scholarly journals Role of Transcription in Plasmid Maintenance in the hpr1Δ Mutant of Saccharomyces cerevisiae

2002 ◽  
Vol 22 (24) ◽  
pp. 8763-8773 ◽  
Author(s):  
Robert J. Merker ◽  
Hannah L. Klein
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Rna Polymerase Ii ◽  
Rna Splicing ◽  
Plasmid Stability ◽  
Transcription Elongation ◽  
Transcription Terminator ◽  
Plasmid Instability ◽  
Mature Mrna

ABSTRACT The Saccharomyces cerevisiae hyperrecombination mutation hpr1Δ results in instability of sequences between direct repeats that is dependent on transcription of the repeat. Here it is shown that the HPR1 gene also functions in plasmid stability in the presence of destabilizing transcription elongation. In the hpr1Δ mutant, plasmid instability results from unchecked transcription elongation, which can be suppressed by a strong transcription terminator. The plasmid system has been used to examine in vivo aspects of transcription in the absence of Hpr1p. Nuclear run-on studies suggest that there is an increased RNA polymerase II density in the hpr1Δ mutant strain, but this is not accompanied by an increase in accumulation of cytoplasmic mRNA. Suppression of plasmid instability in hpr1Δ can also be achieved by high-copy expression of the RNA splicing factor SUB2, which has recently been proposed to function in mRNA export, in addition to its role in pre-mRNA splicing. High-copy-number SUB2 expression is accompanied by an increase in message accumulation from the plasmid, suggesting that the mechanism of suppression by Sub2p involves the formation of mature mRNA. Models for the role of Hpr1p in mature mRNA formation and the cause of plasmid instability in the absence of the Hpr1 protein are discussed.

scholarly journals Requirement for Yeast RAD26, a Homolog of the HumanCSB Gene, in Elongation by RNA Polymerase II

2001 ◽  
Vol 21 (24) ◽  
pp. 8651-8656 ◽  
Author(s):  
Sung-Keun Lee ◽  
Sung-Lim Yu ◽  
Louise Prakash ◽  
Satya Prakash
Keyword(s):  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Transcription Elongation ◽  
Rapid Synthesis ◽  
Developmental Defects ◽  
Developmental Abnormalities ◽  
Developmental Problems ◽  
Uv Lesions

ABSTRACT Mutations in the human CSB gene cause Cockayne syndrome (CS). In addition to increased photosensitivity, CS patients suffer from severe developmental abnormalities, including growth retardation and mental retardation. Whereas a deficiency in the preferential repair of UV lesions from the transcribed strand accounts for the increased photosensitivity of CS patients, the reason for developmental defects in these individuals has remained unclear. Here we provide in vivo evidence for a role of RAD26, the counterpart of the CSB gene in Saccharomyces cerevisiae, in transcription elongation by RNA polymerase II, and in addition we show that under conditions requiring rapid synthesis of new mRNAs, growth is considerably reduced in cells lackingRAD26. These findings implicate a role for CSB in transcription elongation, and they strongly suggest that impaired transcription elongation is the underlying cause of the developmental problems in CS patients.

scholarly journals Evidence that the Elongation Factor TFIIS Plays a Role in Transcription Initiation at GAL1 in Saccharomyces cerevisiae

2005 ◽  
Vol 25 (7) ◽  
pp. 2650-2659 ◽  
Author(s):  
Donald M. Prather ◽  
Erica Larschan ◽  
Fred Winston
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Rna Polymerase Ii ◽  
Transcription Initiation ◽  
Transcription Elongation ◽  
Elongation Factor ◽  
Pol Ii ◽  
Upstream Activating Sequence ◽  
Transcript Cleavage

ABSTRACT TFIIS is a transcription elongation factor that has been extensively studied biochemically. Although the in vitro mechanisms by which TFIIS stimulates RNA transcript cleavage and polymerase read-through have been well characterized, its in vivo roles remain unclear. To better understand TFIIS function in vivo, we have examined its role during Gal4-mediated activation of the Saccharomyces cerevisiae GAL1 gene. Surprisingly, TFIIS is strongly associated with the GAL1 upstream activating sequence. In addition, TFIIS recruitment to Gal4-binding sites is dependent on Gal4, SAGA, and Mediator but not on RNA polymerase II (Pol II). The association of TFIIS is also necessary for the optimal recruitment of TATA-binding protein and Pol II to the GAL1 promoter. These results provide strong evidence that TFIIS plays an important role in the initiation of transcription at GAL1 in addition to its well-characterized roles in transcription elongation.

scholarly journals Divergent Subunit Interactions among Fungal mRNA 5′-Capping Machineries

2002 ◽  
Vol 1 (3) ◽  
pp. 448-457 ◽  
Author(s):  
Toshimitsu Takagi ◽  
Eun-Jung Cho ◽  
Rozmin T. K. Janoo ◽  
Vladimir Polodny ◽  
Yasutaka Takase ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Candida Albicans ◽  
Rna Polymerase Ii ◽  
Subunit Interactions ◽  
Pol Ii ◽  
Mrna Capping ◽  
Capping Enzyme ◽  
Enzyme Subunits ◽  
Carboxyl Terminal

ABSTRACT The Saccharomyces cerevisiae mRNA capping enzyme consists of two subunits: an RNA 5′-triphosphatase (RTPase) and GTP::mRNA guanylyltransferase (GTase). The GTase subunit (Ceg1) binds to the phosphorylated carboxyl-terminal domain of the largest subunit (CTD-P) of RNA polymerase II (pol II), coupling capping with transcription. Ceg1 bound to the CTD-P is inactive unless allosterically activated by interaction with the RTPase subunit (Cet1). For purposes of comparison, we characterize here the related GTases and RTPases from the yeasts Schizosaccharomyces pombe and Candida albicans. Surprisingly, the S. pombe capping enzyme subunits do not interact with each other. Both can independently interact with CTD-P of pol II, and the GTase is not repressed by CTD-P binding. The S. pombe RTPase gene (pct1 +) is essential for viability. Pct1 can replace the S. cerevisiae RTPase when GTase activity is supplied by the S. pombe or mouse enzymes but not by the S. cerevisiae GTase. The C. albicans capping enzyme subunits do interact with each other. However, this interaction is not essential in vivo. Our results reveal an unexpected diversity among the fungal capping machineries.

scholarly journals Loss of Ubp3 increases silencing, decreases unequal recombination in rDNA, and shortens the replicative life span in Saccharomyces cerevisiae

2014 ◽  
Vol 25 (12) ◽  
pp. 1916-1924 ◽  
Author(s):  
David Öling ◽  
Rehan Masoom ◽  
Kristian Kvint
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Rna Polymerase ◽  
Life Span ◽  
Rna Polymerase Ii ◽  
Ribosomal Dna ◽  
Genetic Evidence ◽  
Wild Type ◽  
Replicative Life Span ◽  
Unequal Recombination

Ubp3 is a conserved ubiquitin protease that acts as an antisilencing factor in MAT and telomeric regions. Here we show that ubp3∆ mutants also display increased silencing in ribosomal DNA (rDNA). Consistent with this, RNA polymerase II occupancy is lower in cells lacking Ubp3 than in wild-type cells in all heterochromatic regions. Moreover, in a ubp3∆ mutant, unequal recombination in rDNA is highly suppressed. We present genetic evidence that this effect on rDNA recombination, but not silencing, is entirely dependent on the silencing factor Sir2. Further, ubp3∆ sir2∆ mutants age prematurely at the same rate as sir2∆ mutants. Thus our data suggest that recombination negatively influences replicative life span more so than silencing. However, in ubp3∆ mutants, recombination is not a prerequisite for aging, since cells lacking Ubp3 have a shorter life span than isogenic wild-type cells. We discuss the data in view of different models on how silencing and unequal recombination affect replicative life span and the role of Ubp3 in these processes.

scholarly journals Impact of Pus1 Pseudouridine Synthase on Specific Decoding Events in Saccharomyces cerevisiae

Biomolecules ◽  
2020 ◽  
Vol 10 (5) ◽  
pp. 729
Author(s):  
Bahar Khonsari ◽  
Roland Klassen
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Elevated Temperature ◽  
Synthetic Lethality ◽  
Positive Role ◽  
Functional Impact ◽  
Pseudouridine Synthase ◽  
Decoding Efficiency ◽  
Cognate Trna

Pus1-dependent pseudouridylation occurs in many tRNAs and at multiple positions, yet the functional impact of this modification is incompletely understood. We analyzed the consequences of PUS1 deletion on the essential decoding of CAG (Gln) codons by tRNAGlnCUG in yeast. Synthetic lethality was observed upon combining the modification defect with destabilized variants of tRNAGlnCUG, pointing to a severe CAG-decoding defect of the hypomodified tRNA. In addition, we demonstrated that misreading of UAG stop codons by a tRNAGlnCUG variant is positively affected by Pus1. Genetic approaches further indicated that mildly elevated temperature decreases the decoding efficiency of CAG and UAG via destabilized tRNAGlnCAG variants. We also determined the misreading of CGC (Arg) codons by tRNAHisGUG, where the CGC decoder tRNAArgICG contains Pus1-dependent pseudouridine, but not the mistranslating tRNAHis. We found that the absence of Pus1 increased CGC misreading by tRNAHis, demonstrating a positive role of the modification in the competition against non-synonymous near-cognate tRNA. Part of the in vivo decoding defects and phenotypes in pus1 mutants and strains carrying destabilized tRNAGlnCAG were suppressible by additional deletion of the rapid tRNA decay (RTD)-relevant MET22, suggesting the involvement of RTD-mediated tRNA destabilization.

scholarly journals NC2α Interacts with BTAF1 and Stimulates Its ATP-Dependent Association with TATA-Binding Protein

2004 ◽  
Vol 24 (22) ◽  
pp. 10072-10082 ◽  
Author(s):  
Marcin P. Klejman ◽  
Lloyd A. Pereira ◽  
Hester J. T. van Zeeburg ◽  
Siv Gilfillan ◽  
Michael Meisterernst ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Rna Polymerase Ii ◽  
Transcriptional Activity ◽  
Binding Protein ◽  
Tata Binding Protein ◽  
Present Evidence ◽  
Cell Extracts ◽  
Human Ortholog ◽  
Rna Polymerase Ii Transcription

ABSTRACT Transcriptional activity of the TATA-binding protein (TBP) is controlled by a variety of proteins. The BTAF1 protein (formerly known as TAFII170/TAF-172 and the human ortholog of Saccharomyces cerevisiae Mot1p) and the NC2 complex composed of NC2α (DRAP1) and NC2β (Dr1) are able to bind to TBP directly and regulate RNA polymerase II transcription both positively and negatively. Here, we present evidence that the NC2α subunit interacts with BTAF1. In contrast, the NC2β subunit is not able to associate with BTAF1 and seems to interfere with the BTAF1-TBP interaction. Addition of NC2α or the NC2 complex can stimulate the ability of BTAF1 to interact with TBP. This function is dependent on the presence of ATP in cell extracts but does not involve the ATPase activity of BTAF1 nor phosphorylation of NC2α. Together, our results constitute the first evidence of the physical cooperation between BTAF1 and NC2α in TBP regulation and provide a framework to understand transcription functions of NC2α and NC2β in vivo.

scholarly journals Role of Integrase in Reverse Transcription of the Saccharomyces cerevisiae Retrotransposon Ty1

2005 ◽  
Vol 4 (6) ◽  
pp. 1057-1065 ◽  
Author(s):  
M. Wilhelm ◽  
F.-X. Wilhelm
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Amino Acid ◽  
Specific Activity ◽  
Rnase H ◽  
Amino Acid Residues ◽  
Acidic Amino Acid ◽  
Acidic Domain ◽  
Basic Region

ABSTRACT Reverse transcriptase (RT) with its associated RNase H (RH) domain and integrase (IN) are key enzymes encoded by retroviruses and retrotransposons. Several studies have implied a functional role of the interaction between IN and RT during the replication of retroviral and retrotransposon genomes. In this study, IN deletion mutants were used to investigate the role of IN on the RT activity of the yeast Saccharomyces cerevisiae retrotransposon Ty1. We have identified two domains of Ty1 integrase which have effects on RT activity in vivo. The deletion of a domain spanning amino acid residues 233 to 520 of IN increases the exogenous specific activity of RT up to 20-fold, whereas the removal of a region rich in acidic amino acid residues between residues 521 and 607 decreases its activity. The last result complements our observation that an active recombinant RT protein can be obtained if a small acidic tail mimicking the acidic domain of IN is fused to the RT-RH domain. We suggest that interaction between these acidic amino acid residues of IN and a basic region of RT could be critical for the correct folding of RT and for the formation of an active conformation of the enzyme.

scholarly journals Genome-Wide Relationships between TAF1 and Histone Acetyltransferases in Saccharomyces cerevisiae

2006 ◽  
Vol 26 (7) ◽  
pp. 2791-2802 ◽  
Author(s):  
Melissa Durant ◽  
B. Franklin Pugh
Keyword(s):  
Gene Expression ◽  
Saccharomyces Cerevisiae ◽  
Histone Acetylation ◽  
Rna Polymerase Ii ◽  
Histone Acetyltransferases ◽  
Histone H4 ◽  
Promoter Regions ◽  
Genome Wide ◽  
Acetylated Histone H4

ABSTRACT Histone acetylation regulates gene expression, yet the functional contributions of the numerous histone acetyltransferases (HATs) to gene expression and their relationships with each other remain largely unexplored. The central role of the putative HAT-containing TAF1 subunit of TFIID in gene expression raises the fundamental question as to what extent, if any, TAF1 contributes to acetylation in vivo and to what extent it is redundant with other HATs. Our findings herein do not support the basic tenet that TAF1 is a major HAT in Saccharomyces cerevisiae, nor do we find that TAF1 is functionally redundant with other HATs, including Gcn5, Elp3, Hat1, Hpa2, Sas3, and Esa1, which is in contrast to previous conclusions regarding Gcn5. Our findings do reveal that of these HATs, only Gcn5 and Esa1 contribute substantially to gene expression genome wide. Interestingly, histone acetylation at promoter regions throughout the genome does not require TAF1 or RNA polymerase II, indicating that most acetylation is likely to precede transcription and not depend upon it. TAF1 function has been linked to Bdf1, which binds TFIID and acetylated histone H4 tails, but no linkage between TAF1 and the H4 HAT Esa1 has been established. Here, we present evidence for such a linkage through Bdf1.

scholarly journals Interaction of the Srb10 Kinase with Sip4, a Transcriptional Activator of Gluconeogenic Genes in Saccharomyces cerevisiae

2001 ◽  
Vol 21 (17) ◽  
pp. 5790-5796 ◽  
Author(s):  
Olivier Vincent ◽  
Sergei Kuchin ◽  
Seung-Pyo Hong ◽  
Robert Townley ◽  
Valmik K. Vyas ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Rna Polymerase Ii ◽  
Transcriptional Repression ◽  
Transcriptional Control ◽  
Carbon Sources ◽  
Transcriptional Activator ◽  
Glucose Limitation ◽  
Cell Extracts ◽  
Rna Polymerase Ii Holoenzyme

ABSTRACT Sip4 is a Zn2Cys6 transcriptional activator that binds to the carbon source-responsive elements of gluconeogenic genes in Saccharomyces cerevisiae. The Snf1 protein kinase interacts with Sip4 and regulates its phosphorylation and activator function in response to glucose limitation; however, evidence suggested that another kinase also regulates Sip4. Here we examine the role of the Srb10 kinase, a component of the RNA polymerase II holoenzyme that has been primarily implicated in transcriptional repression but also positively regulates Gal4. We show that Srb10 is required for phosphorylation of Sip4 during growth in nonfermentable carbon sources and that the catalytic activity of Srb10 stimulates the ability of LexA-Sip4 to activate transcription of a reporter. Srb10 and Sip4 coimmunoprecipitate from cell extracts and interact in two-hybrid assays, suggesting that Srb10 regulates Sip4 directly. We also present evidence that the Srb10 and Snf1 kinases interact with different regions of Sip4. These findings support the view that the Srb10 kinase not only plays negative roles in transcriptional control but also has broad positive roles during growth in carbon sources other than glucose.

scholarly journals FCP1, a Phosphatase Specific for the Heptapeptide Repeat of the Largest Subunit of RNA Polymerase II, Stimulates Transcription Elongation

2002 ◽  
Vol 22 (21) ◽  
pp. 7543-7552 ◽  
Author(s):  
Subhrangsu S. Mandal ◽  
Helen Cho ◽  
Sungjoon Kim ◽  
Kettly Cabane ◽  
Danny Reinberg
Keyword(s):  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Transcription Elongation ◽  
Carboxy Terminal Domain ◽  
Transcription Factor Iif ◽  
Transcript Elongation ◽  
Rnap Ii ◽  
Carboxy Terminal

ABSTRACT FCP1, a phosphatase specific for the carboxy-terminal domain of RNA polymerase II (RNAP II), was found to stimulate transcript elongation by RNAP II in vitro and in vivo. This activity is independent of and distinct from the elongation-stimulatory activity associated with transcription factor IIF (TFIIF), and the elongation effects of TFIIF and FCP1 were found to be additive. Genetic experiments resulted in the isolation of several distinct fcp1 alleles. One of these alleles was found to suppress the slow-growth phenotype associated with either the reduction of intracellular nucleotide concentrations or the inhibition of other transcription elongation factors. Importantly, this allele of fcp1 was found to be lethal when combined individually with two mutations in the second-largest subunit of RNAP II, which had been shown previously to affect transcription elongation.

Sign in / Sign up

Export Citation Format

Share Document