Signaling activation and repression of RNA polymerase II transcription in yeast

Reece, Richard J.; Platt, Adam

doi:10.1002/bies.950191110

Cited by 33 publications

(21 citation statements)

References 70 publications

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“…The galactose switch, which induces the synthesis of galactose metabolic enzymes in response to this sugar in the environment, serves as a textbook model of how such coupling can be achieved in a eukaryotic cell (for reviews see Refs. [1][2][3]. In Saccharomyces cerevisiae the regulatory module consists of three proteins, the DNA binding activator Gal4, its negative regulator Gal80, and the signal-transducing Gal3 protein.…”

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The Galactose Switch in Kluyveromyces lactis Depends on Nuclear Competition between Gal4 and Gal1 for Gal80 Binding

Anders

Lilie

Franke

et al. 2006

Journal of Biological Chemistry

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The Gal4 protein represents a universally functional transcription activator, which in yeast is regulated by protein-protein interaction of its transcription activation domain with the inhibitor Gal80. Gal80 inhibition is relieved via galactose-mediated Gal80-Gal1-Gal3 interaction. The Gal4-Gal80-Gal1/3 regulatory module is conserved between Saccharomyces cerevisiae and Kluyveromyces lactis. Here we demonstrate that K. lactis Gal80 (KlGal80) is a nuclear protein independent of the Gal4 activity status, whereas KlGal1 is detected throughout the entire cell, which implies that KlGal80 and KlGal1 interact in the nucleus. Consistently KlGal1 accumulates in the nucleus upon KlGAL80 overexpression. Furthermore, we show that the KlGal80-KlGal1 interaction blocks the galactokinase activity of KlGal1 and is incompatible with KlGal80-KlGal4-AD interaction. Thus, we propose that dissociation of KlGal80 from the AD forms the basis of KlGal4 activation in K. lactis. Quantitation of the dissociation constants for the KlGal80 complexes gives a much lower affinity for KlGal1 as compared with Gal4. Mathematical modeling shows that with these affinities a switch based on competition between Gal1 and Gal4 for Gal80 binding is nevertheless efficient provided two monomeric Gal1 molecules interact with dimeric Gal80. Consistent with such a mechanism, analysis of the sedimentation behavior by analytical ultracentrifugation demonstrates the formation of a heterotetrameric KlGal80-KlGal1 complex of 2:2 stoichiometry.Regulation of transcription requires coupling of gene-specific transcription activators to regulatory signals. The galactose switch, which induces the synthesis of galactose metabolic enzymes in response to this sugar in the environment, serves as a textbook model of how such coupling can be achieved in a eukaryotic cell (for reviews see Refs.

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The Galactose Switch in Kluyveromyces lactis Depends on Nuclear Competition between Gal4 and Gal1 for Gal80 Binding

Anders

Lilie

Franke

et al. 2006

Journal of Biological Chemistry

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“…In noninducing conditions, Gal4p is bound to the upstream activating sequences for galactose (UAS G ) but is prevented from activating transcription by the inhibitor Gal80p (32,43). Rapid induction by galactose requires the product of GAL3 (40, 52, 57), which is a regulatory protein with similarity to the galactokinase encoded by GAL1 (3, 9, 50), although Gal3p does not have galactokinase activity (9).…”

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Multiple Signals Regulate GALTranscription in Yeast

Rohde

Trinh

Sadowski

2000

Molecular and Cellular Biology

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Gal4p activates transcription of the Saccharomyces GAL genes in response to galactose and is phosphorylated during interaction with the RNA polymerase II (Pol II) holoenzyme. One phosphorylation at S699 is necessary for full GAL induction and is mediated by Srb10p/CDK8 of the RNA Pol II holoenzyme mediator subcomplex. Gal4p S699 phosphorylation is necessary for sensitive response to inducer, and its requirement for GAL induction can be abrogated by high concentrations of galactose in strains expressing wild-type GAL2 and GAL3. Gal4p S699 phosphorylation occurs independently of Gal3p and is responsible for the long-term adaptation response observed in gal3 yeast. SRB10 and GAL3 are shown to represent parallel mechanisms for GAL gene induction. These results demonstrate that Gal4p activity is controlled by two independent signals: one that acts through Gal3p-galactose and a second that is mediated by the holoenzyme-associated cyclindependent kinase Srb10p. Since Srb10p is regulated independently of galactose, our results suggest a function for CDK8 in coordinating responses to specific inducers with the environment through the phosphorylation of gene-specific activators.Eukaryotic cells react to their environment by regulating transcription factors bound to promoters of responsive genes (28). Cells grown in culture are typically provided with sufficient essential nutrients and factors to ensure unchecked propagation. However, in their natural environment, cell growth is ordinarily limited by the scarcity of one or more factors or nutrients. In such circumstances, there must be mechanisms to ensure that transcriptional responses to one signal do not surpass what the cell can accommodate with its limited growth potential. This issue has not yet been addressed in eukaryotes. In this report, we demonstrate that the prototypical transactivator Gal4p is regulated by two separate signals represented by the specific inducer galactose and the RNA polymerase II (Pol II) holoenzyme-associated cyclin-dependent kinase Srb10p/ CDK8. These observations suggest a mechanism whereby responses to a specific inducer can be coordinated with the physiological environment.Gal4p regulates expression of the yeast GAL genes in response to galactose. In noninducing conditions, Gal4p is bound to the upstream activating sequences for galactose (UAS G ) but is prevented from activating transcription by the inhibitor Gal80p (32,43). Rapid induction by galactose requires the product of GAL3 (40, 52, 57), which is a regulatory protein with similarity to the galactokinase encoded by GAL1 (3, 9, 50), although Gal3p does not have galactokinase activity (9). Recent experiments demonstrate that Gal3p, when bound to galactose, directly interacts with Gal80p in the presence of ATP (42,50,59,60). Gal3p-galactose is thought to cause induction of the GAL genes by producing a conformational change in the Gal4p-Gal80p complex that allows interaction of the Gal4p activating domains with the general transcription factors (42, 59). The induced conformation may ...

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“…Yeast cells utilize galactose by rapidly activating expression of GAL genes (25,26,32,45). Transcription of GAL genes is dependent on the DNA-binding activator Gal4p (18,21,27,31).…”

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Evidence for Gal3p's Cytoplasmic Location and Gal80p's Dual Cytoplasmic-Nuclear Location Implicates New Mechanisms for Controlling Gal4p Activity in Saccharomyces cerevisiae

Peng¹,

Hopper

2000

Molecular and Cellular Biology

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Genetics and in vitro studies have shown that the direct interaction between Gal3p and Gal80p plays a central role in galactose-dependent Gal4p-mediated GAL gene expression in the yeast Saccharomyces cerevisiae. Precisely how Gal3p-Gal80p interaction effects induction is not clear. It has been assumed that Gal3p interacts with Gal80p in the nucleus upon galactose addition to release Gal80p inhibition of Gal4p. Although Gal80p has been shown to possess nuclear localization signal (NLS) peptides, the subcellular distribution of neither Gal80p nor Gal3p was previously determined. Here we report that Gal3p is located in the cytoplasm and apparently excluded from the nucleus. We show that Gal80p is located in both the cytoplasm and the nucleus. Converting Gal80p into a nucleus-localized protein (NLS-Gal80p) by exogenous NLS addition impairs GAL gene induction. The impaired induction can be partially suppressed by targeting Gal3p to the nucleus (NLS-Gal3p). We document a very rapid association between NLS-Gal3p and Gal80p in vivo in response to galactose, illustrating that the nuclear import of Gal80p is very rapid and efficient. We also demonstrate that nucleus-localized NLS-Gal80p can move out of the nucleus and shuttle between nuclei in yeast heterokaryons. These results are the first indication that the subcellular distribution dynamics of the Gal3 and Gal80 proteins play a role in regulating Gal4p-mediated GAL gene expression in vivo.

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Signaling activation and repression of RNA polymerase II transcription in yeast

Cited by 33 publications

References 70 publications

The Galactose Switch in Kluyveromyces lactis Depends on Nuclear Competition between Gal4 and Gal1 for Gal80 Binding

The Galactose Switch in Kluyveromyces lactis Depends on Nuclear Competition between Gal4 and Gal1 for Gal80 Binding

Multiple Signals Regulate GALTranscription in Yeast

Evidence for Gal3p's Cytoplasmic Location and Gal80p's Dual Cytoplasmic-Nuclear Location Implicates New Mechanisms for Controlling Gal4p Activity in Saccharomyces cerevisiae

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