RNA turnover in eukaryotes : nucleases, pathways and analysis of mRNA decay / edited by Lynne E. Maquat, Megerditch Kiledjian.

Format:

Book

Contributor:

Maquat, Lynne.

Kiledjian, Megerditch.

Series:

Methods in enzymology ; v. 448.

Methods in enzymology ; v. 448

Language:

English

Subjects (All):

Messenger RNA.

Genetic regulation.

Eukaryotic cells.

Nucleases.

Physical Description:

1 online resource (661 p.)

Other Title:

Nucleases, pathways and analysis of mRNA decay

Place of Publication:

San Diego, Calif. ; London : Academic, c2008.

Language Note:

English

Summary:

Specific complexes of protein and RNA carry out many essential biological functions, including RNA processing, RNA turnover, RNA folding, as well as the translation of genetic information from mRNA into protein sequences. Messenger RNA (mRNA) decay is now emerging as an important control point and a major contributor to gene expression. Continuing identification of the protein factors and cofactors, and mRNA instability elements responsible for mRNA decay allow researchers to build a comprehensive picture of the highly orchestrated processes involved in mRNA decay and its regulation.

Contents:

Front Cover; RNA Turnover in Eukaryotes: Nucleases, Pathways and Analysis of mRNA Decay; Copyright Page; Contents; Contributors; Preface; Volumes in Series; Section I: Decapping Analyses; Chapter 1: Analysis of mRNA Decapping; 1. Introduction; Acknowledgment; References; Chapter 2: A Kinetic Assay to Monitor RNA Decapping Under Single- Turnover Conditions; 1. Introduction; 3. Materials; Acknowledgments; Appendix; Chapter 3: Purification and Analysis of the Decapping Activator Lsm1p-7p-Pat1p Complex from Yeast; 1. Introduction; 2. Purification of the Lsm1p-7p-Pat1p Complex

3. Analysis of RNA Binding by the Lsm1p-7p-Pat1p Complex4. Concluding Remarks; Chapter 4: Reconstitution of Recombinant Human LSm Complexes for Biochemical, Biophysical, and Cell Biological Studies; 1. Introduction; 3. Protein Expression and Purification; 4. LSm Complex Reconstitution; 5. LSm Complex Functional Assays; 6. Outlook; Section II: Polyadenylation/Deadenylation Analyses; Chapter 5: Regulated Deadenylation in Vitro; 1. Introduction; 3. Troubleshooting; References; Chapter 6: Cell-Free Deadenylation Assays with Drosophila Embryo Extracts; 2. Preparation of Drosophila Embryo Extracts

3. Preparation of Substrate RNA5. Characterization of Sequence-Dependent Deadenylation in Drosophila Embryo Extracts; Acknowledgments; References; Chapter 7: Measuring CPEB-Mediated Cytoplasmic Polyadenylation-Deadenylation in Xenopus laevis Oocytes and Egg Extracts; 3. Detection of Nuclear and Cytoplasmic Polyadenylation of X. laevis Cyclin B1 mRNA; 5. Detection of Endogenous Cytoplasmic Poly(A) Polymerase Activity; 6. Overexpression of Exogenous mRNAs and Measuring Polyadenylation-Deadenylation

7. Preparation of X. laevis Egg Extracts and Measuring Polyadenylation-Deadenylation with Cyclin B1 RNAChapter 8: The Preparation and Applications of Cytoplasmic Extracts from Mammalian Cells for Studying Aspects of mRNA Decay; 2. Preparation of HeLa-Cell Cytoplasmic Extracts; 3. Preparation of RNA Substrates; 4. Evaluating mRNA Decay with Cytoplasmic Extracts; Acknowledgments; References; Section III: Nucleases in mRNA Decay; Chapter 9: In Vitro Assays of 5' to 3'-Exoribonuclease Activity; 1. Introduction; 2. Purification of Xrn1

4. Degradation of RNA by XRN1 Depends on the Nature of the 5'-End Chapter 10: Reconstitution of RNA Exosomes from Human and Saccharomyces Cerevisiae: Cloning, Expression, Purification, and Activity Assays; 2. Cloning Strategies for Recombinant Protein Expression; 3. PCR and Subcloning Protocols; 5. PCR and Subcloning Protocols for Human cDNA; 6. Expression and Purification of Yeast Exosome Proteins; 7. Expression and Purification of Human Exosome Proteins; 8. Reconstitution and Purification of Human and Yeast Exosomes; 9. Exoribonuclease Assays; 11. Conclusions; References

Chapter 11: Biochemical Studies of the Mammalian Exosome with Intact Cells

Notes:

Description based upon print version of record.

Includes bibliographical references and indexes.

ISBN:

9786612287312

9781282287310

1282287311

9780080922072

0080922074

OCLC:

808777999