Download or read book A Biochemical Characterization of Transcription Elongation Factors that Facilitate the Processivity of RNA Polymerase II written by John Crickard and published by . This book was released on 2016 with total page pages. Available in PDF, EPUB and Kindle. Book excerpt: The processivity of RNA Polymerase II across the body of a gene is a complex anddynamic process that involves the contribution of many transcription elongation factors.These elongation factors play a variety of roles including maintenance of RNAPII activity,maintenance of chromatin, and regulation of chromatin structure. The roles of elongationfactors highlight two strategies utilized by the cell to promote transit of RNAPII acrossthe body of a gene. The overall goal of this work was to gain mechanistic insight, basedon in vitro models, into RNAPII processivity, and how elongation factors contribute to thisprocess. In this dissertation, I demonstrate that a region of the H2B tail domain, knownas the H2B repression domain (HBR), helps Spt16-Pob3 (FACT) remove H2A/B dimersfrom the nucleosome. Then I show that Spt4/5, a universally conserved transcriptionelongation factor, maintains active RNAPII through interactions with the non-templatestrand of DNA. I also find that eukaryotic specific regions of Spt5 may recognize certainconformations of RNAPII. Finally, I identify a novel strategy utilized by Spt4/5 to act onspecific intermediate states to relocate RNAPII with in the nucleosome. This observationsuggests that nucleosomes may not simply act as a roadblock to transcription, but mayserve a regulatory function during transcription elongation. Together these findingsadvance our understanding of how elongation factors contribute to the processivity ofRNAPII, and how that effects gene regulation.
Download or read book Biochemical Characterization of RNA Polymerase II written by Yuichiro Takagi and published by . This book was released on 1997 with total page 230 pages. Available in PDF, EPUB and Kindle. Book excerpt:
Download or read book Structural and Biochemical Studies of the Transcription Termination Machinery written by Peter Hsu and published by . This book was released on 2013 with total page 118 pages. Available in PDF, EPUB and Kindle. Book excerpt: RNA Polymerase II (PolII) dependent transcription of mRNAs is central to gene expression throughout eukaryotes. Transcription is a highly regulated process, with a defined initiation, elongation, and termination phase, all of which are controlled by multiple trans-acting protein factors and cis-acting elements on the template DNA and transcribed RNA. Extensive work has shown that the phosphorylation state of the C- terminal domain (CTD) of PolII plays a central role in the recruitment of trans-acting factors during all phases of transcription. Aberrant phosphorylation can result in a lethal phenotype in yeasts, therefore implying that correct control of phosphorylation by kinases and phosphatases specific for the CTD is critical for life. In addition to the polymerase, conserved cis-acting sequence elements near the 3'-end on pre-mRNAs help to define and recruit various RNA processing machines to the transcription elongation complex in order to properly process and package the pre-mRNA for export to the cytoplasm for translation. In this thesis, I first review current knowledge regarding PolII CTD phosphorylation and its effects on the transcription elongation complex. Additionally, in this first chapter, I will also provide an overview of the 3'-end mRNA processing/transcription termination machinery. In the second part of my thesis, I will describe my doctoral work on the structural and biochemical characterization of Rtr1, a unique PolII CTD phosphatase that represents a novel new member of this class of enzymes. In my studies, I show that Rtr1 is a bona fide phosphatase of unique sequence and structure that is allosterically regulated by its own C-terminus. Additionally, I show that Rtr1 is a dual specificity phosphatase, with activities against both serine and tyrosine residues on the CTD. In chapter 3 of this thesis, I describe my work on the in vitro reconstitution of the Cleavage Stimulation Factor (CstF) responsible for the recognition of sequences downstream of the polyadenylation site on pre-mRNAs that help to define the 3'-end processing reaction that occur at the end of genes. Using highly purified proteins, I show for the first time that CstF is a dimer of trimers, with two copies of each subunit in the entire assembly. In addition, I show that CstF, as a complex, can bind to G/U rich RNAs with nanomolar affinities, in stark contrast with previous studies showing that singly purified proteins from the complex binding with much weaker affinities.
Download or read book Characterization of an RNA Polymerase II Subunit RPB9 and a Transcript Elongation Factor TFIIS from Saccharomyces Cerevisiae written by Rodney Gerard Weilbaecher and published by . This book was released on 1997 with total page 536 pages. Available in PDF, EPUB and Kindle. Book excerpt:
Download or read book Biochemical Characterization of the Interactions Between a Transcription Factor CTCF and Its Partners Y Box Binding Protein 1 and the Large Subunit of RNA Polymerase II written by Shaharum Shamsuddin and published by . This book was released on 2002 with total page 592 pages. Available in PDF, EPUB and Kindle. Book excerpt:
Download or read book Biochemical Characterization of the Interactions Between a Transcription Factor CTCF and Its Partners Y Box Binding Protein 1 and the Large Subunit of RNA Polymerase II written by Shaharum Shamsuddin and published by . This book was released on 2002 with total page 0 pages. Available in PDF, EPUB and Kindle. Book excerpt:
Download or read book Crystal Structure and Biochemical Analysis of Archaeal RNA Polymerase Transcription Elongation Factors written by Brianna Joy Klein and published by . This book was released on 2010 with total page 185 pages. Available in PDF, EPUB and Kindle. Book excerpt:
Download or read book Molecular Mechanisms of Factors that Control RNA Polymerase II Transcription Elongation Dynamics written by Manchuta Dangkulwanich and published by . This book was released on 2015 with total page 137 pages. Available in PDF, EPUB and Kindle. Book excerpt: The expression of a gene begins by transcribing a target region on the DNA to form a molecule of messenger RNA. As transcription is the first step of gene expression, it is there- fore highly regulated. The regulation of transcription is essential in fundamental biological processes, such as cell growth, development and differentiation. The process is carried out by an enzyme, RNA polymerase, which catalyzes the addition of a nucleotide complementary to the template and moves along the DNA one base pair at a time. To complete its tasks, the enzyme functions as a complex molecular machine, possessing various evolutionarily designed parts. In eukaryotes, RNA polymerase has to transcribe through DNA wrapped around histone proteins forming nucleosomes. These structures represent physical barriers to the transcribing enzyme. In chapter 2, we investigated how each nucleosomal component--the histone tails, the specific histone-DNA contacts, and the DNA sequence--contributes to the strength of the barrier. Removal of the tails favors progression of RNA polymerase II into the entry region of the nucleosome by locally increasing the wrapping-unwrapping rates of the DNA around histones. In contrast, point mutations that affect histone-DNA contacts at the dyad abolish the barrier to transcription in the central region by decreasing the local wrapping rate. Moreover, we showed that the nucleosome amplifies sequence-dependent transcriptional pausing, an effect mediated through the structure of the nascent RNA. Each of these nucleosomal elements controls transcription elongation by distinctly affecting the density and duration of polymerase pauses, thus providing multiple and alternative mechanisms for control of gene expression by additional factors. During transcription elongation, RNA polymerase has been assumed to attain equilibrium between pre- and post-translocated states rapidly relative to the subsequent catalysis. Under this assumption, a branched Brownian ratchet mechanism that necessitates a putative secondary nucleotide binding site on the enzyme was proposed. In chapter 3, we challenged individual yeast RNA polymerase II (Pol II) with a nucleosome as a "road block", and separately measured the forward and reverse translocation rates with our single-molecule transcription elongation assay. Surprisingly, we found that the forward translocation rate is comparable to the catalysis rate. This finding reveals a linear, non-branched ratchet mech-anism for the nucleotide addition cycle in which translocation is one of the rate-limiting steps. We further determined all the major on- and off-pathway kinetic parameters in the elongation cycle. This kinetic model provides a framework to study the influence of various factors on transcription dynamics. To further dissect the operation of Pol II, we focused on the trigger loop, a mobile element near the active site of the enzyme. Biochemical and structural studies have demonstrated that the trigger loop makes direct contacts with substrates and promotes nucleotide incorporation. It is also an important regulatory element for transcription fidelity. In chapter 4, we characterized the dynamics of a trigger loop mutant RNA polymerase to elucidate the roles of this element in transcription regulation, and applied the above kinetic framework to quantify the effects of the mutation. In comparison to the wild-type enzyme, we found that the mutant is more sensitive to force, faster at substrate sequestration, and more efficient to return from a pause to active transcription. This work highlighted important roles of regulatory elements in controlling transcription dynamics and fidelity. Moreover, RNA polymerase interacts with various additional factors, which add layers of regulation on transcription. Transcription factors IIS (TFIIS) and IIF (TFIIF) are known to interact with elongating RNA polymerase directly and stimulate transcription. In chapter 5, we studied the effects of these factors on elongation dynamics using our single molecule assay. We found that both TFIIS and TFIIF enhance the overall transcription elongation by reducing the lifetime of transcriptional pauses and that TFIIF also decreases the probability of pause entry. Furthermore, we observed that both factors enhance the efficiency of nucleosomal transcription. Our findings helped elucidate the molecular mechanisms of gene expression modulation by transcription factors. In summary, we have dissected the mechanisms by which the nucleosomal elements regulate transcription, and derived a quantitative kinetic model of transcription elongation in a linear Brownian ratchet scheme with the slow translocation of the enzyme. The corresponding translocation energy landscape shows that the off-pathway states are favored thermodynamically but not kinetically over the on-pathway states. This observation confers the enzyme its high propensity to pause, thus allowing additional regulatory mechanisms during pausing. TFIIS and TFIIF, for example, regulate transcription dynamics by shortening the lifetime of Pol II pauses. On the other hand, the trigger loop of Pol II regulates both the active elongation and pausing. These examples illustrate molecular mechanisms of cis- and trans-acting factors regulate the dynamics of transcription elongation.
Download or read book RNA Exosome written by Torben Heick Jensen and published by Springer Science & Business Media. This book was released on 2011-06-29 with total page 161 pages. Available in PDF, EPUB and Kindle. Book excerpt: The diversity of RNAs inside living cells is amazing. We have known of the more “classic” RNA species: mRNA, tRNA, rRNA, snRNA and snoRNA for some time now, but in a steady stream new types of molecules are being described as it is becoming clear that most of the genomic information of cells ends up in RNA. To deal with the enormous load of resulting RNA processing and degradation reactions, cells need adequate and efficient molecular machines. The RNA exosome is arising as a major facilitator to this effect. Structural and functional data gathered over the last decade have illustrated the biochemical importance of this multimeric complex and its many co-factors, revealing its enormous regulatory power. By gathering some of the most prominent researchers in the exosome field, it is the aim of this volume to introduce this fascinating protein complex as well as to give a timely and rich account of its many functions. The exosome was discovered more than a decade ago by Phil Mitchell and David Tollervey by its ability to trim the 3’end of yeast, S. cerevisiae, 5. 8S rRNA. In a historic account they laid out the events surrounding this identification and the subsequent birth of the research field. In the chapter by Kurt Januszyk and Christopher Lima the structural organization of eukaryotic exosomes and their evolutionary counterparts in bacteria and archaea are discussed in large part through presentation of structures.
Download or read book Molecular Biology of B Cells written by Tasuku Honjo and published by Academic Press. This book was released on 2014-12-22 with total page 0 pages. Available in PDF, EPUB and Kindle. Book excerpt: Molecular Biology of B Cells, Second Edition is a comprehensive reference to how B cells are generated, selected, activated and engaged in antibody production. All of these developmental and stimulatory processes are described in molecular, immunological, and genetic terms to give a clear understanding of complex phenotypes. Molecular Biology of B Cells, Second Edition offers an integrated view of all aspects of B cells to produce a normal immune response as a constant, and the molecular basis of numerous diseases due to B cell abnormality. The new edition continues its success with updated research on microRNAs in B cell development and immunity, new developments in understanding lymphoma biology, and therapeutic targeting of B cells for clinical application. With updated research and continued comprehensive coverage of all aspects of B cell biology, Molecular Biology of B Cells, Second Edition is the definitive resource, vital for researchers across molecular biology, immunology and genetics.
Download or read book Molecular Biology of the Cell written by and published by . This book was released on 2002 with total page 0 pages. Available in PDF, EPUB and Kindle. Book excerpt:
Download or read book Long Noncoding RNAs in Plants written by Santosh Kumar Upadhyay and published by Academic Press. This book was released on 2020-11-13 with total page 392 pages. Available in PDF, EPUB and Kindle. Book excerpt: The growth of human population has increased the demand for improved yield and quality of crops and horticultural plants. However, plant productivity continues to be threatened by stresses such as heat, cold, drought, heavy metals, UV radiations, bacterial and fungal pathogens, and insect pests. Long noncoding RNAs are associated with various developmental pathways, regulatory systems, abiotic and biotic stress responses and signaling, and can provide an alternative strategy for stress management in plants. Long Noncoding RNAs in Plants: Roles in development and stress provides the most recent advances in LncRNAs, including identification, characterization, and their potential applications and uses. Introductory chapters include the basic features and brief history of development of lncRNAs studies in plants. The book then provides the knowledge about the lncRNAs in various important agricultural and horticultural crops such as cereals, legumes, fruits, vegetables, and fiber crop cotton, and their roles and applications in abiotic and biotic stress management. - Includes the latest advances and research in long noncoding RNAs in plants - Provides alternative strategies for abiotic and biotic stress management in horticultural plants and agricultural crops - Focuses on the application and uses of long noncoding RNAs
Download or read book Fundamentals of Molecular Structural Biology written by Subrata Pal and published by Academic Press. This book was released on 2019-08-15 with total page 518 pages. Available in PDF, EPUB and Kindle. Book excerpt: Fundamentals of Molecular Structural Biology reviews the mathematical and physical foundations of molecular structural biology. Based on these fundamental concepts, it then describes molecular structure and explains basic genetic mechanisms. Given the increasingly interdisciplinary nature of research, early career researchers and those shifting into an adjacent field often require a "fundamentals" book to get them up-to-speed on the foundations of a particular field. This book fills that niche.
Download or read book Protein Phosphorylation in Human Health written by Cai Huang and published by BoD – Books on Demand. This book was released on 2012-09-06 with total page 482 pages. Available in PDF, EPUB and Kindle. Book excerpt: 15 chapters on protein phosphorylation and human health written by expert scientists. Covers most important research hot points, such as Akt, AMPK and mTOR. Bridges the basic protein phosphorylation pathways with human health and diseases. Detailed and comprehensive text with excellent figure illustration.
Download or read book Nature written by Sir Norman Lockyer and published by . This book was released on 2002 with total page 1568 pages. Available in PDF, EPUB and Kindle. Book excerpt:
Download or read book Plant RNA Biology written by Dóra Szakonyi and published by Frontiers Media SA. This book was released on 2019-11-18 with total page 167 pages. Available in PDF, EPUB and Kindle. Book excerpt: Discoveries from the past decades revealed that RNA molecules are much more than inert intermediates between the coding DNA sequences and their functional products, proteins. Today, RNAs are recognized as active regulatory molecules influencing gene expression, chromatin organization and genome stability, thus impacting all aspects of plant life including development, growth, reproduction and stress tolerance. Innovations in methodologies, the expanding application of next-generation sequencing technologies, and the creation of public datasets and databases have exposed a new universe of RNA-based mechanisms and led to the discovery of new families of non-coding RNAs, uncovered the large extent of alternative splicing events, and highlighted the potential roles of RNA modifications and RNA secondary structures. Furthermore, considerable advances have been made in identifying RNA-binding and processing factors involved in the synthesis and maturation of different forms of RNA molecules as well as in RNA processing, biochemical modifications or degradation. This Research Topic showcases the broad biological significance of RNAs in plant systems and contains eight original research articles, one review and four mini-reviews, covering various RNA-based mechanisms in higher plants. Emerging new technologies and novel multidisciplinary approaches are empowering the scientific community and will expectedly bring novel insights into our understanding of the mechanisms through which RNA is regulated and regulates biological processes in plant cells.
Download or read book Chromatin Immunoprecipitation Assays written by Philippe Collas and published by Methods in Molecular Biology. This book was released on 2009-08-25 with total page 290 pages. Available in PDF, EPUB and Kindle. Book excerpt: In this book, researchers deeply involved in the development and improvement of chromatin immunoprecipitation assays (ChIP) provide cutting-edge protocols devoted to the most recent progress in ChIP and related subjects.
