An Analysis of Mer1 Function During Meiotic Splicing Regulation in Saccharomyces Cerevisiae PDF Download
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Author: Frederick W. Scherrer (Jr.)
Publisher:
ISBN:
Category : Cell division
Languages : en
Pages : 107
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Book Description
The transition from mitosis to meiosis in the yeast Saccharomyces cerevisiae requires a significant change to gene expression profiles. Regulation of pre-messenger RNA splicing patterns during meiosis assists in this transition by fine tuning expression of essential meiotic genes. Produced only during meiosis, Mer1p is linked to the splicing of at least three mRNAs: MER2, MER3, and AMA1. Previous evidence suggests that Mer1p activates splicing by directly recruiting snRNPs or stabilizing intermediate splicing complexes formed on pre-mRNA that contains an intronic Mer1p enhancer element. However, some splicing factors, especially accessory/non-snRNP factors, have critical roles in retaining unspliced pre-mRNAs in the nucleus. I tested if Mer1p may indirectly regulate splicing by preventing the export of pre-mRNAs to the cytoplasm and also demonstrated that a second subunit of the Retention and Splicing (RES) complex, Bud13p, has transcript-specific effects on Mer1p-activated splicing. The results indicated that Mer1p can retain unspliced pre-mRNA in the nucleus; however, nuclear retention could not be uncoupled from splicing activation. In the absence of Mer1p, the AMA1 pre-mRNA is exported to the cytoplasm, translated, but not subjected to nonsense-mediated decay (NMD) despite a premature stop codon in the intron. A novel role for the Mer1p activation domain was revealed by a two-hybrid interaction with Prp39p, an essential U1 snRNP protein. This suggests the initial contact between Mer1p and the spliceosome occurs during commitment complex assembly. Collectively, these data imply that Mer1p can retain pre-mRNAs in the nucleus only by facilitating their interaction with the spliceosome and support models for cytoplasmic degradation of unspliced pre-mRNAs that fail to assemble into spliceosomes in yeast. A two-hybrid analysis of U1 snRNP proteins and other early splicing factors tested 460 possible interactions and the several novel interactions reported here indicate a revised model for U1snRNP structure.
Author: Frederick W. Scherrer (Jr.)
Publisher:
ISBN:
Category : Cell division
Languages : en
Pages : 107
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Book Description
The transition from mitosis to meiosis in the yeast Saccharomyces cerevisiae requires a significant change to gene expression profiles. Regulation of pre-messenger RNA splicing patterns during meiosis assists in this transition by fine tuning expression of essential meiotic genes. Produced only during meiosis, Mer1p is linked to the splicing of at least three mRNAs: MER2, MER3, and AMA1. Previous evidence suggests that Mer1p activates splicing by directly recruiting snRNPs or stabilizing intermediate splicing complexes formed on pre-mRNA that contains an intronic Mer1p enhancer element. However, some splicing factors, especially accessory/non-snRNP factors, have critical roles in retaining unspliced pre-mRNAs in the nucleus. I tested if Mer1p may indirectly regulate splicing by preventing the export of pre-mRNAs to the cytoplasm and also demonstrated that a second subunit of the Retention and Splicing (RES) complex, Bud13p, has transcript-specific effects on Mer1p-activated splicing. The results indicated that Mer1p can retain unspliced pre-mRNA in the nucleus; however, nuclear retention could not be uncoupled from splicing activation. In the absence of Mer1p, the AMA1 pre-mRNA is exported to the cytoplasm, translated, but not subjected to nonsense-mediated decay (NMD) despite a premature stop codon in the intron. A novel role for the Mer1p activation domain was revealed by a two-hybrid interaction with Prp39p, an essential U1 snRNP protein. This suggests the initial contact between Mer1p and the spliceosome occurs during commitment complex assembly. Collectively, these data imply that Mer1p can retain pre-mRNAs in the nucleus only by facilitating their interaction with the spliceosome and support models for cytoplasmic degradation of unspliced pre-mRNAs that fail to assemble into spliceosomes in yeast. A two-hybrid analysis of U1 snRNP proteins and other early splicing factors tested 460 possible interactions and the several novel interactions reported here indicate a revised model for U1snRNP structure.
Author: Katherine Anne Senn
Publisher:
ISBN:
Category :
Languages : en
Pages : 0
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Book Description
Pre-mRNA splicing, the removal of noncoding introns to form a protein coding mRNA, is a crucial step in eukaryotic gene expression. The spliceosome, a macromolecular complex made up of five small nuclear RNAs (snRNAs) and dozens of proteins organized into small nuclear ribonucleoproteins (snRNPs), catalyzes splicing in two steps. These steps are highly regulated to ensure correct mRNA isoform production and protein expression. Core components of the spliceosome as well as additional auxiliary factors tune splicing for accuracy and regulation of alternative splicing outcomes. Because of the complexity of splicing regulation, there are many open questions about the factors involved and the regulation of alternative splicing. This thesis addresses the functions of two yeast proteins involved in splicing regulation and an alternatively spliced gene in humans.In chapters 2 and 3, I discuss the novel yeast splicing factor Fyv6. Genetic and biochemical evidence supports a role for Fyv6 in the second step of splicing, exon ligation. A collaborator has determined the structure of a Fyv6-containing spliceosome by cryo-electron microscopy. Using this structure, we have used biochemical and transcriptomics experiments to elucidate how Fyv6 affects 3' splice site choice. Yeast splicing regulatory factor Mer1 interacts with introns and the U1 snRNP to enhance splicing of a subset of meiotically-regulated pre-mRNAs by an as-yet undetermined mechanism. With the goal of elucidating the mechanism, I recapitulated Mer1-dependent splicing in vitro and used colocalization single molecule microscopy to examine how Mer1 affects the dynamics of U1 binding to pre-mRNAs (Chapter 4). Our results indicate Mer1 promotes splicing without significantly altering the observed distributions of U1 binding events. This study lays the groundwork for future studies of splicing regulation by Mer1 and other factors. Finally, I studied the expression and alternative splicing of human glucose transporter family member GLUT8 (Chapter 5). Multiple alternatively spliced GLUT8 mRNA isoforms exist, but only some encode viable proteins. One that does not is upregulated in cancers, which reduces GLUT8 protein expression. Collaborators showed that based on its localization, inability to transport glucose, and release of a cleaved peptide, the main functional isoform of GLUT8 may be a metabolic sensor.
Author: Javier Armisen Garrido
Publisher:
ISBN:
Category : Biochemistry
Languages : en
Pages : 155
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Book Description
In eukaryotes, genes are presented in a series of coding and non-coding DNA regions (exons/introns) that are transcribed into a premature RNA (pre-mRNA). Introns can be removed from the mature premRNA, before its translation into proteins, in a process called splicing. The splicing reaction occurs in two highly regulated transesterification reactions inside of the cell nucleus, and it is catalyzed by the Spliceosome, involving the binding and release of five small nuclear ribonucleoprotein particles (snRNPs). While some introns are constitutively spliced, others can be alternatively spliced, giving different exon combinations and therefore different proteins, increasing the protein diversity of the species. In humans, misregulation of alternative splicing can result in the production of aberrant proteins, some of which may produce cancer or other severe diseases. In yeast, alternative splicing is regulated by different splicing factors, such as Mer1p. Mer1p is expressed during meiosis in the yeast Saccharomyces cerevisiae and activates the splicing in at least three different genes (AMA1, MER2, and MER3), which contain a conserved intronic splicing enhancer sequence. Previous results have shown that Mer1p is able to interact with the pre-mRNA and with specific proteins of the U1 and U2 snRNPs. However, the specific molecular mechanisms by which Mer1p activates splicing remained unknown. The objective of this work is to determine how Mer1p regulates the splicing of its targets, and how different splicing factors modulate Mer1p activity. Using biochemistry and genetics, the data presented in this work indicate that Mer1p recruits the snRNPs U1, U2 and U6, to pre-mRNA. This recruitment of the snRNPs is dependent of the U1 snRNP protein Nam8p and the U2 snRNP protein Snu17p, but independent on the branchpoint region or ATP. Furthermore, Mer1p accelerates and stabilizes the formation of the early complexes of the spliceosome. Finally, U1 and U2 are recruited to the pre-mRNA at the same time, emerging a new alternative hypothesis of splicing regulation that can be applied to other enhancer regulators and that differs from the classical model of stepwise assembly of the snRNP.
Author: Zhicheng Qiu
Publisher:
ISBN:
Category :
Languages : en
Pages : 318
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Book Description
Meiotic splicing is a form of regulated splicing that promotes intron removal from pre-mRNAs during meiosis. In addition to transcription regulation, meiotic splicing provides an extra layer of regulation that allows expression of a subset of meiotic genes essential for the meiotic developmental program in Saccharomyces cerevisiae . Several factors play important roles in meiotic splicing. Mer1 is a protein produced only in meiotic cells and activates splicing by binding to an intronic Mer1-enhancer element present in the AMA1 , MER2 and MER3 pre-mRNAs. Nam8, a component of yeast U1 snRNP, is optional for mitotic growth but required during meiosis, because Nam8 collaborates with Mer1 to promote splicing of essential meiotic mRNAs including AMA1 , MER2 and MER3 . Here I identify SPO22 and PCH2 as novel targets of Nam8-dependent meiotic splicing. Whereas SPO22 splicing is co-dependent on Mer1, PCH2 is not. In addition, I find that Tgs1, the enzyme that converts m 7 G RNA caps to the 2,2,7-trimethylguanosine (TMG) caps characteristic of spliceosomal snRNAs, is another essential factor for meiotic splicing and meiosis. tgs1 ? cells are especially defective in splicing PCH2 and SAE3 meiotic pre-mRNAs. I also show that four essential meiotic pre-mRNAs: MER2 , MER3 , SPO22 and AMA1 are sufficient to rescue the sporulation defects of nam8 ? and mer1 ?. Whereas Nam8 is also necessary for PCH2 splicing, PCH2 cDNA is not needed for sporulation by nam 8?, diploids. These results suggest that there are no essential intron-containing RNAs missing from the known roster of Mer1 and Nam8 targets.
Author: Mei Li Benni
Publisher:
ISBN:
Category :
Languages : en
Pages : 292
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Author: Louise Joanna Newnham
Publisher:
ISBN:
Category :
Languages : en
Pages :
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The Synaptonemal Complex (SC) is a proteinaceous structure that connects homologous chromosomes lengthwise during meiotic prophase. In budding yeast, the SC consists of two parallel axes that become connected by the central element protein, Zip1 that extends along the chromosome axes (Sym, Engebrecht et al. 1993). Extension of the SC is coordinated to crossover formation by a group of proteins known as the 'ZMM's (Zip1, Zip2, Zip3, Zip4, Msh4, Msh5 and Mer3) (Borner, Kleckner et al. 2004). Work outlined here demonstrates a role for the mismatch repair paralogue, Msh4 in preventing SC extension from being de-coupled from crossover formation. Furthermore, increased temperature serves as a positive effector for this decoupling. These findings suggest that SC extension is highly regulated to ensure that it is coupled with crossing over. As well as its role in crossover formation (Storlazzi, Xu et al. 1996), the work outlined here demonstrates an independent role for Zip1 in promoting the segregation of non-exchange chromosome pairs (NECs). Zip1 pairs the centromeres of NECs in pachytene through to metaphase I, where it aids their segregation at the first meiotic division. The localisation and function of Zip1 at the centromeres of non-exchange chromosomes depends on Zip3 and Zip2, respectively. Zip1 is observed at the centromeres of all chromosomes following SC disassembly through to the first meiotic division, where it promotes the segregation of exchange pairs also. A model is suggested whereby Zip1 promotes the segregation of both non-exchange and exchange chromosome pairs by tethering homologous centromeres throughout meiotic prophase. Finally, a parallel pathway for NEC segregation is also described that depends upon the spindle checkpoint component, Mad3. When both ZIP1 and MAD3 are deleted, NECs segregate at random.
Author:
Publisher:
ISBN:
Category : Dissertations, Academic
Languages : en
Pages : 850
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Author:
Publisher: Academic Press
ISBN: 012375173X
Category : Science
Languages : en
Pages : 961
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Book Description
This fully updated edition of the bestselling three-part Methods in Enzymology series, Guide to Yeast Genetics and Molecular Cell Biology is specifically designed to meet the needs of graduate students, postdoctoral students, and researchers by providing all the up-to-date methods necessary to study genes in yeast. Procedures are included that enable newcomers to set up a yeast laboratory and to master basic manipulations. This volume serves as an essential reference for any beginning or experienced researcher in the field. - Provides up-to-date methods necessary to study genes in yeast - Includes proceedures that enable newcomers to set up a yeast laboratory and to master basic manipulations - Serves as an essential reference for any beginning or experienced researcher in the field
Author: Lisa Reneé Treviño
Publisher:
ISBN:
Category :
Languages : en
Pages : 206
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Book Description
Author:
Publisher: ScholarlyEditions
ISBN: 1464963797
Category : Medical
Languages : en
Pages : 2673
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Book Description
Issues in Genetic Medicine / 2011 Edition is a ScholarlyEditions™ eBook that delivers timely, authoritative, and comprehensive information about Genetic Medicine. The editors have built Issues in Genetic Medicine: 2011 Edition on the vast information databases of ScholarlyNews.™ You can expect the information about Genetic Medicine in this eBook to be deeper than what you can access anywhere else, as well as consistently reliable, authoritative, informed, and relevant. The content of Issues in Genetic Medicine: 2011 Edition has been produced by the world’s leading scientists, engineers, analysts, research institutions, and companies. All of the content is from peer-reviewed sources, and all of it is written, assembled, and edited by the editors at ScholarlyEditions™ and available exclusively from us. You now have a source you can cite with authority, confidence, and credibility. More information is available at http://www.ScholarlyEditions.com/.
