Binding of TBP to promoters in vivo is stimulated by activators and requires Pol II holoenzyme

Kuras, Laurent; Struhl, Kevin

doi:10.1038/21239

Cited by 439 publications

(465 citation statements)

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“…Recruitment has also been inferred from the results of "activator bypass" experiments in which artificial recruitment of TBP to promoter sites near the TATA element resulted in transcriptional activation, implying that TBP recruitment is a rate-limiting step in transcriptional activation in vivo (10,39,79). Most convincingly, chromatin immunoprecipitation experiments revealed TBP to be physically associated with promoters of numerous genes under activated, but not nonactivated, conditions, implying that recruitment accompanies activation (43,46).…”

mentioning

confidence: 86%

“…Transcription in eukaryotes is often initiated by activator proteins that bind to upstream sites in promoters, and a strong case, built on an impressive wealth of evidence, has been made that these activators use protein-protein interactions to recruit components of the transcriptional apparatus (6,18,21,43,46,60). However, discussions of the recruitment model have not generally explicitly considered how this mechanism allows repressive effects of chromatin to be overcome.…”

Section: Discussionmentioning

confidence: 99%

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Artificially Recruited TATA-Binding Protein Fails To Remodel Chromatin and Does Not Activate Three Promoters That Require Chromatin Remodeling

Ryan

Stafford

et al. 2000

Molecular and Cellular Biology

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Transcriptional activators are believed to work in part by recruiting general transcription factors, such as TATA-binding protein (TBP) and the RNA polymerase II holoenzyme. Activation domains also contribute to remodeling of chromatin in vivo. To determine whether these two activities represent distinct functions of activation domains, we have examined transcriptional activation and chromatin remodeling accompanying artificial recruitment of TBP in yeast (Saccharomyces cerevisiae). We measured transcription of reporter genes with defined chromatin structure by artificial recruitment of TBP and found that a reporter gene whose TATA element was relatively accessible could be activated by artificially recruited TBP, whereas two promoters, GAL10 and CHA1, that have accessible activator binding sites, but nucleosomal TATA elements, could not. A third reporter gene containing the HIS4 promoter could be activated by GAL4-TBP only when a RAP1 binding site was present, although RAP1 alone could not activate the reporter, suggesting that RAP1 was needed to open the chromatin structure to allow activation. Consistent with this interpretation, artificially recruited TBP was unable to perturb nucleosome positioning via a nucleosomal binding site, in contrast to a true activator such as GAL4, or to perturb the TATA-containing nucleosome at the CHA1 promoter. Finally, we show that activation of the GAL10 promoter by GAL4, which requires chromatin remodeling, can occur even in swi gcn5 yeast, implying that remodeling pathways independent of GCN5, the SWI-SNF complex, and TFIID can operate during transcriptional activation in vivo.Transcriptional activators are thought to stimulate transcription of TATA-containing promoters in part by recruiting TFIID, a multiprotein complex consisting of the TATA-binding protein (TBP) and TBP-associated factors (TAFs), to the TATA box (25,60). Several in vitro and in vivo studies support this model. For example, transcription initiated at a mutated TATA element by induced synthesis of TBP with altered specificity was enhanced in both rate and extent in the presence of an activator that could bind upstream of the relevant promoter, consistent with activator-mediated recruitment (40). Recruitment has also been inferred from the results of "activator bypass" experiments in which artificial recruitment of TBP to promoter sites near the TATA element resulted in transcriptional activation, implying that TBP recruitment is a rate-limiting step in transcriptional activation in vivo (10,39,79). Most convincingly, chromatin immunoprecipitation experiments revealed TBP to be physically associated with promoters of numerous genes under activated, but not nonactivated, conditions, implying that recruitment accompanies activation (43,46).A potential obstacle to recruitment of TBP in vivo is posed by chromatin. Binding of TBP to nucleosomal TATA elements is greatly impeded in vitro (23, 32). Transcription is also strongly repressed by chromatin, but this repression can be largely alleviated by binding of act...

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confidence: 86%

Section: Discussionmentioning

confidence: 99%

Artificially Recruited TATA-Binding Protein Fails To Remodel Chromatin and Does Not Activate Three Promoters That Require Chromatin Remodeling

Ryan

Stafford

et al. 2000

Molecular and Cellular Biology

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“…ChIP Analysis-ChIP was performed essentially as described (32). Briefly, 100-ml cultures were grown to log phase (A 600 ϳ0.8 -0.9).…”

Section: Methodsmentioning

confidence: 99%

Actively Transcribed GAL Genes Can Be Physically Linked to the Nuclear Pore by the SAGA Chromatin Modifying Complex

Luthra

Kerr

Harreman

et al. 2007

Journal of Biological Chemistry

119

112

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Recent work has demonstrated that some actively transcribed genes closely associate with nuclear pore complexes (NPC) at the nuclear periphery. The Saccharomyces cerevisiae Mlp1 and Mlp2 proteins are components of the inner nuclear basket of the nuclear pore that mediate interactions with these active genes. To investigate the physical link between the NPC and active loci, we identified proteins that interact with the carboxyl-terminal globular domain of Mlp1 by tandem affinity purification coupled with mass spectrometry. This analysis led to the identification of several components of the Spt-Ada-Gcn5-acetyltransferase (SAGA) histone acetyltransferase complex, Gcn5, Ada2, and Spt7. We utilized co-immunoprecipitation and in vitro binding assays to confirm the interaction between the Mlp proteins and SAGA components. Chromatin immunoprecipitation experiments revealed that Mlp1 and SAGA components associate with the same region of the GAL promoters. Critically, this Mlp-promoter interaction depends on the integrity of the SAGA complex. These results identify a physical association between SAGA and the NPC, and support previous results that relied upon visualization of GAL loci at the nuclear periphery by microscopy (Cabal, G. G. Genovesio, A., Rodriguez-Navarro, S., Zimmer, C., Gadal, O., Lesne, A., Buc, H., Feuerbach-Fournier, F., Olivo-Marin, J.-C., Hurt, E. C., and Nehrbass, U. (2006) Nature 441, 770 -773). We propose that a physical interaction between nuclear pore components and the SAGA complex can link the actively transcribed GAL genes to the nuclear pore.

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“…G1-arrested cells were released into fresh media and ϳ20 OD of cells were harvested every 12 min. Cells were fixed and chromatin immunoprecipitates were prepared essentially as described (Kuras and Struhl, 1999), using IgG-agarose to immunoprecipitate TAP-tagged Mcm2 from wild-type and TetO 7 -SMC4 strains. Immunoprecipitated DNA was analyzed by PCR using four primer pairs for specific regions at and around ARS1 as described (Tanaka et al, 1997).…”

Section: Chromatin Immunoprecipitationmentioning

confidence: 99%

A Survey of Essential Gene Function in the Yeast Cell Division Cycle

Peña‐Castillo

Mnaimneh

et al. 2006

MBoC

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Mutations impacting specific stages of cell growth and division have provided a foundation for dissecting mechanisms that underlie cell cycle progression. We have undertaken an objective examination of the yeast cell cycle through flow cytometric analysis of DNA content in TetO 7 promoter mutant strains representing 75% of all essential yeast genes. More than 65% of the strains displayed specific alterations in DNA content, suggesting that reduced function of an essential gene in most cases impairs progression through a specific stage of the cell cycle. Because of the large number of essential genes required for protein biosynthesis, G1 accumulation was the most common phenotype observed in our analysis. In contrast, relatively few mutants displayed S-phase delay, and most of these were defective in genes required for DNA replication or nucleotide metabolism. G2 accumulation appeared to arise from a variety of defects. In addition to providing a global view of the diversity of essential cellular processes that influence cell cycle progression, these data also provided predictions regarding the functions of individual genes: we identified four new genes involved in protein trafficking (NUS1, PHS1, PGA2, PGA3), and we found that CSE1 and SMC4 are important for DNA replication. INTRODUCTIONCell division is a fundamental biological process that in all organisms consists of a series of closely coordinated events. In the budding yeast Saccharomyces cerevisiae, the cell division cycle begins with an initial growth phase, G1, during which time the cell increases in mass and volume. The transition from G1 into S phase is marked by progression through Start (the point of commitment to cell division), initiation of nuclear DNA synthesis, and the emergence of a bud that will form the new daughter cell. S phase is followed by a second growth phase, G2, which is in turn followed by nuclear division, and then cell separation. A very similar series of events occurs in all eukaryotes, with the obvious exception of bud emergence. Because of the fundamental biological importance of cell cycle progression and its importance in both human development and diseases such as cancer, identification of the molecular determinants of specific stages of the eukaryotic cell cycle has been a subject of intense study for several decades.The morphological landmarks of the cell cycle stages in budding yeast, most notably the size of the bud relative to the size of the mother cell, allows the identification of mutants blocked at specific stages of the cell cycle and thereby forms the basis for the classic cell cycle screens (Hartwell et al., 1970;Culotti and Hartwell, 1971;Hartwell, 1971aHartwell, , 1971bHartwell, , 1973Moir et al., 1982). These genetic screens, using conditional temperature-sensitive mutants, identified more than 50 genes that are required for specific stages in the cell division cycle and so were termed CDC genes. On average 4.6 alleles were identified for each CDC gene in the original screens, suggesting the number of cdc mutan...

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Binding of TBP to promoters in vivo is stimulated by activators and requires Pol II holoenzyme

Cited by 439 publications

References 28 publications

Artificially Recruited TATA-Binding Protein Fails To Remodel Chromatin and Does Not Activate Three Promoters That Require Chromatin Remodeling

Artificially Recruited TATA-Binding Protein Fails To Remodel Chromatin and Does Not Activate Three Promoters That Require Chromatin Remodeling

Actively Transcribed GAL Genes Can Be Physically Linked to the Nuclear Pore by the SAGA Chromatin Modifying Complex

A Survey of Essential Gene Function in the Yeast Cell Division Cycle

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