phosphodiesterase Methods and Protocols offers cell biologists, molecular biologists, pharmacologists, and medicinal chemists a broad ranging survey of the advanced tools and concepts needed to understand therole of PDEs in physiological functions, their implications in several critical pathologies, and the opportunities they offer for new drug design.

Inhaltsverzeichnis

Study of Cyclic Adenosine Monophosphate Microdomains in Cells
Marco Mongillo, Anna Terrin, Sandrine Evellin, Valentina Lissandron, and Manuela Zaccolo
High-Resolution Measurements of Cyclic Adenosine Monophosphate Signals in 3D Microdomains
Jeffrey W. Karpen and Thomas C. Rich
Cygnets: In Vivo Characterization of Novel cGMP Indicators and In Vivo Imaging of Intracellular cGMP
Akira Honda, Carolyn L. Sawyer, Sharon M. Cawley, and Wolfgang R. G. Dostmann
High-Throughput Screening of Phosphodiesterase Activity in Living Cells
Thomas C. Rich and Jeffrey W. Karpen
Assessment of Phosphodiesterase Isozyme Contribution in Cell and Tissue Extracts
Thérèse Keravis, Rima Thaseldar-Roumié, and Claire Lugnier
Localization of the Cyclic Guanosine 39,59-Monophosphate- Hydrolyzing Phosphodiesterase Type 9 in Rat Brain by Nonradioactive In Situ Hybridization
Wilma C. G. van Staveren and Marjanne Markerink-van Ittersum
Determination of Ca2+/Calmodulin-Stimulated Phosphodiesterase Activity in Intact Cells
Chen Yan
Adenovirus-Mediated Overexpression of Murine Cyclic Nucleotide Phosphodiesterase 3B
Faiyaz Ahmad, Linda Härndahl, Yan Tang, Lena Stenson Holst, and Vincent C. Manganiello
Identification of Promoter Elements in the 5'-Flanking Region of Murine Cyclic Nucleotide Phosphodiesterase 3B Gene
Hanguan Liu, Jing Rong Tang, Eva Degerman, and Vincent C. Manganiello
Purification of PDE6 Isozymes From Mammalian Retina
Dana C. Pentia, Suzanne Hosier, Rachel A. Collupy, Beverly A. Valeriani, and Rick H. Cote
Cyclic Guanosine 59-Monophosphate Binding to Regulatory GAF Domains of Photoreceptor Phosphodiesterase
Rick H. Cote
Renaturation of the Catalytic Domain of PDE4A Expressed in Escherichia coli as Inclusion Bodies
Wito Richter, Thomas Hermsdorf, and Dietrich Dettmer
Determining the Subunit Structure of Phosphodiesterases Using Gel Filtration and Sucrose Density Gradient Centrifugation
Wito Richter
Crystallization of Cyclic Nucleotide Phosphodiesterases
Hengming Ke, Qing Huai, and Robert X. Xu
Generation of PDE4 Knockout Mice by Gene Targeting
S.-L. Catherine Jin, Anne M. Latour, and Marco Conti
Immunoprecipitation of PDE2 Phosphorylated and Inactivated by an Associated Protein Kinase
J. Kelley Bentley
Investigation of Extracellular Signal-Regulated Kinase 2 Mitogen-Activated Protein Kinase Phosphorylation and Regulation of Activity of PDE4 Cyclic Adenosine Monophosphate-Specific Phosphodiesterases
Elaine V. Hill, Miles D. Houslay, and George S. Baillie
Radiolabeled Ligand Binding to the Catalytic or Allosteric Sites of PDE5 and PDE11
James L. Weeks II, Mitsi A. Blount, Alfreda Beasley, Roya Zoraghi, Melissa K. Thomas, Konjeti Raja Sekhar, Jackie D. Corbin, and Sharron H. Francis
Analysis of Dimerization Determinants of the PDE6 Catalytic Subunits
Khakim G. Muradov, Kimberly K. Boyd, and Nikolai O. Artemyev
Interaction Between Catalytic and Inhibitory Subunits of PDE6
Nikolai O. Artemyev
Purification, Reconstitution on Lipid Vesicles, and Assays of PDE6 and Its Activator G Protein, Transducin
Theodore G. Wensel, Feng He, and Justine A. Malinski
Index

Zusammenfassung

Research leaders in the PDE field describe new concepts and techniques for investigating the role of PDEs in orchestrating normal and pathophysiological responses. Presented in step-by-step detail, these readily reproducible methods allow the measurement of cyclic nucleotide variations in living cells, as well as their visualization in a spatio-temporal manner, the localization and characterization of their activities in tissues and living cells, and the assessment of targeted PDEs in creating specific tools and drugs.

Zusatztext

"...interesting and thought provoking, but technically challenging...excellent guide to current research techniques in the field of phosphodiesterase." - Doody's Health Sciences Book Review Journal

Produktdetails

• Mitarbeit: Claire Lugnier (Herausgeber)
• Verlag: Springer, Berlin
• Sprache: Englisch
• Produktform: Fester Einband
• Erschienen: 17.04.2009
• EAN: 9781588293145
• ISBN: 978-1-58829-314-5
• Seiten: 336
• Gewicht: 1 g
• Illustration: 92 SW-Abb., 1 Farbabb., 22 Duoton-Abb., 70 SW-Zeichn.
• Serien: Methods in Molecular Biology; Methods in Molecular Biology
• Themen: Naturwissenschaften, Medizin, Informatik, Technik > Biologie > Biochemie, Biophysik; B, hybridization, biochemistry, Biomedical and Life Sciences, Biochemistry, general, genes, NUCLEOTIDE, termination, Promoter