Elsevier S & T

Osmosensing and Osmosignaling

Sies, Helmut

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Table of contents
  • Contentsv
  • Contributorsxv
  • Prefacexxiii
  • Volumes in Seriesxxv
  • Chapter 1: Water Compartments in Cells1
  • 1. Introduction2
  • 2. The Stoichiometric Hydration Model (SHM)3
  • 2.1. Water bridges3
  • 2.2. Methods used to calculate compartmental capacities3
  • 2.3. Solvent-accessible surface area calculation method5
  • 2.4. SASA compensation assuming compact folding of globular proteins8
  • 2.5. SASA estimate for globular proteins based on the hydration model9
  • 3. Predictions Using the SHM9
  • 3.1. Prediction of hydration capacities hRa, hB, hPr, and hM(native)9
  • 3.2. Comparison of hydration model predictions to measured capacities11
  • 4. Biophysical Measurements of Compartmental Hydration Capacities11
  • 4.1. Selection of methods11
  • 4.2. Gravimetric measurement of hydration12
  • 4.3. Protein rehydration rate method13
  • 4.4. Differential scanning calorimetry method15
  • 4.5. Proton NMR titration method15
  • 4.6. Osmotic compression method15
  • 5. Relationship of the Hydration Model to Osmosensing and Osmosignaling18
  • 6. Relationship of the SH Model to Enzyme Function19
  • 6.1. Enzyme hydration and activity19
  • 6.2. Neutron scattering assessment of enzyme motion and activity21
  • 6.3. The SH model predicts both vibrational dynamics and enzyme activity changes23
  • 7. Relationship of the SH Model to Cellular Function24
  • 7.1. Cellular function depends on water24
  • 7.2. Hydration compartments and cellular systems correlate with SH model predictions24
  • 8. Summary and Conclusions25
  • References25
  • Chapter 2: Yeast Osmoregulation29
  • 1. Time Line: Response to Osmotic Shock30
  • 2. Glycerol Metabolism and the Aquaglyceroporin Fps131
  • 3. The HOG Signaling System33
  • 4. Transcriptional Responses35
  • 5. Studying the Control of Signaling37
  • 6. Future Directions41
  • Acknowledgments41
  • References42
  • Chapter 3: Physiological Analysis of Bacterial Mechanosensitive Channels47
  • 1. Introduction47
  • 2. Analysis of Native Channel Function49
  • 3. Protocol54
  • 4. Variants on the Basic Viability Assay56
  • 5. Analyzing Mutant Channels57
  • References59
  • Chapter 4: Control of Cell Cycle in Response to Osmostress: Lessons from Yeast63
  • 1. Introduction64
  • 2. Regulation of the Yeast Cell Cycle64
  • 3. Osmostress Response in Budding and Fission Yeast66
  • 4. Control of the G1 Phase by SAPKs68
  • 5. Control of the G2 Phase by SAPKs70
  • 6. Control of the Anaphase-Telophase Transition71
  • 7. Regulation of Exit from Mitosis by SAPKs72
  • 8. Conclusions and Perspectives73
  • References74
  • Chapter 5: Bacterial Osmosensing TransportersCover
  • 1. Introduction78
  • 1.1. Bacterial osmotolerance, halotolerance, and osmoregulation78
  • 1.2. Osmosensing and osmosignaling82
  • 2. Estimation of Key Solution Properties and of Turgor Pressure85
  • 2.1. Osmotic pressure, water activity, osmolality, and osmolarity85
  • 2.2. Ionic strength86
  • 2.3. Macromolecular crowding, volume exclusion, and protein hydration87
  • 2.4. Turgor pressure87
  • 3. Assessment of Osmotolerance88
  • 4. Identification of Osmoprotectants, Compatible Solutes, and Organic Osmolytes89
  • 5. Identification and Distribution of Osmosensory and Osmoregulatory Transporters90
  • 5.1. Transporter identification90
  • 5.2. Transporter gene distribution95
  • 6. In Vitro Systems96
  • 6.1. Membrane vesicle preparation96
  • 6.2. Proteoliposome preparation97
  • 6.3. Membrane composition and topology in cells, membrane vesicles, and proteoliposomes97
  • 7. Measurement of Osmosensory Transporter Activity98
  • 8. Conclusion100
  • References101
  • Chapter 6: Plant Gene Networks in Osmotic Stress Response: From Genes to Regulatory Networks109
  • 1. Introduction110
  • 2. Identification of Stress-Responsive Genes113
  • 2.1. Plant preparation and treatments113
  • 2.2. RNA purification113
  • 2.3. Preparation of cDNA library and screening using differential hybridization114
  • 2.4. Arabidopsis full-length cDNA microarray analysis114
  • 2.5. Agilent Arabidopsis 2 oligo microarray analysis114
  • 2.6. RNA gel blot analyses115
  • 2.7. Real-time PCR115
  • 3. Identification of cis-Acting Regulatory Elements: Use of GUS, LUC, and GFP Reporter Genes115
  • 3.1. Methods for promoter analysis and screening of mutants using GUS reporter gene117
  • 3.2. Methods for promoter analysis and screening of mutants using LUC reporter gene117
  • 3.3. Methods for application of GFP reporter gene117
  • 3.4. In silico analysis117
  • 3.5. Histochemical assay118
  • 4. Identification of Trans-Acting Regulatory Elements118
  • 4.1. Strategy for isolation of cDNAs encoding TFs using yeast one-hybrid system119
  • 4.2. Verification of specific binding of TFs to cis elements120
  • 5. Characterization of TFs121
  • 5.1. Transient transactivation assay121
  • 5.2. Nature of the TF121
  • 5.3. Localization of DNA-binding domain121
  • 5.4. Localization of activation domain122
  • 5.5. Interaction of TFs122
  • 6. Functional Analysis of TFs In Planta122
  • 6.1. Post-translational modification of TF by phosphorylation122
  • 6.2. Gain-of-function study of TFs in plants123
  • 6.3. Loss-of-function study of TFs in plants123
  • Acknowledgments124
  • References125
  • Chapter 7: Osmosensing by Integrins in Rat Liver129
  • 1. Integrins in Mechanotransduction and Growth Factor Signaling130
  • 2. Cell Hydration and Cell Function132
  • 3. The Perfused Rat Liver: An Intact Organ Model of the Liver133
  • 3.1. Measurement of cell hydration133
  • 3.2. Measurement of proteolysis135
  • 3.3. Measurement of potassium fluxes135
  • 3.4. Measurement of canalicular bile acid excretion136
  • 3.5. Tissue processing for Western blot analysis136
  • 4. Activation of beta1 Integrin Subunit by Hypoosmolarity and Insulin in Perfused Rat Liver136
  • 5. Integrins in the Regulation of Autophagic Proteolysis in the Liver136
  • 6. Integrins in the Regulation of Canalicular Bile Acid Excretion in the Liver138
  • 7. Integrins in Hepatocyte Volume Regulation139
  • 8. Conclusion139
  • References140
  • Chapter 8: Hyperosmotic Activation of the CD95 System145
  • 1. Introduction146
  • 2. Hyperosmotic Activation of the CD95 System in Hepatocytes148
  • 3. Concluding Remarks156
  • Acknowledgments156
  • References156
  • Chapter 9: New Approaches for Determining Apoptotic Volume Decrease in Cells161
  • 1. Introduction162
  • 2. Classical Ways to Study Cell Shrinkage or AVD During Apoptosis163
  • 3. Instrument Setup and Calibration for the Cell Lab Quanta SC165
  • 3.1. Instrument start-up and calibration166
  • 3.2. Fluorescence calibration protocol166
  • 3.3. Volume calibration protocol167
  • 4. General Considerations in Studying AVD and Apoptosis on the Cell Lab Quanta SC168
  • 5. Preparation of Cells Undergoing Apoptosis169
  • 6. AVD and Externalization of Phosphatidylserine During Apoptosis169
  • 7. AVD and Caspase Activity During Apoptosis170
  • 7.1. Preparing CaspaTag reagents170
  • 7.2. Protocol for CaspaTag/cell size analysis on the Cell Lab Quanta SC171
  • 8. AVD and Plasma Membrane Potential During Apoptosis171
  • 8.1. Preparing DiBAC4 reagent171
  • 8.2. Protocol for DiBAC4/cell size analysis on the Cell Lab Quanta’SC172
  • 9. Data Acquisition for the Cell Lab Quanta SC172
  • 10. Data Analysis for AVD and Apoptotic Characteristics on the Cell Lab Quanta SC172
  • 11. Conclusion180
  • Acknowledgments63
  • References63
  • Chapter 10: Transient Receptor Potential Channels in Mechanosensing and Cell Volume Regulation183
  • 1. Introduction184
  • 2. General Mechanisms of Mechano- or Osmosensing by Membrane Proteins184
  • 3. TRP Channels in Mechano- and Osmosensing188
  • 4. TRP Channels in Cell Volume Regulation192
  • 5. Experimental Procedures193
  • 5.1. Electrophysiological recordings used to assess mechanosensitivity of TRP channels193
  • 5.2. Electrophysiological recordings to assess osmosensitivity of TRP channels195
  • 5.3. Assessing the contribution of TRP channels to cell volume regulation: simultaneous measurements197
  • 5.4. Combined patch clamp and cell volume measurements200
  • Acknowledgments201
  • References201
  • Chapter 11: Cell Volume Regulatory Ion Channels in Cell Proliferation and Cell Death209
  • 1. Introduction210
  • 2. Anion Channels, Osmolyte Transport, and pH Regulation211
  • 3. Ca2+ and Unselective Cation Channels212
  • 4. K+ channels213
  • 5. Switching from Cell Proliferation to Suicidal Cell Death215
  • Acknowledgments63
  • References63
  • Chapter 12: Actin Cytoskeleton Architecture and Signaling in Osmosensing227
  • 1. Introduction228
  • 2. Morphological Analysis of Actin Cytoskeleton During Cell Volume Changes229
  • 3. Quantitative Biochemical Analysis of Actin Cytoskeleton Dynamics During Cell Volume Changes230
  • 4. Signaling Pathways Linking Actin Reorganization and Cell Volume Regulation231
  • 5. Quantitative Assessment of Cellular Actin Cytoskeleton Dynamics232
  • 5.1. Overview232
  • 6. Quantification of Cellular Monomeric and Total Actin Using the DNase I Inhibition Assay233
  • 7. DNase I Inhibition Assay Protocol234
  • 8. Quantification of Filamentous Actin Using Rhodamine-Phalloidin Fluorescence Measurements of Actin235
  • 9. Filamentous (F-) Actin Quantification Protocol235
  • 10. Quantitative Immunoblot Analysis of Triton X-100 Insoluble Cytoskeletal Pellets and Correspondin236
  • 11. Triton Soluble/Insoluble Actin Ratio Determination236
  • 11.1. Protocol I236
  • 11.2. Protocol II237
  • References237
  • Chapter 13: Osmotic Stress and DNA Damage241
  • 1. Introduction242
  • 2. Detection of DNA Damage by Comet Assay242
  • 2.1. Overview of the technique242
  • 2.2. Alkaline comet assay243
  • 2.3. Measurements of oxidative base damage by the comet assay245
  • 2.4. Detection of DNA double-strand breaks by the comet assay245
  • 3. Detection of DNA Breaks by In Situ End Labeling246
  • 3.1. Overview of techniques246
  • 3.2. Labeling of DNA breaks in paraffin-embedded tissue sections by the TUNEL assay247
  • 3.3. Apoptotic vs nonapoptotic DNA breaks: technical considerations for increasing sensitivity to de249
  • References250
  • Chapter 14: Transcriptional Activator TonE-Binding Protein in Cellular Protection and Differentiatio253
  • 1. Introduction254
  • 2. Immunological Detection of TonEBP255
  • 2.1. Immunoblotting256
  • 2.2. Single immunoprecipitation256
  • 2.3. Coimmunoprecipitation257
  • 2.4. Immunofluorescence detection in cultured cells257
  • 2.5. Immunohistochemical detection of TonEBP in the kidney by preembedding technique257
  • 3. Functional Analyses of TonEBP258
  • 3.1. TonEBP-driven luciferase reporter gene258
  • 3.2. Overexpression of TonEBP and DN-TonEBP259
  • 3.3. RNA interference of TonEBP259
  • 3.4. Electrophoretic mobility shift assay (EMSA)260
  • 4. How to Identify TonEBP Target Genes261
  • 4.1. Primer extension analysis263
  • 4.2. Construction of promoter-reporter263
  • 5. Biochemical Analysis of TonEBP265
  • Acknowledgments265
  • References266
  • Chapter 15: Desiccation Response of Mammalian Cells: Anhydrosignaling269
  • 1. Introduction270
  • 2. Cell Cultures272
  • 3. Desiccation273
  • 4. Cell Viability Test273
  • 5. Western Blotting273
  • 6. Relative Quantification of Gene Expression275
  • Acknowledgment276
  • References63
  • Chapter 16: Tonicity-Regulated Gene Expression279
  • 1. Introduction280
  • 1.1. Genes that TonEBP/OREBP transactivates in response to increased tonicity281
  • 1.2. Mechanisms of tonicity-dependent activation of TonEBP/OREBP282
  • 1.3. Cell culture285
  • 1.4. Transfection285
  • 2. Quantitation of the Nuclear:Cytoplasmic Distribution Ratio of the Transcription Factor TonEBP/ORE286
  • 2.1. Overview286
  • 2.2. Nuclear:Cytoplasmic distribution ratio protocol286
  • 3. Nonradioactive EMSA of Nuclear Extracts of HEK293 Cells for Binding of the Complex of TonEBP/OREB288
  • 3.1. Overview288
  • 3.2. Nuclear protein extraction288
  • 3.3. Electromobility shift assay with supershift288
  • 4. Quantitation of Transcriptional and Transactivational Activities of TonEBP/OREBP Using Transient290
  • 4.1. Reporter constructs290
  • 4.2. Overview291
  • 4.3. Transient transfections291
  • 4.4. Stable transfection292
  • 4.5. Reporter assays using stably transfected reporter cells293
  • Acknowledgment293
  • References293
  • Chapter 17: Hyperosmotic Induction of Mitogen-Activated Protein Kinase Scaffolding297
  • 1. Introduction298
  • 2. RNA Interference300
  • 2.1. Electroporation of siRNA into adherent cells301
  • 2.2. Transfection of siRNA into adherent cells301
  • 3. In Vitro Kinase Assays302
  • 3.1. MEKK3 assay302
  • 3.2. p38 MAPK assay303
  • 4. Measuring Dynamic Interactions Between MAPK Scaffold Members Using FRET303
  • 4.1. Considerations for constructing fluorescent chimeric proteins304
  • 4.2. Selecting a cell system to answer meaningful biological questions305
  • 4.3. Microscope configuration306
  • 4.4. Cell culture306
  • 4.5. Setting image acquisition parameters307
  • 4.6. Image acquisition307
  • 4.7. Image processing308
  • 4.8. Determining spectral cross talk coefficients308
  • 4.9. Calculating FRET indices308
  • 4.10. Data interpretation309
  • References310
  • Chapter 18: Osmoregulation of Bile Formation313
  • 1. Networks of Transport Systems314
  • 2. Osmoregulation of Canalicular Secretion315
  • 3. Osmosensing and Osmosignaling Pathways Toward Canalicular Secretion317
  • 4. Osmoregulation of Sinusoidal Bile Acid Uptake318
  • 5. Methods Used to Study Osmoregulated Transporter Insertion/Retrieval318
  • 6. Concluding Remarks320
  • Acknowledgments63
  • References63
  • Chapter 19: Osmosignaling and Volume Regulation in Intestinal Epithelial Cells325
  • 1. Introduction326
  • 2. Properties of Ion Channels and.Transporters Activated During RVD in Intestinal Epithelial Cells326
  • 3. Osmosensing and Signaling328
  • 4. Model Systems330
  • 5. Measuring Ionic Responses331
  • 5.1. Whole cell patch clamp331
  • 5.2. Isotope efflux assay333
  • 5.3. Ussing chamber experiments335
  • 5.4. Fluorometric quantification of intracellular ion concentrations337
  • 6. Concluding Remarks338
  • References338
  • Chapter 20: Osmotic Regulation of Cellular Glucose Uptake343
  • 1. Introduction344
  • 2. Hyperosmolarity and Glucose Transport347
  • 2.1. Differentiation of 3T3-L1 adipocytes347
  • 2.2. Glucose uptake induced in response to.hyperosmolarity347
  • 2.3. Study of pathways involved in the hyperosmotic effect on glucose uptake348
  • 2.4. Inhibition of insulin-stimulated glucose uptake by hyperosmotic stress349
  • 3. Hyperosmolarity and Membrane Ruffling349
  • 3.1. Membrane ruffling assay349
  • 4. Hyperosmolarity and Signaling Pathways350
  • 4.1. Preparation of total cell lysates350
  • 4.2. Immunoprecipitation of docking proteins (Gab1 or IRS1)351
  • 4.3. Western blotting assays351
  • 5. Hyperosmolarity and Phosphatidylinositol 3-Kinase Activity351
  • 5.1. Preparation of phosphatidylinositol352
  • 5.2. Immunoprecipitation of phosphatidylinositol 3-kinase352
  • 5.3. Measurement of PI 3-kinase activity in immunoprecipitation353
  • 6. Conclusion353
  • Acknowledgments353
  • References354
  • Chapter 21: Effects of Osmolytes on Protein Folding and Aggregation in Cells355
  • 1. Introduction356
  • 2. Analysis of Effects of Osmolytes In Vivo358
  • 2.1. Culture growth and induction of osmolyte uptake in vivo360
  • 2.2. Partitioning into insoluble and soluble cell fractions361
  • 2.3. Variations in the effect of different osmolytes in vivo362
  • 3. Comparison of the Influence of Osmolytes In Vivo with That In Vitro367
  • 3.1. Expression and purification367
  • 3.2. In vitro stability measurements368
  • 3.3. In vitro aggregation kinetics368
  • 3.4. Different osmolytes have different effects on folding and aggregation in vitro368
  • 4. Conclusions370
  • Acknowledgments370
  • References370
  • Chapter 22: Simulations of Macromolecules in Protective and Denaturing Osmolytes: Properties of Mixe63
  • 1. Introduction374
  • 2. Methods375
  • 2.1. Urea376
  • 2.2. Trimethylamine N-oxide376
  • 2.3. Chymotrypsin inhibitor 2376
  • 2.4. Molecular dynamics simulations377
  • 2.5. Analyses377
  • 3. Results379
  • 3.1. Pure water379
  • 3.2. Urea in water379
  • 3.3. Trimethylamine N-oxide in water382
  • 3.4. Chymotrypsin inhibitor 2 in water, urea, and TMAO386
  • 4. Discussion389
  • 5. Conclusions392
  • Acknowledgments393
  • References393
  • Chapter 23: Application of the Transfer Model to Understand How Naturally Occurring Osmolytes Affect397
  • 1. Introduction398
  • 2. Initial Observations of Osmolyte Effects on Proteins398
  • 3. The Transfer-Free Energy Model401
  • 4. How Transfer-Free Energies Are Measured403
  • 5. The Issue of Activity Coefficients: Solution Nonideality at the Solubility Limit406
  • 6. Additive, Concentration Scale-Independent and Model Compound-Independent Transfer-Free Energies o408
  • 7. The Transfer-Free Energy of Amino Acid Side Chains412
  • 8. Solvent-Accessible Surface Areas of Native and Denatured States: Calculating Deltaalphai413
  • 9. Putting It All Together: Predicting m Values for the Energetics of Osmolyte-Induced Protein Foldi414
  • 10. Summary415
  • Acknowledgments
  • References416
  • Chapter 24: Mechanisms of High Salinity Tolerance in Plants419
  • 1. Introduction420
  • 2. Adverse Effect of Salinity Stress421
  • 3. Generic Pathway for Plant Response to Stress422
  • 4. Ion Pumps, Calcium, and SOS Pathways in Relation to Salinity Stress424
  • 5. Abscisic Acid and Transcription Factors in Salinity Stress Tolerance427
  • 6. Mitogen-Activated Protein Kinases and Salinity Stress430
  • 7. Glycine Betaine and Proline in Salinity Stress430
  • 8. Reactive Oxygen Species in Salinity Stress431
  • 9. DEAD-Box Helicases in Salinity Stress Tolerance432
  • 10. Cross-Tolerances Between Stresses434
  • 11. Future Prospective435
  • Acknowledgments
  • References
  • Chapter 25: Phenotype of the Taurine Transporter Knockout Mouse439
  • 1. Targeted Disruption of the taut Gene440
  • 2. Reduced Taurine Levels Lead to Various Diseases in taut-/- Mice441
  • 3. Reduced Exercise Capacity in taut-/- Mice442
  • 4. Pathophysiological Changes in Brain, Retina, Olfactory Bulb, and Inner Ear of taut-/- Mice444
  • 4.1. Deficit in taurine-evoked synaptic enhancement and changes in neuroreceptor expression in taut-444
  • 4.2. Apoptosis leads to retinal degeneration in taut-/- mice445
  • 4.3. Olfactory dysfunction of taut-/- mice447
  • 4.4. Age-dependent loss of hearing in taut-/- mice448
  • 5. Renal Function in taut-/- Mice448
  • 6. Chronic Liver Disease in taut-/- Mice449
  • 7. Available Organic Osmolyte Transporter Transgene and Knockout Mice452
  • References452
  • Chapter 26: Molecular Basis of Osmolyte Effects on Protein and Metabolites459
  • 1. Introduction460
  • 1.1. Importance of chemical activities for life460
  • 1.2. Solvation vs binding460
  • 1.3. Solvation, solution structure, and water structure461
  • 1.4. Overall solvation derived from pair correlations462
  • 1.5. Experimental data useful for deriving thermodynamic solvation463
  • 1.6. Useful approximations464
  • 2. General Kirkwood-Buff Equations467
  • 2.1. Working in the molal scale467
  • 2.2. Elimination of parameters based on the given system468
  • 2.3. Working in the molar scale468
  • 3. Case 1: Osmolyte Solvation470
  • 3.1. Derivation of equations470
  • 3.2. Application of equations472
  • 4. Case 2: m Values474
  • 4.1. Derivation of equations474
  • 4.2. Interpretation476
  • 5. Case 3: Specific Binding of an Osmolyte479
  • 5.1. Specific binding as a solvation phenomenon479
  • 5.2. Specific binding explicitly taken into account480
  • 6. Case 4: Specific Binding of a Dilute Ligand482
  • Acknowledgments483
  • References484
  • Chapter 27: Methods of Changing Biopolymer Volume Fraction and Cytoplasmic Solute Concentrations for487
  • 1. Introduction488
  • 2. Varying Biopolymer Volume Fraction and.Cytoplasmic Solute Concentrations Osmotically488
  • 3. Biophysical Studies In Vivo Using Permeable Solutes: Protein Unfolding by Urea Titration495
  • 4. Effects of Biopolymer Volume Fraction and/or Other In Vivo Factors on Protein Diffusion in the E.498
  • 5. Summary501
  • Acknowledgments501
  • References501
  • Chapter 28: Characterization of Plant Aquaporins505
  • 1. Molecular Function of Plant Aquaporins506
  • 1.1. Aquaporin function in plant membranes507
  • 1.2. Analysis of plant aquaporins in heterologous and artificial expression systems509
  • 1.3. Single cell analysis514
  • 2. Aquaporin Function in Plants517
  • 2.1. Expression analysis517
  • 2.2. Modification of aquaporin expression518
  • 2.3. Hydraulic conductivity of plant organs and single cells521
  • 2.4. Development523
  • 2.5. Water and salt stress524
  • 2.6. Photosynthesis525
  • Acknowledgment526
  • References526
  • Author Index533
  • Subject Index571
Book details
  • Vendor Elsevier S & T
  • SKU 9780123739216
  • ISBN-13 9780080552118
  • Author Sies, Helmut
  • Category Science
  • Subject Molecular Biology

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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 osmosensing and osmosignaling written by experts in the field.