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Publisher:
ISBN:
Category :
Languages : en
Pages : 276
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During RNA synthesis, RNA polymerase (RNAP) can temporally halt nucleotide addition by pausing. Transcriptional pausing plays a central role in regulation of transcription and may involve conformational changes in RNAP due to interactions of RNAP with intrinsic signals encoded in DNA and RNA, changes in translocation register, or both. However, basic mechanisms of transcriptional pausing and the role of RNAP translocation in regulation of transcript elongation are poorly understood. Here, we present work to investigate pause mechanisms, specifically how the pause signal is integrated/transmitted from the RNA exit channel of the enzyme to the enzymefs active site, and the contributions of RNAP translocation to pausing. We determined that the identity of RNA:DNA nucleotides in the active site are strong determinants of translocation bias, with the 3L RNA nucleotide favoring the pretranslocated state in the order U>C>A>G. Transcript elongation in bacteria is regulated in part by structures that form in the nascent RNA transcript and interact with RNAP in the RNA exit channel of the enzyme. A pause RNA hairpin structure makes direct contact with the flap domain of the RNAP f subunit, and slows nucleotide addition by factors of 10-20. We investigated the effects of different length and type of duplexes in the RNAP exit channel on pausing and response to regulators. We show that 8-bp RNA duplexes can stimulate pausing and those with less than 8-bp duplexes do not give full hairpin effect. We also investigated how regulatory communication between the formation of secondary structure in the RNA exit channel and the active site of RNAP modulate the catalytic center function. The opening of the RNAP clamp has been proposed to occur during pausing and stabilize the pause state. Our findings indicate that the formation of the paused 8-bp RNA:RNA duplex in the RNA exit channel causes movement of RNAPfs clamp and the flap domains, which in turn inhibits RNAP translocation and the catalysis of rapid nucleotide transfer. Transcription factors like NusA and RfaH tune and provide additional function to this communication network by affecting the clamp movements, translocation, or both functions.
Author:
Publisher:
ISBN:
Category :
Languages : en
Pages : 276
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Book Description
During RNA synthesis, RNA polymerase (RNAP) can temporally halt nucleotide addition by pausing. Transcriptional pausing plays a central role in regulation of transcription and may involve conformational changes in RNAP due to interactions of RNAP with intrinsic signals encoded in DNA and RNA, changes in translocation register, or both. However, basic mechanisms of transcriptional pausing and the role of RNAP translocation in regulation of transcript elongation are poorly understood. Here, we present work to investigate pause mechanisms, specifically how the pause signal is integrated/transmitted from the RNA exit channel of the enzyme to the enzymefs active site, and the contributions of RNAP translocation to pausing. We determined that the identity of RNA:DNA nucleotides in the active site are strong determinants of translocation bias, with the 3L RNA nucleotide favoring the pretranslocated state in the order U>C>A>G. Transcript elongation in bacteria is regulated in part by structures that form in the nascent RNA transcript and interact with RNAP in the RNA exit channel of the enzyme. A pause RNA hairpin structure makes direct contact with the flap domain of the RNAP f subunit, and slows nucleotide addition by factors of 10-20. We investigated the effects of different length and type of duplexes in the RNAP exit channel on pausing and response to regulators. We show that 8-bp RNA duplexes can stimulate pausing and those with less than 8-bp duplexes do not give full hairpin effect. We also investigated how regulatory communication between the formation of secondary structure in the RNA exit channel and the active site of RNAP modulate the catalytic center function. The opening of the RNAP clamp has been proposed to occur during pausing and stabilize the pause state. Our findings indicate that the formation of the paused 8-bp RNA:RNA duplex in the RNA exit channel causes movement of RNAPfs clamp and the flap domains, which in turn inhibits RNAP translocation and the catalysis of rapid nucleotide transfer. Transcription factors like NusA and RfaH tune and provide additional function to this communication network by affecting the clamp movements, translocation, or both functions.
Author: Tasuku Honjo
Publisher: Academic Press
ISBN: 9780123979339
Category : Science
Languages : en
Pages : 0
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Book Description
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.
Author: Michael Justin Blau
Publisher:
ISBN:
Category :
Languages : en
Pages :
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Author: Chang-Hui Shen
Publisher: Elsevier
ISBN: 0323986099
Category : Science
Languages : en
Pages : 590
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Book Description
Diagnostic Molecular Biology, Second Edition describes the fundamentals of molecular biology in a clear, concise manner with each technique explained within its conceptual framework and current applications of clinical laboratory techniques comprehensively covered. This targeted approach covers the principles of molecular biology, including basic knowledge of nucleic acids, proteins and chromosomes; the basic techniques and instrumentations commonly used in the field of molecular biology, including detailed procedures and explanations; and the applications of the principles and techniques currently employed in the clinical laboratory. Topics such as whole exome sequencing, whole genome sequencing, RNA-seq, and ChIP-seq round out the discussion. Fully updated, this new edition adds recent advances in the detection of respiratory virus infections in humans, like influenza, RSV, hAdV, hRV but also corona. This book expands the discussion on NGS application and its role in future precision medicine. - Provides explanations on how techniques are used to diagnosis at the molecular level - Explains how to use information technology to communicate and assess results in the lab - Enhances our understanding of fundamental molecular biology and places techniques in context - Places protocols into context with practical applications - Includes extra chapters on respiratory viruses (Corona)
Author: Michael F. Carey
Publisher: CSHL Press
ISBN: 9780879696351
Category : Medical
Languages : en
Pages : 684
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Book Description
In the genome era, the analysis of gene expression has become a critical requirement in many laboratories. But there has been no comprehensive source of strategic, conceptual, and technical information to guide this often complex task. Transcriptional Regulation in Eukaryotes answers that need. Written by two experienced investigators, Michael Carey and Stephen Smale at the UCLA School of Medicine, and based in part on the Gene Expression course taught at Cold Spring Harbor Laboratory, this book directly addresses all the concerns of a laboratory studying the regulation of a newly isolated gene and the biochemistry of a new transcription factor. This important and unique book is essential reading for anyone pursuing the analysis of gene expression in model systems or disease states.
Author: Ron Milo
Publisher: Garland Science
ISBN: 1317230698
Category : Science
Languages : en
Pages : 400
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Book Description
A Top 25 CHOICE 2016 Title, and recipient of the CHOICE Outstanding Academic Title (OAT) Award. How much energy is released in ATP hydrolysis? How many mRNAs are in a cell? How genetically similar are two random people? What is faster, transcription or translation?Cell Biology by the Numbers explores these questions and dozens of others provid
Author: Torben Heick Jensen
Publisher: Springer Science & Business Media
ISBN: 1441978410
Category : Medical
Languages : en
Pages : 161
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Book Description
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.
Author: Manchuta Dangkulwanich
Publisher:
ISBN:
Category :
Languages : en
Pages : 137
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Book Description
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.
Author:
Publisher: Academic Press
ISBN: 0128145161
Category : Science
Languages : en
Pages : 4109
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Book Description
Encyclopedia of Virology, Fourth Edition, Five Volume Set builds on the solid foundation laid by the previous editions, expanding its reach with new and timely topics. In five volumes, the work provides comprehensive coverage of the whole virosphere, making this a unique resource. Content explores viruses present in the environment and the pathogenic viruses of humans, animals, plants and microorganisms. Key areas and concepts concerning virus classification, structure, epidemiology, pathogenesis, diagnosis, treatment and prevention are discussed, guiding the reader through chapters that are presented at an accessible level, and include further readings for those needing more specific information. More than ever now, with the Covid19 pandemic, we are seeing the huge impact viruses have on our life and society. This encyclopedia is a must-have resource for scientists and practitioners, and a great source of information for the wider public. Offers students and researchers a one-stop shop for information on virology not easily available elsewhere Fills a critical gap of information in a field that has seen significant progress in recent years Authored and edited by recognized experts in the field, with a range of different expertise, thus ensuring a high-quality standard
Author: Samantha Fowler
Publisher:
ISBN: 9781739015503
Category :
Languages : en
Pages : 0
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Book Description
Black & white print. Concepts of Biology is designed for the typical introductory biology course for nonmajors, covering standard scope and sequence requirements. The text includes interesting applications and conveys the major themes of biology, with content that is meaningful and easy to understand. The book is designed to demonstrate biology concepts and to promote scientific literacy.
