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Author: Boris Zinshteyn
Publisher:
ISBN:
Category :
Languages : en
Pages : 159
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Book Description
The goal of this thesis is to elucidate the mechanisms that govern translational efficiency (TE) - the amount of protein produced from each molecule of mRNA. While the mechanisms regulating the TE of a few specific messages are well understood, the general contribution of translational control to differences in cellular protein levels is currently unclear. Recent advances have enabled the direct measurement of protein levels and translation rates genome-wide, and studies in multiple organisms have found varying degrees of translation regulation, both at steady state, and in response to stress or developmental cues. Despite this influx of high-throughput data, the mechanisms underlying the differences in gene-specific and condition-dependent TE remain largely unknown. In this thesis, I describe the roles of two different components of the translational machinery in regulating translational efficiency. In Chapter 1, I discuss the features of mRNA coding sequences that can affect TE, thereby introducing Chapter 2, in which I investigate the role of a conserved anticodon tRNA modification in determining the rate of translation elongation and the phenotypic consequences of its loss for budding yeast. In Chapter 3, I discuss the regulation of translation initiation to introduce Chapter 4, in which I explore how the RNA binding specificity of the core translation factor, yeast eukaryotic initiation factor 4G (eIF4G), contributes to genome-wide competition between mRNAs. Finally, I will discuss future directions for this work.
Author: Boris Zinshteyn
Publisher:
ISBN:
Category :
Languages : en
Pages : 159
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Book Description
The goal of this thesis is to elucidate the mechanisms that govern translational efficiency (TE) - the amount of protein produced from each molecule of mRNA. While the mechanisms regulating the TE of a few specific messages are well understood, the general contribution of translational control to differences in cellular protein levels is currently unclear. Recent advances have enabled the direct measurement of protein levels and translation rates genome-wide, and studies in multiple organisms have found varying degrees of translation regulation, both at steady state, and in response to stress or developmental cues. Despite this influx of high-throughput data, the mechanisms underlying the differences in gene-specific and condition-dependent TE remain largely unknown. In this thesis, I describe the roles of two different components of the translational machinery in regulating translational efficiency. In Chapter 1, I discuss the features of mRNA coding sequences that can affect TE, thereby introducing Chapter 2, in which I investigate the role of a conserved anticodon tRNA modification in determining the rate of translation elongation and the phenotypic consequences of its loss for budding yeast. In Chapter 3, I discuss the regulation of translation initiation to introduce Chapter 4, in which I explore how the RNA binding specificity of the core translation factor, yeast eukaryotic initiation factor 4G (eIF4G), contributes to genome-wide competition between mRNAs. Finally, I will discuss future directions for this work.
Author:
Publisher:
ISBN:
Category :
Languages : en
Pages : 9
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Book Description
Killer virus of Saccharomyces cerevisiae is a cytoplasmically- inherited virus that confers on persistently-infected yeast cells the ability to secrete a protein toxin which kills uninfected yeast cells but to which infected cells, denoted killers, are resistant (reviewed by Tipper and Bostian, 1984; Wickner, 1986). Killer toxin production can be assayed by halos of killing of a lawn of uninfected cells around killer colonies on a petri dish. The genome of the virus consists of two segments of double-stranded (ds) RNA that are separately encapsidated in cytoplasmic virions localized in the cytoplasm of infected cells. The L-A dsRNA segment is 4.9 kbp in length and encodes the major capsid protein (81 kd) of the virions and possibly a larger capsid-RNA polymerase fusion product by a -1 frame shift occurring at the translational level, analogous to that producing retrovirus gag-pol fusion protein (Icho and Wickner, 1988; Diamond et al, 1989).
Author: Caitlin E. Gamble
Publisher:
ISBN:
Category :
Languages : en
Pages : 124
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Book Description
Synonymous codon choice modulates translation, but the properties of codons or codon combinations that result in impaired translation are not understood. We scored expression of 35,811 three-codon insertions in GFP in Saccharomyces cerevisiae and evaluated these variants for codon usage and RNA structure effects on GFP fluorescence levels. We have established that codon pairs affect translation elongation and efficiency in yeast in a manner distinct from the effects of individual codon tRNA abundance. Also, similar to previous studies in bacteria, we have found that the base-pairing status of nucleotides near the translation start site is likely to impair translation initiation. Both inhibitory codon pairs and 5’ mRNA structure can impose substantial limitations on translation efficiency through synonymous variation. For 17 inhibitory codon pairs, we show that it is the pair, rather than the dipeptide, the 6-base sequence, or the two individual codons, that is responsible for inhibition. Variants from the GFP insertion library that had an inhibitory pair had significantly lower expression than variants in which: the 6 base sequences were out of frame; the two codons were present but separated; or one of the codons of the pair was instead an optimal codon. We find that the inhibitory pairs act in translation, based on both suppression of inhibition by over expressed tRNA (11/12 tested) and the reduction in translation speeds relative to synonymous dipeptide sequences, as observed from ribosome occupancies along yeast transcripts. Furthermore, for 12 of the 17 pairs, preserving the order of codons in the pair was required for strong inhibitory effects. Thus the position of inhibitory pairs within the ribosome is likely a key factor in translation efficiency. Moreover, the identity of codons in inhibitory pairs is inconsistent with an inhibition mechanism governed primarily by limited tRNA supply. Rather, our data implicates wobble decoding and interactions between adjacent sites in the ribosome. The high-throughput experimental analysis described here has resulted in the direct and extensive identification of multiple inhibitory codon pairs, a quantitative analysis of their relative effects on translation in vivo, and tests of their activity as modulators of translation.
Author: Claude Kipnis
Publisher: Springer Science & Business Media
ISBN: 3662037521
Category : Mathematics
Languages : en
Pages : 453
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Book Description
This book has been long awaited in the "interacting particle systems" community. Begun by Claude Kipnis before his untimely death, it was completed by Claudio Landim, his most brilliant student and collaborator. It presents the techniques used in the proof of the hydrodynamic behavior of interacting particle systems.
Author: Dustin Howard Hite
Publisher:
ISBN:
Category :
Languages : en
Pages :
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The central dogma of biology states that DNA, the genetic information, is transcribed into RNA, an information containing intermediate, which is then translated into proteins, actionable molecules which perform the majority of tasks required for life. To synthesize proteins, the cell employs a massive, macromolecular machine, the ribosome, and a myriad of protein factors to successfully translate an mRNA. My graduate studies have focused both on the ribosome and the protein translation factors that interact with the ribosome to facilitate translation initiation, elongation, and termination. First, utilizing recent advances in high throughput sequencing, we discovered that sequencing of ribosome protected fragments could illuminate in vivo dynamics of ribosome structural changes in Saccharomyces cerevisiae. We demonstrated that the ribosome protects two distinct sizes of fragments and assigned each fragment population to approximate stages of the translation elongation cycle where large structural rearrangements of the ribosome are known to occur. Once these assignments were made, we were able to model elongation speed and demonstrated that, contrary to previous reports, tRNA abundance and codon optimality were not the major determinants of elongation speed; surprisingly our data indicated that the polarity of the amino acid being decoded dictated elongation rates under these conditions, with polar amino acids acting to slow elongation rates. This study also implicated Dom34, a known NO GO decay factor, as a novel component of canonical translation termination and ribosome recycling. Second, we used another genome-wide assay of translation, "gradient encoding" microarray analysis, to interrogate the genome-wide effects of depleting five individual translation factors. Based on the current understanding of the molecular mechanisms of each translation factor, we hypothesized that the depletion of each factor would result in differential translation of mRNAs based on the physical properties of each mRNA species. However, we were startled to observe that the translational program of S. cerevisiae was relatively unperturbed by the depletion of three initiation factors, one elongation factor, and one termination factor. Further investigation revealed that yeast were actively compensating for the deficiency of each factor by either increasing or decreasing translation initiation rates such that the depleted factor was no longer limiting. This tuning was mediated by changes in eIF2[alpha] phosphorylation levels, a known modulator of translation initiation. Overall, we have leveraged high throughput technologies to provide novel understanding of in vivo structural dynamics of the ribosome and reveal a novel, unexpected robustness of the translational program in S. cerevisiae.
Author: Chia-Lin Wei (Ph.D. in microbiology)
Publisher:
ISBN:
Category :
Languages : en
Pages : 422
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Author: Mick F. Tuite
Publisher: Springer
ISBN:
Category : Science
Languages : en
Pages : 552
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Author: Wanfu Hou
Publisher:
ISBN:
Category :
Languages : en
Pages : 0
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While we have learned that mRNA sequence strongly influences translation and co-translational pathways, the exact mechanisms by which this sequence and the RNA structures encoded impact these steps of gene expression are less understood. Therefore, more investigations are needed to reveal how translation elongation and efficiency is influenced by RNA sequence and how the affected translation regulates and determines a variety of co-translational pathways. In this dissertation, I explore the effects of translational kinetics on a series of co-translational pathways using the budding yeast Saccharomyces cerevisiae as a model organism. In Chapter 2, I developed an in-vivo elongation reporter in Saccharomyces cerevisiae to quantitatively monitor translation elongation duration and protein expression. Using this elongation reporter, I investigated the effects of elongation stalls induced by different types of genetic factors on gene expression and demonstrated that distinct ribosomal stalls may trigger distinct co-translational pathways. In Chapter 3, I studied co-translational mRNA localization to mitochondrion, and proposed that translational kinetics, such as ribosomal stall caused by polyprolines, play an important role in mediating co-translational import. In addition, I further studied the effects of elongation stalls on mitochondrial import stress and triggering of relevant quality control pathways. In Chapter 4, I investigated the mechanism of cytosolic mRNP granule formation under glucose deprivation condition. In this study, I use CRISPRi to knockdown expression of RVB2 and confirm its essential role in deciding the fate of mRNA localization and translatability after glucose depletion. Finally in Chapter 5, I address the enhancements made to the developed method, outline potential future directions for the research presented in this dissertation, and conclude with my final remarks.
Author: Nahum Sonenberg
Publisher: CSHL Press
ISBN: 9780879696184
Category : Gene expression
Languages : en
Pages : 1034
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Book Description
Since the 1996 publication of Translational Control, there has been fresh interest in protein synthesis and recognition of the key role of translation control mechanisms in regulating gene expression. This new monograph updates and expands the scope of the earlier book but it also takes a fresh look at the field. In a new format, the first eight chapters provide broad overviews, while each of the additional twenty-eight has a focus on a research topic of more specific interest. The result is a thoroughly up-to-date account of initiation, elongation, and termination of translation, control mechanisms in development in response to extracellular stimuli, and the effects on the translation machinery of virus infection and disease. This book is essential reading for students entering the field and an invaluable resource for investigators of gene expression and its control.
Author: David C. Amberg
Publisher: CSHL Press
ISBN: 0879697288
Category : Genetics
Languages : en
Pages : 250
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Book Description
"Methods in Yeast Genetics" is a course that has been offered annually at Cold Spring Harbor for the last 30 years. This provides a set of teaching experiments along with the protocols and recipes for the standard techniques and reagents used in the study of yeast biology.
