The<i>PCF1–1</i>mutation increases the activity of the transcription factor (TF) IIIB fraction from<i>Saccharomyces cerevisae</i>

Willis, Ian M.; Oksman, Alexander; López-De-León, Alfredo

doi:10.1093/nar/20.14.3725

Cited by 19 publications

(13 citation statements)

References 34 publications

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“…The biochemical basis of the PCFI-l rate effect is currently under investigation. In addition to increasing the rate of preinitiation complex assembly, single-round initiation assays have demonstrated that the PCFJ-1 mutation also increases the number of transcriptionally active complexes in crude cell extracts (38). Our present findings that TFIIIB fractions purified from mutant cells contain more TFIIIB70 and have elevated TFIIIB" activity suggest that these observations are related.…”

Section: Tfiiib90supporting

confidence: 49%

“…7, lanes 4 and 5). With recombinant TBP as a standard, both fractions were found to contain a significant molar excess (about 100-fold for the mutant TFIIIB fraction) of TBP polypeptide relative to TFIIIB (based on activity measurements [30] cell extracts is reduced from about six-to threefold (38,39).…”

Section: Tfiiib90mentioning

confidence: 99%

“…Both the rate of formation of preinitiation complexes and the number of such complexes which are transcriptionally competent are increased in mutant compared with wild-type cell extracts (38,39). Additionally, fractionation of cell extracts has shown that PCF1-1 augments the activity of a crude TFIIIB fraction and that the activities of the TFIIIC and Pol III fractions are unaffected (38). Since these studies did not establish whether the PCFI gene product was present in the TFIIIB fraction, the factor was proposed to be either a component or a regulator of TFIIIB.…”

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confidence: 99%

“…Studies carried out to date indicate that the PCFl-1 mutation influences two steps in the transcription process. Both the rate of formation of preinitiation complexes and the number of such complexes which are transcriptionally competent are increased in mutant compared with wild-type cell extracts (38,39). Additionally, fractionation of cell extracts has shown that PCF1-1 augments the activity of a crude TFIIIB fraction and that the activities of the TFIIIC and Pol III fractions are unaffected (38).…”

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confidence: 99%

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A mutation in the second largest subunit of TFIIIC increases a rate-limiting step in transcription by RNA polymerase III.

Rameau¹,

Puglia²,

Crowe³

et al. 1994

Mol. Cell. Biol.

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In previous studies, we have shown that the PCFI-I mutation of Saccharomyces cerevisiae suppresses the negative effect of a tRNA gene A block promoter mutation in vivo and increases the transcription of a variety of RNA polymerase III genes in vitro. Here, we report that PCF1 encodes the second largest subunit of transcription factor IIIC (TFIIIC) and that the PCFI-1 mutation causes an amino acid substitution in a novel protein structural motif, a tetratricopeptide repeat, in this subunit. polypeptide is the only TFIIIC subunit that can be photocross-linked to the region footprinted by TFIIIB. This subunit is a likely candidate to mediate the recruitment of TFIIIB. Similarly, the 95-and 55-kDa subunits may interact with the A block, and the 138-kDa subunit may interact with the B block. To help resolve the roles of individual TFIIIC subunits in transcription complex assembly, a major effort to clone these genes has been under way. So far, this has been achieved for the 95-, 131-, and 138-kDa polypeptides (23,26,29,34).Studies with yeast and human cells indicate that TFIIIB is a multisubunit factor (20,24,35,36). Detailed characterization of the yeast factor suggests that it comprises three components: the TATA-binding protein (TBP), a 70-kDa TFIIB-like polypeptide (TFIIIB70), and a 90-kDa polypeptide (TFIIIB90). These three polypeptides are stably associated under most conditions and copurify as a complex with TFIIIB activity over numerous columns. However, by using strong cation exchangers, specifically MonoS, TFIIIB activity has been separated into two fractions, designated TFIIIB ' and TFIIIB" (18). By photocross-linking, these two fractions were found to contain the TFIIIB70 and TFIIIBgo polypeptides, respectively, which had been identified previously in less pure TFIIIB fractions with this technique (3). Following the demonstration of a universal role for TBP in eukaryotic transcription (7, 32), TBP was identified as a component of TFIIIB and traced by Western blotting (immunoblotting) to the TFIIIB' fraction (20). This result together with the cloning of the gene for TFIIIB70 (5,6,25) has permitted the demonstration that these proteins are the only TFIIIB components in the TFIIIB' fraction. The two proteins expressed in bacteria can replace the TFIIIB' fraction in a reconstituted transcription system (20). The TFIIIB" fraction has not been purified to homogeneity. However, TFIIIB90 may be the only TFIIIB subunit in this fraction, since TFIIIB" transcription activity can be provided by sodium dodecyl sulfate (SDS) gel-eluted and renatured proteins in the 90-kDa size range (20

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Section: Tfiiib90supporting

confidence: 49%

Section: Tfiiib90mentioning

confidence: 99%

mentioning

confidence: 99%

mentioning

confidence: 99%

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A mutation in the second largest subunit of TFIIIC increases a rate-limiting step in transcription by RNA polymerase III.

Rameau¹,

Puglia²,

Crowe³

et al. 1994

Mol. Cell. Biol.

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“…One intriguing idea is that the TFIIIB factor, which binds immediately upstream of the pol III transcription initiation site, might be inhibiting pol II transcription by serving as a competitive ligand for components of the pol II transcription machinery. TFIIIB is composed of several subunits, including the TATA-binding protein (3,40,50,53,66,67,92,103,104). If this factor, or any other component of the pol III complex (7,9,17,19,21,48,51,63,91,105), bound tightly to the activation domains of factors bound at the UAS elements, interactions with the pol II basal promoter complex could be competed for (Fig.…”

Section: Resultsmentioning

confidence: 99%

tRNA genes as transcriptional repressor elements.

Hull¹,

Erickson²,

Johnston³

et al. 1994

Mol. Cell. Biol.

120

129

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Eukaryotic genomes frequently contain large numbers of repetitive RNA polymerase III (pol III) promoter elements interspersed between and within RNA pol II transcription units, and in several instances a regulatory relationship between the two types of promoter has been postulated. In the budding yeast Saccharomyces cerevisiae, tRNA genes are the only known interspersed pol III promoter-containing repetitive elements, and we find that they strongly inhibit transcription from adjacent pol II promoters in vivo. This inhibition requires active transcription of the upstream tRNA gene but is independent of its orientation and appears not to involve simple steric blockage of the pol II upstream activator sites. Evidence is presented that different pol II promoters can be repressed by different tRNA genes placed upstream at varied distances in both orientations.To test whether this phenomenon functions in naturally occurring instances in which tRNA genes and pol II promoters are juxtaposed, we examined the sigma and Ty3 elements. This class of retrotransposons is always found integrated immediately upstream of different tRNA genes. Weakening tRNA gene transcription by means of a temperature-sensitive mutation in RNA pol III increases the pheromone-inducible expression of sigma and Ty3 elements up to 60-fold.Many eukaryotic genomes contain families of moderately to highly repeated DNA elements containing RNA polymerase III (pol III) promoters (reviewed in references 74 and 75). Frequently these elements resemble the intragenic pol III promoter class found in tRNA and 7SL RNA genes, which consist of consensus A-box and B-box sequences downstream from the transcription start sites. These elements can be found either dispersed as individual copies or as highly reiterated tandem copies, especially in heterochromatic regions. The pol III elements are not generally transcribed into stable RNA commensurate with their copy number in vivo, although they can usually be transcribed in vitro, and there are numerous reports of condition-specific or development-specific activation in vivo (10,27,61,90,95,97,101). Several hypotheses have been put forward regarding possible functions for these sequences, but one particularly interesting suggestion is that dispersed RNA pol III promoters might exert either a positive or negative influence on the transcriptional activity of overlapping or nearby RNA pol II promoters (11,12,15,89,90,96). In some cases, cryptic pol III promoter elements directly interfere with factor binding sites in the pol II promoter upstream region or with the pol II initiation site itself. In at least one report, however, repression was achieved by an Alu repetitive element, in which case there was no obvious steric overlap with the neighboring pol II promoter (96).In this report, the question of whether RNA pol III promoters can exert negative transcriptional regulation on neighboring DNA has been approached by studying the budding yeast Saccharomyces cerevisiae. Although this yeast does not appear to have any Alu-ty...

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A suppressor of mutations in the class III transcription system encodes a component of yeast TFIIIB.

et al. 1996

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Class III genes depend on TFIIIB for recruitment of RNA polymerase III. Yeast TFIIIB is comprised of three components: TBP, TFIIIB70 and a 90 kDa polypeptide contained in the fraction B″. We report the isolation of the yeast gene TFC7 which, based on genetic and biochemical evidence, encodes the 90 kDa polypeptide. TFC7 was isolated as a multicopy suppressor of temperature‐sensitive mutations in the two largest subunits of TFIIIC. It is an essential gene, encoding a polypeptide of 68 kDa migrating with an apparent size of approximately 90 kDa. In gel shift assays, recombinant TFC7 protein (rTFC7) alone did not bind detectably to DNA, or to the TFIIIC‐DNA complex even in the presence of TBP or TFIIIB70, but it was required to assemble the TFIIIB‐TFIIIC‐DNA complex. The two‐hybrid assay pointed to an interaction between TFC7 protein and tau 131, the second largest subunit of TFIIIC (that also interacts with TFIIIB70). rTFC7p can replace the B″ component of TFIIIB for synthesis of U6 RNA in a system reconstituted with recombinant TBP and TFIIIB70 polypeptides and highly purified RNA polymerase III. Surprisingly, specific transcription of the SUP4 tRNATyr gene promoted by rTFC7p was much weaker than with B″. An additional factor activity, provided by the recently identified TFIIIE fraction, was required to restore control levels of transcription.

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ThePCF1–1mutation increases the activity of the transcription factor (TF) IIIB fraction fromSaccharomyces cerevisae

Cited by 19 publications

References 34 publications

A mutation in the second largest subunit of TFIIIC increases a rate-limiting step in transcription by RNA polymerase III.

A mutation in the second largest subunit of TFIIIC increases a rate-limiting step in transcription by RNA polymerase III.

tRNA genes as transcriptional repressor elements.

A suppressor of mutations in the class III transcription system encodes a component of yeast TFIIIB.

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