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Methods in Enzymology, Volume 435 : Volume 435 - Helmut Sies

Methods in Enzymology, Volume 435

Volume 435

Hardcover Published: 8th November 2007
ISBN: 9780123739704
Number Of Pages: 592

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For over fifty years the Methods in Enzymology series has been the critically acclaimed laboratory standard and one of the most respected publications in the field of biochemistry. The highly relevant material makes it an essential publication for researchers in all fields of life and related sciences. This volume features articles on the topic of oxygen biology and hypoxia.

Contributorsp. xiii
Prefacep. xxi
Volumes in Seriesp. xxiii
Hypoxia-Inducible Factorp. 1
Hypoxia-Inducible Factors P[superscript ER]/ARNT/S[superscript IM] Domains: Structure and Functionp. 3
Introductionp. 4
Delineation of the HIF PAS Domainsp. 5
Expression and Characterization of HIF PAS Domainsp. 7
Assessing PAS Domain Protein-Protein Interactionsp. 11
Discussionp. 18
Acknowledgmentsp. 20
Referencesp. 21
Hypoxia-Inducible Factor Prolyl-Hydroxylase: Purification and Assays of PHD2p. 25
Introductionp. 26
Preparationp. 28
Assaying of PHD2 Activityp. 29
Indirect Measurements of PHD2 Activityp. 29
Direct Measurements of PHD2 Hydroxylation Activityp. 35
Binding Assaysp. 37
Comparison of Assay Formatsp. 38
Referencesp. 39
Determination and Modulation of Prolyl-4-Hydroxylase Domain Oxygen Sensor Activityp. 43
Introductionp. 44
Production of Functionally Active PHDsp. 48
Determination of PHD by VHL Binding to Peptides Derived from the HIF-1[alpha] ODD Domainp. 48
Determination of Prolyl-4-Hydroxylation by Oxidative Decarboxylation of 2-Oxoglutaratep. 51
Crude Tissue Extracts are not a Suitable Source of PHD Activity for the 2-Oxoglutarate Conversion Assayp. 53
Thin Layer Chromatography to Assess the Purity of [5-[superscript 14] C]2-Oxoglutaratep. 53
Application of the 2-Oxoglutarate Conversion Assay to Protein Targetsp. 55
Conclusionsp. 55
Acknowledgmentsp. 57
Referencesp. 57
Characterization of Ankyrin Repeat-Containing Proteins as Substrates of the Asparaginyl Hydroxylase Factor Inhibiting Hypoxia-Inducible Transcription Factorp. 61
Introductionp. 62
Experimental Techniquesp. 65
Discussion/Conclusionp. 82
Acknowledgmentsp. 83
Referencesp. 83
Transgenic Models to Understand Hypoxia-Inducible Factor Functionp. 87
Introductionp. 88
Hypoxia Response Pathway Genes and Developmentp. 90
HIF in Physiologyp. 93
HIF Function in Tumor Biologyp. 98
Summaryp. 100
Referencesp. 101
The Silencing Approach of the Hypoxia-Signaling Pathwayp. 107
A Brief History of RNAip. 108
The Hypoxia-Signaling Pathwayp. 109
HIF-[alpha] Stabilityp. 110
HIF Activityp. 114
HIF-1/HIF-2 Target Gene Specificityp. 116
RNAi as a New Potential Therapeutic Strategyp. 116
Acknowledgmentsp. 118
Referencesp. 118
Cellular and Developmental Adaptations to Hypoxia: A Drosophila Perspectivep. 123
Introductionp. 124
Drosophila melanogaster as a Model System to Study Physiological Responses to Hypoxiap. 124
Experimental Advantages of the Model Systemp. 125
The Drosophila Respiratory Systemp. 126
Occurrence of a Drosophila System Homologous to Mammalian HIFp. 128
Regulation of Sima by Oxygen Levelsp. 131
Role of Sima and Fatiga in Drosophila Developmentp. 132
Hypoxia-Inducible Genes and the Adaptation of Drosophila to Oxygen Starvationp. 134
Regulation of Sima by the PI3K and TOR Pathwaysp. 134
Role of the HIF System in Growth Control and Cell Size Determinationp. 136
Concluding Remarksp. 138
Acknowledgmentsp. 139
Referencesp. 139
Erythropoietinp. 145
Constitutively Overexpressed Erythropoietin Reduces Infarct Size in a Mouse Model of Permanent Coronary Artery Ligationp. 147
Introductionp. 148
Material and Methodsp. 149
Resultsp. 151
Discussionp. 153
Acknowledgmentsp. 154
Referencesp. 154
Use of Gene-Manipulated Mice in the Study of Erythropoietin Gene Expressionp. 157
Introductionp. 158
Materialsp. 161
Methods and Resultsp. 163
Conclusionp. 173
Acknowledgmentsp. 173
Referencesp. 174
Control of Erythropoietin Gene Expression and its Use in Medicinep. 179
Introductionp. 180
Native EPO Gene Expression and its Pharmacologic Stimulationp. 180
EPO Gene Transferp. 184
Recombinant EPOSp. 187
Conclusionsp. 190
Referencesp. 191
Role of Hypoxia-Inducible Factor-2[alpha] in Endothelial Development and Hematopoiesisp. 199
Introductionp. 200
Vasculogenesis/Angiogenesis and HIFsp. 200
HIF-1[beta]/ARNT Null Micep. 204
Neovascularization and HIFsp. 205
Hematopoiesis and HIFsp. 211
Conclusionp. 214
Referencesp. 214
Hypoxia and Adaptationp. 219
Organ Protection by Hypoxia and Hypoxia-Inducible Factorsp. 221
Introductionp. 222
From Ischemic to Hypoxic Preconditioningp. 222
Hypoxia-Inducible Transcription Factorsp. 223
Strategies to Activate HIF and HIF Target Genesp. 227
Hypoxic Preconditioning and HIFp. 231
HIF in Chronic Hypoxic/Ischemic Diseasesp. 236
Conclusions and Perspectivesp. 237
Referencesp. 238
Hypoxia and Regulation of Messenger RNA Translationp. 247
Introductionp. 248
Changes in Global mRNA Translation During Hypoxiap. 249
Molecular Mechanisms that Regulate mRNA Translation During Hypoxiap. 251
Methods Employed to Study mRNA Translation During Hypoxiap. 256
Protocolsp. 268
Referencesp. 271
Hypoxia and the Unfolded Protein Responsep. 275
Introductionp. 276
Methods Employed in Detecting Hypoxic Induction of ER Stressp. 282
Acknowledgmentsp. 289
Referencesp. 290
Hypoxia and Tumor Biologyp. 295
Tumor Hypoxia in Cancer Therapyp. 297
Hypoxia in Human Tumorsp. 298
The Dynamic Nature of Hypoxia in Tumorsp. 300
Consequences of Tumor Hypoxia for Cancer Treatmentp. 300
Size of the Oxygen Effect with Radiationp. 302
The Influence of Tumor Hypoxia on Cancer Treatment by Radiotherapyp. 303
Influence of Tumor Hypoxia on Response to Chemotherapyp. 307
Exploiting Hypoxia in Cancer Treatmentp. 308
Referencesp. 315
HIF Gene Expression in Cancer Therapyp. 323
Introductionp. 324
Experimental Proceduresp. 326
Conclusionsp. 337
Acknowledgmentsp. 337
Referencesp. 338
Analysis of Hypoxia-Inducible Factor-1[alpha] Expression and its Effects on Invasion and Metastasisp. 347
Introductionp. 347
Protocol 1: HIF-1[alpha] Immunohistochemistryp. 349
Protocol 2: Invasion Assayp. 350
Protocol 3: Transepithelial Resistance Measurement of Cell-Cell Adhesionp. 351
Protocol 4: Analysis of MRNA Expression by QRT-PCRp. 351
Referencesp. 352
Macrophage Migration Inhibitory Factor Manipulation and Evaluation in Tumoral Hypoxic Adaptationp. 355
Introductionp. 356
Modulation of MIF Levels by Targeted shRNAs and Assessment of Knockdown Efficiencyp. 357
Analysis of MIF-Dependent CSN5 and COP9 Signalosome Functionp. 362
Determination of Tumor-Associated MIF Expression and MIF Polymorphic Disparityp. 364
Conclusionsp. 367
Referencesp. 367
The Von Hippel-Lindau Tumor Suppressor Protein: An Updatep. 371
Introductionp. 372
Regulation of Epithelial Differentiation by pVHLp. 373
Crosstalk Between c-Met and VHLp. 374
Regulation of Neuronal Apoptosis by pVHLp. 375
Possible Links Between p53 and pVHLp. 376
Regulation of pVHL by Phosphorylationp. 376
Polyubiquitylation of pVHLp. 377
Mouse Models for Studying pVHL Functionp. 377
Referencesp. 378
Hypoxia-Inducible Factor 1 Inhibitorsp. 385
Introductionp. 386
Cell-Based High Throughput Screensp. 387
Cell-Free Assaysp. 393
Bioassay-Directed Isolation of Natural Product HIF-1 Inhibitorsp. 398
Conclusionsp. 399
Acknowledgmentsp. 399
Referencesp. 400
Hypoxia and Inflammatory Mediatorsp. 403
Regulation of Hypoxia-Inducible Factors During Inflammationp. 405
Introductionp. 406
Regulation of HIF at the Transcriptional Levelp. 408
Regulation of HIF at the Translational Levelp. 410
Regulation of HIF-1[alpha] at the Posttranslational Levelp. 411
Regulation of HIF-1 Activityp. 413
Perspectivesp. 414
Conclusionsp. 414
Acknowledgmentsp. 415
Referencesp. 415
Superoxide and Derived Reactive Oxygen Species in the Regulation of Hypoxia-Inducible Factorsp. 421
Introductionp. 422
Reactive Oxygen Species Act as Signaling Moleculesp. 423
HIFs are Sensitive to Oxygenp. 424
Reactive Oxygen Species Modulate HIFp. 425
How are HIFs Regulated by Reactive Oxygen Species?p. 427
Summaryp. 431
Methodsp. 431
The Cytochrome C Reduction Assay for Detection of Extracellular Reactive Oxygen Speciesp. 431
Chemiluminescence Assay for Detection of Extracellular Reactive Oxygen Speciesp. 432
Measuring Intracellular Production of Reactive Oxygen Species using Fluorescent Dyesp. 433
Detection of Reactive Oxygen Species by Electron Paramagnetic Resonancep. 436
Acknowledgmentsp. 438
Referencesp. 438
Genetics of Mitochondrial Electron Transport Chain in Regulating Oxygen Sensingp. 447
Introductionp. 448
Detecting HIF-1[alpha] Protein Levelsp. 449
Detecting Intracellular ROS Levelsp. 452
Method 1: Examining Hypoxic Stabilization of HIF-1[alpha] Protein in Cells Containing RNAI against the Rieske Fe-S Proteinp. 454
Method 2: Examining the Role of ROS Generated from Mitochondrial Electron Transport in Hypoxic Stabilization of HIF-1[alpha] Proteinp. 458
Concluding Remarksp. 459
Acknowledgmentsp. 459
Referencesp. 460
Hypoxia-Inducible Factor-1[alpha] Under the Control of Nitric Oxidep. 463
HIF-1 and Oxygen Sensingp. 464
Nitric Oxide: A Multifunctional Messengerp. 465
Accumulation of HIF-1[alpha] and Activation of HIF-1 by NOp. 467
Superoxide Stabilizes HIF-1[alpha] but Antagonizes NO Actionsp. 470
Hypoxic Signal Transmission is Antagonized by NOp. 472
Summary and Conclusionsp. 473
Acknowledgmentsp. 475
Referencesp. 475
Hypoxic Regulation of NF-[kappa]B Signalingp. 479
Backgroundp. 480
Treatment Protocols for Cellular Hypoxia Studiesp. 481
Measurement of NF-[kappa]B Activity in Cultured Cellsp. 484
Summary/Conclusionsp. 491
Referencesp. 491
Author Indexp. 493
Subject Indexp. 535
Table of Contents provided by Ingram. All Rights Reserved.

ISBN: 9780123739704
ISBN-10: 0123739705
Series: Methods in Enzymology
Audience: Professional
Format: Hardcover
Language: English
Number Of Pages: 592
Published: 8th November 2007
Publisher: Elsevier Science Publishing Co Inc
Country of Publication: US
Dimensions (cm): 22.9 x 15.2  x 2.79
Weight (kg): 0.92
Edition Number: 435

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