TFIIA Plays a Role in the Response to Oxidative Stress

ABSTRACT To characterize the role of the general transcription factor TFIIA in the regulation of gene expression by RNA polymerase II, we examined the transcriptional profiles of TFIIA mutants of Saccharomyces cerevisiae using DNA microarrays. Whole-genome expression profiles were determined for three different mutants with mutations in the gene coding for the small subunit of TFIIA, TOA2. Depending on the particular mutant strain, approximately 11 to 27% of the expressed genes exhibit altered message levels. A search for common motifs in the upstream regions of the pool of genes decreased in all three mutants yielded the binding site for Yap1, the transcription factor that regulates the response to oxidative stress. Consistent with a TFIIA-Yap1 connection, the TFIIA mutants are unable to grow under conditions that require the oxidative stress response. Underexpression of Yap1-regulated genes in the TFIIA mutant strains is not the result of decreased expression of Yap1 protein, since immunoblot analysis indicates similar amounts of Yap1 in the wild-type and mutant strains. In addition, intracellular localization studies indicate that both the wild-type and mutant strains localize Yap1 indistinguishably in response to oxidative stress. As such, the decrease in transcription of Yap1-dependent genes in the TFIIA mutant strains appears to reflect a compromised interaction between Yap1 and TFIIA. This hypothesis is supported by the observations that Yap1 and TFIIA interact both in vivo and in vitro. Taken together, these studies demonstrate a dependence of Yap1 on TFIIA function and highlight a new role for TFIIA in the cellular mechanism of defense against reactive oxygen species.

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Comparative phytochemical analysis of Phlogacanthus thyrsiflorus Nees: Implications on attenuation of pro-oxidants and pathogen virulence in Caenorhabditis elegans model system

Asian Journal of Pharmaceutical and Clinical Research ◽

10.22159/ajpcr.2017.v10i5.17319 ◽

2017 ◽

Vol 10 (5) ◽

pp. 361

Author(s):

Kitlangki Suchiang ◽

Nitasha H Kayde

Keyword(s):

Oxidative Stress ◽

Free Radicals ◽

Free Radical ◽

Caenorhabditis Elegans ◽

Model System ◽

Wild Type ◽

C Elegans ◽

Mutant Strains

Background: Phlogacanthus thyrsiflorus Nees (P. thyrsiflorus) of Acanthaceae family is endogenous to sub-tropical Himalayas. It has been reported to be used traditionally in Jaintia tribe of Meghalaya, India for treatment of many ailments.Objectives: The aim was to detect the active compounds present in the leaves for evaluation of in vitro free radicals scavenging potentials. Leaves protective actions in vivo will be investigated using Caenorhabditis elegans (C. elegans) model system utilizing wild type and mutant strains and the phenomena of host-pathogens interactions.Materials and methods: Gas chromatography/ Mass spectrometry (GC/MS) was used for detection of different compounds present. The versatility of leaf extracts to scavenge different free radicals generated in vitro was assessed with different in vitro methods. Survival analysis of wild type and mutant strains C. elegans under enhanced pro-oxidants exposure was investigated in vivo. Fast killing assay was also performed to study the extracts modulatory activity on host C. elegans survival under pathogen Pseudomonas aeruginosa infection.Results: Forty compounds were detected in methanolic fraction of the extract with variable percentages. Both aqueous and methanol extract possessed remarkable, versatile free radical scavenging activity irrespective of the types of free radical generated. The in vivo experiments are in compliance, with observable increased survival ability percentage of C. elegans under intense exogenous oxidative stress and pathogen infection.Conclusion: Our findings enlightened the different compounds present with versatility of P. thyrsiflorus in tackling different free radicals generated both in vitro and in vivo that highly support for its candidature as a good antioxidant source. Our findings may justify the historical relevance of this plant in herbal remedies that could form the basis for inquiry of new active principles.Keywords: Free radicals, Oxidative stress, Caenorhabditis elegans, Phlogacanthus thyrsiflorus, Phytochemicals

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Evidence for the Involvement of the Glc7-Reg1 Phosphatase and the Snf1-Snf4 Kinase in the Regulation of INO1 Transcription in Saccharomyces cerevisiae

Genetics ◽

10.1093/genetics/152.1.73 ◽

1999 ◽

Vol 152 (1) ◽

pp. 73-87 ◽

Cited By ~ 6

Author(s):

Margaret K Shirra ◽

Karen M Arndt

Keyword(s):

Rna Polymerase Ii ◽

Glucose Repression ◽

Snf1 Kinase ◽

Glucose Levels ◽

Recessive Mutations ◽

Mutant Strains ◽

Rna Polymerase Ii Transcription ◽

Two Hybrid

AbstractBinding of the TATA-binding protein (TBP) to the promoter is a pivotal step in RNA polymerase II transcription. To identify factors that regulate TBP, we selected for suppressors of a TBP mutant that exhibits promoter-specific defects in activated transcription in vivo and severely reduced affinity for TATA boxes in vitro. Dominant mutations in SNF4 and recessive mutations in REG1, OPI1, and RTF2 were isolated that specifically suppress the inositol auxotrophy of the TBP mutant strains. OPI1 encodes a repressor of INO1 transcription. REG1 and SNF4 encode regulators of the Glc7 phosphatase and Snf1 kinase, respectively, and have well-studied roles in glucose repression. In two-hybrid assays, one SNF4 mutation enhances the interaction between Snf4 and Snf1. Suppression of the TBP mutant by our reg1 and SNF4 mutations appears unrelated to glucose repression, since these mutations do not alleviate repression of SUC2, and glucose levels have little effect on INO1 transcription. Moreover, mutations in TUP1, SSN6, and GLC7, but not HXK2 and MIG1, can cause suppression. Our data suggest that association of TBP with the TATA box may be regulated, directly or indirectly, by a substrate of Snf1. Analysis of INO1 transcription in various mutant strains suggests that this substrate is distinct from Opi1.

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Direct Interaction between the Subunit RAP30 of Transcription Factor IIF (TFIIF) and RNA Polymerase Subunit 5, Which Contributes to the Association between TFIIF and RNA Polymerase II

Journal of Biological Chemistry ◽

10.1074/jbc.m009634200 ◽

2001 ◽

Vol 276 (15) ◽

pp. 12266-12273 ◽

Cited By ~ 31

Author(s):

Wenxiang Wei ◽

Dorjbal Dorjsuren ◽

Yong Lin ◽

Weiping Qin ◽

Takahiro Nomura ◽

...

Keyword(s):

Rna Polymerase ◽

Rna Polymerase Ii ◽

Middle Part ◽

Wild Type ◽

Binding Region ◽

Pol Ii ◽

Transcription Factor Iif ◽

B Virus

The general transcription factor IIF (TFIIF) assembled in the initiation complex, and RAP30 of TFIIF, have been shown to associate with RNA polymerase II (pol II), although it remains unclear which pol II subunit is responsible for the interaction. We examined whether TFIIF interacts with RNA polymerase II subunit 5 (RPB5), the exposed domain of which binds transcriptional regulatory factors such as hepatitis B virus X protein and a novel regulatory protein, RPB5-mediating protein. The results demonstrated that RPB5 directly binds RAP30in vitrousing purified recombinant proteins andin vivoin COS1 cells transiently expressing recombinant RAP30 and RPB5. The RAP30-binding region was mapped to the central region (amino acids (aa) 47–120) of RPB5, which partly overlaps the hepatitis B virus X protein-binding region. Although the middle part (aa 101–170) and the N-terminus (aa 1–100) of RAP30 independently bound RPB5, the latter was not involved in the RPB5 binding when RAP30 was present in TFIIF complex. Scanning of the middle part of RAP30 by clustered alanine substitutions and then point alanine substitutions pinpointed two residues critical for the RPB5 binding inin vitroandin vivoassays. Wild type but not mutants Y124A and Q131A of RAP30 coexpressed with FLAG-RAP74 efficiently recovered endogenous RPB5 to the FLAG-RAP74-bound anti-FLAG M2 resin. The recovered endogenous RPB5 is assembled in pol II as demonstrated immunologically. Interestingly, coexpression of the central region of RPB5 and wild type RAP30 inhibited recovery of endogenous pol II to the FLAG-RAP74-bound M2 resin, strongly suggesting that the RAP30-binding region of RPB5 inhibited the association of TFIIF and pol II. The exposed domain of RPB5 interacts with RAP30 of TFIIF and is important for the association between pol II and TFIIF.

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Cystine/glutamate antiporter xCT (SLC7A11) facilitates oncogenic RAS transformation by preserving intracellular redox balance

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1821323116 ◽

2019 ◽

Vol 116 (19) ◽

pp. 9433-9442 ◽

Cited By ~ 47

Author(s):

Jonathan K. M. Lim ◽

Alberto Delaidelli ◽

Sean W. Minaker ◽

Hai-Feng Zhang ◽

Milena Colovic ◽

...

Keyword(s):

Oxidative Stress ◽

Transcription Factor ◽

Redox Balance ◽

Gene Encoding ◽

Oncogenic Ras ◽

Opposing Effects ◽

Whole Transcriptome ◽

Intracellular Redox

The RAS family of proto-oncogenes are among the most commonly mutated genes in human cancers and predict poor clinical outcome. Several mechanisms underlying oncogenic RAS transformation are well documented, including constitutive signaling through the RAF-MEK-ERK proproliferative pathway as well as the PI3K-AKT prosurvival pathway. Notably, control of redox balance has also been proposed to contribute to RAS transformation. However, how homeostasis between reactive oxygen species (ROS) and antioxidants, which have opposing effects in the cell, ultimately influence RAS-mediated transformation and tumor progression is still a matter of debate and the mechanisms involved have not been fully elucidated. Here, we show that oncogenic KRAS protects fibroblasts from oxidative stress by enhancing intracellular GSH levels. Using a whole transcriptome approach, we discovered that this is attributable to transcriptional up-regulation of xCT, the gene encoding the cystine/glutamate antiporter. This is in line with the function of xCT, which mediates the uptake of cystine, a precursor for GSH biosynthesis. Moreover, our results reveal that the ETS-1 transcription factor downstream of the RAS-RAF-MEK-ERK signaling cascade directly transactivates the xCT promoter in synergy with the ATF4 endoplasmic reticulum stress-associated transcription factor. Strikingly, xCT was found to be essential for oncogenic KRAS-mediated transformation in vitro and in vivo by mitigating oxidative stress, as knockdown of xCT strongly impaired growth of tumor xenografts established from KRAS-transformed cells. Overall, this study uncovers a mechanism by which oncogenic RAS preserves intracellular redox balance and identifies an unexpected role for xCT in supporting RAS-induced transformation and tumorigenicity.

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Calreticulin Mediates Anesthetic Sensitivity in Drosophila melanogaster

Anesthesiology ◽

10.1097/00000542-200310000-00019 ◽

2003 ◽

Vol 99 (4) ◽

pp. 867-875 ◽

Cited By ~ 16

Author(s):

Sumiko Gamo ◽

Junya Tomida ◽

Katsuyuki Dodo ◽

Dai Keyakidani ◽

Hitoshi Matakatsu ◽

...

Keyword(s):

Drosophila Melanogaster ◽

Double Mutant ◽

Developmental Stages ◽

Northern Blotting ◽

General Anesthetics ◽

Wild Type ◽

Mutant Strains ◽

Low Expression

Background Various species, e.g., Caenorhabditis elegans, Drosophila melanogaster, and mice, have been used to explore the mechanisms of action of general anesthetics in vivo. The authors isolated a Drosophila mutant, ethas311, that was hypersensitive to diethylether and characterized the calreticulin (crc) gene as a candidate of altered anesthetic sensitivity. Methods Molecular analysis of crc included cloning and sequencing of the cDNA, Northern blotting, and in situ hybridization to accomplish the function of the gene and its mutation. For anesthetic phenotype assay, the 50% anesthetizing concentrations were determined for ethas311, revertants, and double-mutant strains (wild-type crc transgene plus ethas311). Results Expression of the crc 1.4-kb transcript was lower in the mutant ethas311 than in the wild type at all developmental stages. The highest expression at 19 h after pupation was observed in the brain of the wild type but was still low in the mutant at that stage. The mutant showed resistance to isoflurane as well as hypersensitivity to diethylether, whereas it showed the wild phenotype to halothane. Both mutant phenotypes were restored to the wild type in the revertants and double-mutant strains. Conclusion ethas311 is a mutation of low expression of the Drosophila calreticulin gene. The authors demonstrated that hypersensitivity to diethylether and resistance to isoflurane are associated with low expression of the gene. In Drosophila, calreticulin seems to mediate these anesthetic sensitivities, and it is a possible target for diethylether and isoflurane, although the predicted anesthetic targets based on many studies in vitro and in vivo are the membrane proteins, such as ion channels and receptors.

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Pathophysiology of Thrombocytopenia and Anemia in Mice Lacking Transcription Factor NF-E2

Blood ◽

10.1182/blood.v94.9.3037 ◽

1999 ◽

Vol 94 (9) ◽

pp. 3037-3047 ◽

Cited By ~ 69

Author(s):

Jack Levin ◽

Jin-Peng Peng ◽

Georgiann R. Baker ◽

Jean-Luc Villeval ◽

Patrick Lecine ◽

...

Keyword(s):

Transcription Factor ◽

Blood Cell ◽

Red Blood Cell ◽

Fetal Life ◽

Wild Type ◽

Cell Lineages ◽

Cell Fragments ◽

Proliferation Potential

Abstract Expression of the p45 subunit of transcription factor NF-E2 is restricted to selected blood cell lineages, including megakaryocytes and developing erythrocytes. Mice lacking p45 NF-E2 show profound thrombocytopenia, resulting from a late arrest in megakaryocyte differentiation, and a number of red blood cell defects, including anisocytosis and hypochromia. Here we report results of studies aimed to explore the pathophysiology of these abnormalities. Mice lacking NF-E2 produce very few platelet-like particles that display highly disorganized ultrastructure and respond poorly to platelet agonists, features consistent with the usually lethal hemorrhage in these animals. Thrombocytopenia was evident during fetal life and was not corrected by splenectomy in adults. Surprisingly, fetal NF-E2–deficient megakaryocyte progenitors showed reduced proliferation potential in vitro. Thus, NF-E2 is required for regulated megakaryocyte growth as well as for differentiation into platelets. All the erythroid abnormalities were reproduced in lethally irradiated wild-type recipients of hematopoietic cells derived from NF-E2-null fetuses. Whole blood from mice lacking p45 NF-E2 showed numerous small red blood cell fragments; however, survival of intact erythrocytes in vivo was indistinguishable from control mice. Considered together, these observations indicate a requirement for NF-E2 in generating normal erythrocytes. Despite impressive splenomegaly at baseline, mice lacking p45 NF-E2 survived splenectomy, which resulted in increased reticulocyte numbers. This reveals considerable erythroid reserve within extra-splenic sites of hematopoiesis and suggests a role for the spleen in clearing abnormal erythrocytes. Our findings address distinct aspects of the requirements for NF-E2 in blood cell homeostasis and establish its roles in proper differentiation of megakaryocytes and erythrocytes.

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GA Binding Protein (GABP) Is Required for Development and Maturation of Myeloid Cells.

Blood ◽

10.1182/blood.v104.11.776.776 ◽

2004 ◽

Vol 104 (11) ◽

pp. 776-776

Author(s):

Zhongfa Yang ◽

Alan G. Rosmarin

Keyword(s):

Cell Cycle ◽

Transcription Factor ◽

Target Genes ◽

Embryonic Stem ◽

Myeloid Cells ◽

Transactivation Domain ◽

Wild Type ◽

Floxed Allele

Abstract GABP is an ets transcription factor that regulates transcription of key myeloid genes, including CD18 (beta2 leukocyte integrin), neutrophil elastase, lysozyme, and other key mediators of the inflammatory response; it is also known to regulate important cell cycle control genes. GABP consists of two distinct and unrelated proteins that, together, form a functional transcription factor complex. GABPalpha (GABPa) is an ets protein that binds to DNA; it forms a tetrameric complex by recruiting its partner, GABPbeta (GABPb), which contains the transactivation domain. GABPa is a single copy gene in both the human and murine genomes and it is the only protein that can recruit GABPb to DNA. We cloned GABPa from a murine genomic BAC library and prepared a targeting vector in which exon 9 (which encodes the GABPa ets domain) was flanked by loxP (floxed) recombination sites. The targeting construct was electroporated into embryonic stem cells, homologous recombinants were implanted into pseudopregnant mice, heterozygous floxed GABPa mice were identified, and intercrossing yielded expected Mendelian ratios of wild type, heterozygous, and homozygous floxed GABPa mice. Breeding of heterozygous floxed GABPa mice to CMV-Cre mice (which express Cre recombinase in all tissues) yielded expected numbers of hemizygous mice (only one intact GABPa allele), but no nullizygous (GABPa−/−) mice among 64 pups; we conclude that homozygous deletion of GABPa causes an embryonic lethal defect. To determine the effect of GABPa deletion on myeloid cell development, we bred heterozygous and homozygous floxed mice to LysMCre mice, which express Cre only in myeloid cells. These mice had a normal complement of myeloid cells but, unexpectedly, PCR indicated that their Gr1+ myeloid cells retained an intact (undeleted) floxed GABPa allele. We detected similar numbers of in vitro myeloid colonies from bone marrow of wild type, heterozygous floxed, and homozygous floxed progeny of LysMCre matings. However, PCR of twenty individual in vitro colonies from homozygous floxed mice indicated that they all retained an intact floxed allele. Breeding of floxed GABPa/LysMCre mice with hemizygous mice indicated that retention of a floxed allele was not due to incomplete deletion by LysMCre; rather, it appears that only myeloid cells that retain an intact GABPa allele can survive to mature in vitro or in vivo. We prepared murine embryonic fibroblasts from homozygous floxed mice and efficiently deleted GABPa in vitro. We found striking abnormalities in proliferation and G1/S phase arrest. We used quantitative RT-PCR to identify mechanisms that account for the altered growth of GABPa null cells. We found dramatically reduced expression of known GABP target genes that regulate DNA synthesis and cell cycle that appear to account for the proliferative defect. We conclude that GABPa is required for growth and maturation of myeloid cells and we identified downstream targets that may account for their failure to proliferate and mature in vitro and in vivo.

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GABP Transcription Factor Is Required for Myeloid Cell Development In Vivo.

Blood ◽

10.1182/blood.v110.11.374.374 ◽

2007 ◽

Vol 110 (11) ◽

pp. 374-374 ◽

Cited By ~ 2

Author(s):

Zhong-fa Yang ◽

Karen Drumea ◽

Alan G. Rosmarin

Keyword(s):

Bone Marrow ◽

Transcription Factor ◽

T Cell ◽

Myeloid Cells ◽

Myeloid Cell ◽

Cell Development ◽

Wild Type ◽

Ets Domain

Abstract GABP is an ets transcription factor that regulates genes that are required for innate immunity, including CD18 (β2 leukocyte integrin), lysozyme, and neutrophil elastase. GABP consists of two distinct and unrelated proteins. GABPα binds to DNA through its ets domain and recruits GABPβ, which contains the transactivation domain; together, they form a functional tetrameric transcription factor complex. We recently showed that GABP is required for entry into S phase of the cell cycle through its regulation of genes that are required for DNA synthesis and cyclin dependent kinase inhibitors (Yang, et al. Nature Cell Biol9:339, 2007). Furthermore, GABP is an essential component of a retinoic acid responsive myeloid enhanceosome (Resendes and Rosmarin Mol Cell Biol26:3060, 2006). We cloned Gabpa (the gene that encodes mouse Gabpα) from a mouse genomic BAC library and prepared a targeting vector in which the ets domain is flanked by loxP recombination sites (floxed allele). Deletion of both floxed Gabpa alleles causes an early embryonic lethal defect. In order to define the role of Gabpα in myelopoiesis, we bred floxed Gabpa mice to mice that bear the Mx1-Cre transgene, which drives expression of Cre recombinase in response to injection of the synthetic polynucleotide, poly I-C. Deletion of Gabpa dramatically reduced granulocytes and monocytes in the peripheral blood, spleen, and bone marrow, but myeloid cells recovered within weeks. In vitro colony forming assays indicated that myeloid cells in these mice were derived only from Gabpa replete myeloid precursors (that failed to delete both Gabpa alleles), suggesting strong pressure to retain Gabpα in vivo. We used a novel competitive bone marrow transplantation approach to determine if Gabp is required for myeloid cell development in vivo. Sub-lethally irradiated wild-type recipient mice bearing leukocyte marker CD45.1 received equal proportions of bone marrow from wild type CD45.1 donor mice and floxed-Mx1-Cre donor mice that bear CD45.2. Both the CD45.2 (floxed-Mx1-Cre) and CD45.1 (wild type) bone marrow engrafted well. Mice were then injected with pI-pC to induce Cre-mediated deletion of floxed Gabpa. The mature myeloid and T cell compartments were derived almost entirely from wild type CD45.1 cells. This indicates that the proliferation and/or differentiation of myeloid and T cell lineages requires Gabp. In contrast, B cell development was not impaired. We conclude that Gabpa disruption causes a striking loss of myeloid cells in vivo and corroborates prior in vitro data that GABP plays a crucial role in proliferation of myeloid progenitor cells.

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c-Jun Homodimers Can Function as a Context-Specific Coactivator

Molecular and Cellular Biology ◽

10.1128/mcb.00936-06 ◽

2007 ◽

Vol 27 (8) ◽

pp. 2919-2933 ◽

Cited By ~ 33

Author(s):

Benoit Grondin ◽

Martin Lefrancois ◽

Mathieu Tremblay ◽

Marianne Saint-Denis ◽

André Haman ◽

...

Keyword(s):

Gene Expression ◽

Transcription Factors ◽

Dna Binding ◽

Rna Polymerase Ii ◽

Protein Interactions ◽

Expression Profiles ◽

Basic Domain ◽

Interaction Mode

ABSTRACT Transcription factors can function as DNA-binding-specific activators or as coactivators. c-Jun drives gene expression via binding to AP-1 sequences or as a cofactor for PU.1 in macrophages. c-Jun heterodimers bind AP-1 sequences with higher affinity than homodimers, but how c-Jun works as a coactivator is unknown. Here, we provide in vitro and in vivo evidence that c-Jun homodimers are recruited to the interleukin-1β (IL-1β) promoter in the absence of direct DNA binding via protein-protein interactions with DNA-anchored PU.1 and CCAAT/enhancer-binding protein β (C/EBPβ). Unexpectedly, the interaction interface with PU.1 and C/EBPβ involves four of the residues within the basic domain of c-Jun that contact DNA, indicating that the capacities of c-Jun to function as a coactivator or as a DNA-bound transcription factor are mutually exclusive. Our observations indicate that the IL-1β locus is occupied by PU.1 and C/EBPβ and poised for expression and that c-Jun enhances transcription by facilitating a rate-limiting step, the assembly of the RNA polymerase II preinitiation complex, with minimal effect on the local chromatin status. We propose that the basic domain of other transcription factors may also be redirected from a DNA interaction mode to a protein-protein interaction mode and that this switch represents a novel mechanism regulating gene expression profiles.

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Mutations in RNA Polymerase II and Elongation Factor SII Severely Reduce mRNA Levels in Saccharomyces cerevisiae

Molecular and Cellular Biology ◽

10.1128/mcb.18.10.5771 ◽

1998 ◽

Vol 18 (10) ◽

pp. 5771-5779 ◽

Cited By ~ 39

Author(s):

J. Cale Lennon ◽

Megan Wind ◽

Laura Saunders ◽

M. Benjamin Hock ◽

Daniel Reines

Keyword(s):

Rna Polymerase ◽

Rna Polymerase Ii ◽

Genetic Interaction ◽

Elongation Factor ◽

Mrna Levels ◽

Wild Type ◽

Mutant Cells ◽

Rna Levels

ABSTRACT Elongation factor SII interacts with RNA polymerase II and enables it to transcribe through arrest sites in vitro. The set of genes dependent upon SII function in vivo and the effects on RNA levels of mutations in different components of the elongation machinery are poorly understood. Using yeast lacking SII and bearing a conditional allele of RPB2, the gene encoding the second largest subunit of RNA polymerase II, we describe a genetic interaction between SII and RPB2. An SII gene disruption or therpb2-10 mutation, which yields an arrest-prone enzyme in vitro, confers sensitivity to 6-azauracil (6AU), a drug that depresses cellular nucleoside triphosphates. Cells with both mutations had reduced levels of total poly(A)+ RNA and specific mRNAs and displayed a synergistic level of drug hypersensitivity. In cells in which the SII gene was inactivated, rpb2-10 became dominant, as if template-associated mutant RNA polymerase II hindered the ability of wild-type polymerase to transcribe. Interestingly, while 6AU depressed RNA levels in both wild-type and mutant cells, wild-type cells reestablished normal RNA levels, whereas double-mutant cells could not. This work shows the importance of an optimally functioning elongation machinery for in vivo RNA synthesis and identifies an initial set of candidate genes with which SII-dependent transcription can be studied.

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