The Role of Protein-RNA Interactions in 3' Splice Site Selection During Pre-mRNA Splicing PDF Download
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Author: James G. Umen
Publisher:
ISBN:
Category : Protein binding
Languages : en
Pages : 596
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Book Description
Author: James G. Umen
Publisher:
ISBN:
Category : Protein binding
Languages : en
Pages : 596
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Book Description
Author: Karli Lipinski
Publisher:
ISBN:
Category :
Languages : en
Pages : 0
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Book Description
Pre-mRNA splicing is a fundamental process governing gene expression in eukaryotes and is orchestrated by the spliceosome, a dynamic ribonucleoprotein complex. Spliceosomes catalyze splicing in two steps and are composed of both small nuclear RNAs (snRNAs; U1, U2, U4, U5, U6) and numerous proteins. Dynamic interactions between snRNAs, proteins, and the pre-mRNA substrate occur during splicing which are central to regulating splice site recognition, catalysis, and fidelity. This thesis examines multiple facets of RNA-protein interactions within spliceosome complexes beginning with snRNA biogenesis, through formation of the spliceosome active site, and during the second step of chemistry, exon ligation. Functional mechanisms into splicing, such as selection of intron recognition sites, are examined. Function of the main catalytic component of the spliceosome, the U6 snRNA, is first explored. Transcription and post-transcriptional processing of U6 is unique among snRNAs and therefore may be required to generate a functional U6. Transcription by RNAP II instead of RNAP III produces a functional U6 molecule. Defects in stability, likely a result of incorrect post-transcriptional processing and binding of stabilizing proteins, result in changes to in vivo distributions of spliceosome sub particles called snRNPs. Transcription of U6 by RNAP II is useful for the incorporation of genetic tags for endogenous fluorescent labeling or purification. Additionally, new applications of endogenous fluorescent labeling techniques within the U4 snRNA are presented, paving the way for single molecule studies of snRNA dynamics and Brr2 helicase function during activation. Mango and MS2 tags are well tolerated in the yeast U4 snRNA and with several of the tagged U4 constructs also minimally impacting splicing activities. Future single molecule experiments will examine the timing of the U4 snRNA release from the spliceosome compared to the release of Prp3, a protein associated with U6 and U4 snRNAs. Finally, validation of a proposed novel splicing factor, Fyv6, and study of its influence on 3' SS represent major contributions to the field of splicing. Utilizing a new high resolution P complex structure of the spliceosome containing Fyv6 solved by Max Wilkinson, I examined multiple contacts of Fyv6 with other splicing factors, notably Prp22. Genetic studies in yeast show that interactions with the protein Syf1 and Prp22 are important for Fyv6 function. The absence of Fyv6 from spliceosomes results in transcriptome-wide splicing defects, largely due to changes in 3' SS usage. From these studies, Fyv6 can be added to the list of splicing factors that impact the second step of splicing and affect fidelity of 3' SS selection.
Author: Philippe Jeanteur
Publisher: Springer Science & Business Media
ISBN: 3540344497
Category : Science
Languages : en
Pages : 265
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Book Description
Splicing of primary RNA transcript is a quasi-systematic step of gene expression in higher organisms. This is the first book to highlight the medical implications, i.e. diseases, caused by alternative splicing. Alternative splicing not only vastly increases protein diversity but also offers numerous opportunities for aberrant splicing events with pathological consequences. The book also outlines possible targets for therapy.
Author: B. D. Hames
Publisher: Oxford University Press, USA
ISBN:
Category : Music
Languages : en
Pages : 238
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Book Description
This book gives a co-ordinated review of our present knowledge of eukaryotic RNA synthesis.
Author: Susan R. Haynes
Publisher: Springer Science & Business Media
ISBN: 1592596762
Category : Science
Languages : en
Pages : 485
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Book Description
The molecular characterization of RNA and its interactions with proteins is an important and exciting area of current research. Organisms utilize a variety of RNA–protein interactions to regulate the expression of their genes. This is particularly true for eukaryotes, since newly synthesized messenger RNA must be extensively modified and transported to the cytoplasm before it can be used for protein synthesis. The realization that posttranscriptional processes are critical components of gene regulation has sparked an explosion of interest in both stable ribonucleoprotein (RNP) complexes and transient RNA–protein interactions. RNA is conformationally flexible and can adopt complex structures that provide diverse surfaces for interactions with proteins. The fact that short RNA molecules (aptamers; see Chapter 16) can be selected to bind many different types of molecules is evidence of the structural variability of RNA. RNA molecules are rarely entirely single- or double-stranded, but usually contain multiple short duplexes interrupted by single-stranded loops and bulges; in some RNAs, such as tRNAs, the short duplexes stack on each other. Further variability is generated by the presence of non-Watson-Crick base pairs, modified nucleotides, and more complex structures, such as pseudoknots and triple-strand interactions.
Author: Frank J. Schmidt
Publisher: Humana Press
ISBN: 9781493918959
Category : Medical
Languages : en
Pages : 0
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Book Description
In this volume expert researchers in the field detail many of the methods which are now commonly used to study RNA. These methods are presented as a guidebook to scientists who are experienced with RNA research and want to brush up on a new technique. Written in the highly successful Methods in Molecular Biology series format, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and key tips on troubleshooting and avoiding known pitfalls. Thorough and intuitive, RNA-RNA Interactions: Methods and Protocols guides scientists investigating biological systems and studying RNA.
Author: Christopher W.J. Smith
Publisher: Oxford University Press, UK
ISBN: 0191591629
Category :
Languages : en
Pages : 370
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Book Description
RNA-protein interactions play a fundamental role in gene expression and protein synthesis. Recent research into the role of RNA in cells has elucidated many more vital interactions with proteins. This book provides an up-to-date and comprehensive guide to a wide range of laboratory procedures to investigate the interactions between RNA and proteins. - ;RNA-protein interactions play a vital role in gene transcription and protein expression. Interactions such as the synthesis of mRNA by RNA polymerases, to the essential modification of RNA by the proteins of the spliceosome complex, and the highly catalytic action of the ribosome in protein synthesis, are established as being fundamental to the function of RNA. Recent research into, for example, the role of RNA as a catalyst, has elucidated many more interactions with proteins that are vital to cell function. RNA - Protein Interactions: A Practical Approach provides a clear and comprehensive guide to the experimental procedures used in studying RNA - protein interactions. The approaches covered range from those initially used to detect a novel RNA-protein interaction, various biochemical and genetic approaches to purifying and cloning RNA binding proteins, through to methods for an in depth analysis of the structural basis of the interaction. The volume includes a number of procedures that have not previously been covered in this type of manual. These include the production of site-specifically modified RNAs by enzymatic and chemical methods and in vivo screening for novel RNA - protein interactions in yeast and E. coli . This is the first volume to gather in one place this wide array of approaches for studying RNA - protein interactions. As is customary for the Practical Approach series, the writing is characterized by a clear explanatory style with many detailed protocols. This informative book will be a valuable aid to laboratory workers in biochemistry and molecular biology - graduate students, postdoctoral and senior scientists - whose research encompasses this field. -
Author: Rosanna Asselta
Publisher: Frontiers Media SA
ISBN: 2889662357
Category : Science
Languages : en
Pages : 197
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Book Description
This eBook is a collection of articles from a Frontiers Research Topic. Frontiers Research Topics are very popular trademarks of the Frontiers Journals Series: they are collections of at least ten articles, all centered on a particular subject. With their unique mix of varied contributions from Original Research to Review Articles, Frontiers Research Topics unify the most influential researchers, the latest key findings and historical advances in a hot research area! Find out more on how to host your own Frontiers Research Topic or contribute to one as an author by contacting the Frontiers Editorial Office: frontiersin.org/about/contact.
Author: Deming Xu
Publisher:
ISBN:
Category :
Languages : en
Pages : 0
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Book Description
Nuclear pre-mRNA splicing proceeds in a protein-RNA complex, the spliceosome, via a two-step transesterification reaction. Five spliceosomal small nuclear RNAs; (snRNAs) are brought to the pre-mRNA substrate in a process known as spliceosome assembly. Extensive RNA-RNA interactions and conformational changes occur during spliceosome assembly and maturation. However, very little is known about how RNA conformational changes are coordinated between the two steps of the splicing reaction, and how recognition of the 3' splice site by other factors is connected to the function of U5 loop 1 prior to and following the first splicing step. I performed a genetic screen in order to identify factors that are important for the U2/U6 helix II interaction and/or the function of the 5' -end of U2 snRNA, on the basis of synthetic lethality with an 11nt nucleotide substitution in the 5'-end region of U2 snRNA that could also perturb the U2/U6 helix II interaction. Six Slt (synthetic lethal with U2 snRNA) factors were isolated in this screen. Slt11p and Slt22p are new splicing factors, while the remainder correspond to previously identified splicing factors, Slt15p/Prp17p, Slt16p/Smd3p, Slt17p/Slu7p and Slt21p/Prp8p. Slt22p is a large RNA-dependent ATPase, whose activity is preferentially stimulated by pre-annealed U2/U6 snRNAs. Biochemical and genetic analyses indicated that its function is associated with U2/U6 snRNA interaction and that it may unwind intermolecular helix II prior to the formation of the active spliceosome. Slt2lp/Prp8p has been shown to be required for both steps of the splicing reaction. The 'slt'21/'prp'8-21 mutation has been mapped to a domain in Prp8p that is important for the recognition of the polypyrimidine tract preceding the 3' splice site. It is synthetically lethal with mutations in only the U2 part of U2/U6 helix II. Consistent with the notion that the 5'-end of U2 snRNA plays a role in the second splicing step, I found that U2 mutations in this region are synthetically lethal with several second-step mutations, including 'slt15'/'prp17-100', ' slu4'/'prp17-2', 'slt17'/' slu7-100', and 'slu7-1'. Slt11p is a putative RNA-binding protein containing two Zn-finger motifs. Although 'SLT11' is essential for viability only at elevated temperatures, its product is required for the efficiency of the splicing reaction. In the absence of Slt11p, 'slt17'/'slu7-100' and mutations in specific regions of U2, U5 and U6 snRNAs become lethal. (Abstract shortened by UMI.).
Author: Stefan Stamm
Publisher: John Wiley & Sons
ISBN: 3527326065
Category : Science
Languages : en
Pages : 660
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Book Description
Der definitive Leitfaden zum RNA-Splicing - ideal für alle Kliniker, die sich mit genetischen Erkrankungen befassen, insbesondere natürlich für RNA-Forscher. Website: www.wiley-vch.de/home/splicing
