TBP mutants defective in activated transcription in vivo.

Arndt, Karen M.; Ricupero-Hovasse, S L; Winston, Fred

doi:10.1002/j.1460-2075.1995.tb07135.x

Cited by 86 publications

(95 citation statements)

References 75 publications

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“…The lack of TATA element binding in vitro does not conflict with our conclusion that the TBP mutants directly associate with TATA elements in vivo. In particular, there are a number of TBP derivatives that do not detectably associate with TATA elements in vitro yet are fully competent to support yeast cell viability and transcription in vivo (1,41). This apparent discrepancy is probably due to the fact that TBP association with TATA elements in vivo is stabilized by TAFs and other components of the Pol II machinery (40,44) and hence is not simply due to its inherent DNA-binding affinity.…”

Section: Resultsmentioning

confidence: 99%

“…Direct physical analysis by protein-DNA cross-linking in vivo indicates that, in the vast majority of cases tested, TBP does not associate with core promoters in the absence of a functional activator (40,44). TBP mutants that weaken TATA element binding (1,41) or TFIIA association (4,53) can selectively affect the response to certain activators. Such selective effects on activator-dependent transcription might be related to the observation in vitro that the TATA element is particularly important for supporting multiple rounds of transcription from a preinitiation complex (50,63).…”

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confidence: 99%

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TATA-Binding Protein Mutants That Increase Transcription from Enhancerless and Repressed Promoters In Vivo

Geisberg

Struhl

2000

Molecular and Cellular Biology

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Using a genetic screen, we isolated three TATA-binding protein (TBP) mutants that increase transcription from promoters that are repressed by the Cyc8-Tup1 or Sin3-Rpd3 corepressors or that lack an enhancer element, but not from an equivalently weak promoter with a mutated TATA element. Increased transcription is observed when the TBP mutants are expressed at low levels in the presence of wild-type TBP. These TBP mutants are unable to support cell viability, and they are toxic in strains lacking Rpd3 histone deacetylase or when expressed at higher levels. Although these mutants do not detectably bind TATA elements in vitro, genetic and chromatin immunoprecipitation experiments indicate that they act directly at promoters and do not increase transcription by titration of a negative regulatory factor(s). The TBP mutants are mildly defective for associating with promoters responding to moderate or strong activators; in addition, they are severely defective for RNA polymerase (Pol) III but not Pol I transcription. These results suggest that, with respect to Pol II transcription, the TBP mutants specifically increase expression from core promoters. Biochemical analysis indicates that the TBP mutants are unaffected for TFIID complex formation, dimerization, and interactions with either the general negative regulator NC2 or the N-terminal inhibitory domain of TAF130. We speculate that these TBP mutants have an unusual structure that allows them to preferentially access TATA elements in chromatin templates. These TBP mutants define a criterion by which promoters repressed by Cyc8-Tup1 or Sin3-Rpd3 resemble enhancerless, but not TATA-defective, promoters; hence, they support the idea that these corepressors inhibit the function of activator proteins rather than the Pol II machinery.

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Section: Resultsmentioning

confidence: 99%

mentioning

confidence: 99%

TATA-Binding Protein Mutants That Increase Transcription from Enhancerless and Repressed Promoters In Vivo

Geisberg

Struhl

2000

Molecular and Cellular Biology

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“…Furthermore, npl3-27 exerts a dominant effect on spt15-ts1 cells and not wild-type cells in that expression of exogenous npl3-27 over endogenous wild-type NPL3 causes an mRNA export defect in spt15-ts1 cells. We tested other ts − alleles of spt15 (spt15-328 and spt15-341; Arndt et al 1995) in combination with npl3-27 and found that these alleles also displayed nuclear accumulation of Npl3-27 and mRNA. To determine the specificity of this genetic interaction, we examined strains mutated for RNA Pol II that decrease transcription (rpb1-1, rpb1⌬101, rpb1⌬103), but these strains do not display Npl3-27 or mRNA export defects (data not shown).…”

Section: A Screen For Npl3 Export Factors Identifies Spt15 Which Encmentioning

confidence: 99%

Messenger RNAs are recruited for nuclear export during transcription

Lei¹,

Krebber²,

Silver³

2001

Genes Dev.

205

195

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Following transcription and processing, eukaryotic mRNAs are exported from the nucleus to the cytoplasm for translation. Here we present evidence that mRNAs are targeted for nuclear export cotranscriptionally. Combined mutations in the Saccharomyces cerevisiae hnRNP Npl3 and TATA-binding protein (TBP) block mRNA export, implying that cotranscriptional recruitment of Npl3 is required for efficient export of mRNA. Furthermore, Npl3 can be found in a complex with RNA Pol II, indicating that Npl3 associates with the transcription machinery. Finally, Npl3 is recruited to genes in a transcription dependent manner as determined by chromatin immunoprecipitation. Another mRNA export factor, Yra1, also associates with chromatin cotranscriptionally but appears to be recruited at a later step. Taken together, our results suggest that export factors are recruited to the sites of transcription to promote efficient mRNA export. Compartmentalization of the eukaryotic cell necessitates the movement of macromolecules through the nuclear envelope. In particular, mRNAs must be transported from the site of transcription in the nucleus to the cytoplasm for translation to occur. Export of mRNAs requires processing, packaging by RNA-binding proteins, recognition by export factors, and translocation through the nuclear pore complex (NPC) into the cytoplasm. mRNAs transcribed by RNA Pol II undergo rapid processing in the nucleus. Pre-mRNA processing steps include addition of a 5Ј monomethyl cap, splicing of introns, and 3Ј cleavage and polyadenylation. 5Ј capping occurs on nascent transcripts soon after transcription initiation (Salditt-Georgieff et al. 1980;Jove and Manley 1984;Rasmussen and Lis 1993). Furthermore, capping enzyme is associated with Pol II and can be found associated with promoters of transcribed genes (Cho et al. 1997;McCracken et al. 1997;Komarnitsky et al. 2000;Schroeder et al. 2000). Additionally, the transcription machinery interacts with splicing and 3Ј cleavage and polyadenylation factors, suggesting that these events occur either cotranscriptionally or soon after transcription termination (Mortillaro et al. 1996;Yuryev et al. 1996;Dantonel et al. 1997). Coordination of mRNA processing events and transcription is thought to be primarily through Pol II and its carboxy-terminal domain (CTD) in a manner that can be dependent on the state of CTD phosphorylation (for review, see Hirose and Manley 2000).In the nucleus, hnRNPs and other RNA-binding proteins package mRNAs into ribonucleoprotein particles (RNPs), and a subset of these proteins remain in the nucleus whereas others accompany the RNP into the cytoplasm, dissociate, and move back into the nucleus for further rounds of export. In the visually tractable Chironomus tentans, it is possible to study the very large (35-40 kb) Balbiani ring (BR) RNA throughout its various maturation stages, including transcription, nuclear export, and translation. Previous immunoelectron microscopy studies showed that the hnRNPs hrp36, hrp45, and hrp23 associate with the BR particle ...

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“…MATa and MATa strains used in this study were derived from S. cerevisiae ScTEB652 (MATa ade1 ile leu2-3,112 ura3-52 trp1-HIII his3-D1 MEL1) and FY630 (MATa his4-917d lys2-173R2 leu2D1 ura3-52 trp1D63), respectively (Arndt et al 1995;Blank et al 1997). Details of strain construction will be provided upon request.…”

Section: Strainsmentioning

confidence: 99%

Rapid GAL Gene Switch of Saccharomyces cerevisiae Depends on Nuclear Gal3, Not Nucleocytoplasmic Trafficking of Gal3 and Gal80

Egriboz¹,

Jiang

Hopper

2011

Genetics

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The yeast transcriptional activator Gal4 localizes to UAS GAL sites even in the absence of galactose but cannot activate transcription due to an association with the Gal80 protein. By 4 min after galactose addition, Gal4-activated gene transcription ensues. It is well established that this rapid induction arises through a galactose-triggered association between the Gal80 and Gal3 proteins that decreases the association of Gal80 and Gal4. How this happens mechanistically remains unclear. Strikingly different hypotheses prevail concerning the possible roles of nucleocytoplasmic distribution and trafficking of Gal3 and Gal80 and where in the cell the initial Gal3-Gal80 association occurs. Here we tested two conflicting hypotheses by evaluating the subcellular distribution and dynamics of Gal3 and Gal80 with reference to induction kinetics. We determined that the rates of nucleocytoplasmic trafficking for both Gal80 and Gal3 are slow relative to the rate of induction. We find that depletion of the nuclear pool of Gal3 slows the induction kinetics. Thus, nuclear Gal3 is critical for rapid induction. Fluorescence-recovery-after-photobleaching experiments provided data suggesting that the Gal80-Gal4 complex exhibits kinetic stability in the absence of galactose. Finally, we detect Gal3 at the UAS GAL only if Gal80 is covalently linked to the DNA-binding domain. Taken altogether, these new findings lead us to propose that a transient interaction of Gal3 with Gal4-associated Gal80 could explain the rapid response of this system. This notion could also explain earlier observations.

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TBP mutants defective in activated transcription in vivo.

Cited by 86 publications

References 75 publications

TATA-Binding Protein Mutants That Increase Transcription from Enhancerless and Repressed Promoters In Vivo

TATA-Binding Protein Mutants That Increase Transcription from Enhancerless and Repressed Promoters In Vivo

Messenger RNAs are recruited for nuclear export during transcription

Rapid GAL Gene Switch of Saccharomyces cerevisiae Depends on Nuclear Gal3, Not Nucleocytoplasmic Trafficking of Gal3 and Gal80

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