Saccharomyces cerevisiae RNA Polymerase II Is Affected by Kluyveromyces lactis Zymocin

Jablonowski, Daniel; Schaffrath, Raffael

doi:10.1074/jbc.m203354200

Cited by 16 publications

(12 citation statements)

References 47 publications

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“…In all cases, the mutant cells exhibited zymocin hypersensitivity. Moreover, hypersensitivity was also caused by truncation of the RNAPII CTD itself, further supporting the idea that a functional link between zymocin and RNAPII exists (13).…”

supporting

confidence: 53%

“…In apparent agreement with this idea, low resolution hybridization showed that global poly(A) ϩ mRNA levels generally decline in the presence of zymocin, and Northern blot analysis showed significantly reduced levels of specific RNAPII-generated transcripts (3,5). In an attempt to further reinforce the notion that zymocin action is linked to RNAPII, Jablonowski and Schaffrath studied the effects on zymocin toxicity of genetic conditions that would be supposed to directly impair polymerase activity, such as mutations in the RNAPII kinase encoded by the BUR1/BUR2 genes, deletion of the SRB10 CTD kinase gene, and inactivation of the kinase activity of TFIIH (13). In all cases, the mutant cells exhibited zymocin hypersensitivity.…”

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confidence: 99%

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Physical and Functional Interaction between Elongator and the Chromatin-associated Kti12 Protein

Petrakis

Soegaard

Erdjument‐Bromage

et al. 2005

Journal of Biological Chemistry

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Cells lacking KTI12 or Elongator (ELP) genes are insensitive to the toxin zymocin and also share more general phenotypes. Moreover, data from low stringency immunoprecipitation experiments suggest that Elongator and Kti12 may interact. However, the precise relationship between these factors has not been determined. Here we use a variety of approaches to investigate the possibility that Elongator and Kti12 functionally overlap. Native Kti12 purified to virtual homogeneity under stringent conditions is a single polypeptide, but depletion of Kti12 from a yeast extract results in co-depletion of Elongator, indicating that these factors do interact. Indeed, biochemical evidence suggests that Elongator and Kti12 form a fragile complex under physiological salt conditions. Purified Kti12 does not affect Elongator histone acetyltransferase activity in vitro. However, a variety of genetic experiments comparing the effects of mutation in ELP3 and KTI12 alone and in combination with other transcription factor mutations clearly demonstrate a significant functional overlap between Elongator and Kti12 in vivo. Intriguingly, chromatin immunoprecipitation experiments show that Kti12 is associated with chromatin throughout the genome, even in non-transcribed regions and in the absence of Elongator. Conversely, RNA-immunoprecipitation experiments indicate that Kti12 only plays a minor role for Elongator association with active genes. Together, these experiments indicate a close physical and functional relationship between Elongator and the highly conserved Kti12 protein.

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supporting

confidence: 53%

mentioning

confidence: 99%

Physical and Functional Interaction between Elongator and the Chromatin-associated Kti12 Protein

Petrakis

Soegaard

Erdjument‐Bromage

et al. 2005

Journal of Biological Chemistry

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“…Instead, its function may be modified in such a way that pol II activity is poisoned. Data indicating down-regulation of pol II-dependent transcription in the presence of zymocin, expression of partial zymocin resistance on overproduction of Fcp1p -a phosphatase that acts on the C-terminal domain (CTD) of the largest subunit of pol II -and possible interference of zymocin with recycling of pol II all support this notion (Jablonowski et al 2001b;Jablonowski and Schaffrath 2002).…”

Section: Introductionmentioning

confidence: 60%

Defects in yeast RNA polymerase II transcription elicit hypersensitivity to G1 arrest induced by Kluyveromyces lactis zymocin

Jablonowski

Nagase

et al. 2002

Mol Gen Genomics

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The G1 cell cycle arrest imposed by Kluyveromyces lactis zymocin on Saccharomyces cerevisiae requires a functional RNA polymerase II (pol II) TOT/Elongator complex. In a study of zymocin's mode of action, genetic scenarios known to impair transcription or affect the pol II machinery itself were found to elicit hypersensitivity to zymocin. Thus, mutations in components of SAGA, SWI/SNF, Mediator and Ccr4-Not, complexes involved in transcriptionally relevant functions such as nucleosome modification, chromatin remodelling and formation of the preinitiation complex, make yeast cells hypersensitive to the lethal effects of zymocin. The defects at the level of transcriptional elongation displayed by rtf1Delta, ctk1, fcp1 and rpb2 mutants also result in zymocin hypersensitivity. Intriguingly, inactivation of histone deacetylase (HDAC) activity, which is expected to reduce the demand for the histone acetyltransferase (HAT) function of TOT/Elongator, also reduces sensitivity to zymocin. Thus, zymocin interferes with pol II-dependent transcription, and this effect requires the HAT function of TOT, presumably while the Elongator complex is associated with pol II.

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“…Together with the finding that the TOT function can be dissociated from Elongator by mutagenesis of its HAT gene without affecting other Elongator properties (9), the HAT function of Elongator plays a key role in mediating zymocicity. As judged from the observations that (i) pol II-driven transcription is down-regulated in zymocin-treated cells (8,9), (ii) pol II underassembly and general pol II defects elicit zymocin-hypersensitivity (9,28), (iii) interfering with pol II C-terminal domain (CTD) modification alters the response of a cell to zymocin (9), and (iv) the phospho-states of pol II are imbalanced in zymocin-treated cells (29), zymocin may work by hijacking the TOT function of Elongator to convert it into a global pol II inhibitor. Genetic analyses have shown that deletion of any one of the ELP/TOT genes phenocopies the full range of elp/tot phenotypes induced by elp3 point mutations that drastically reduce the HAT activity of Elongator (8,12,25).…”

mentioning

confidence: 99%

Subunit Communications Crucial for the Functional Integrity of the Yeast RNA Polymerase II Elongator (γ-Toxin Target (TOT)) Complex

Frohloff

Jablonowski

Fichtner

et al. 2003

Journal of Biological Chemistry

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In response to the Kluyveromyces lactis zymocin, the ␥-toxin target (TOT) function of the Saccharomyces cerevisiae RNA polymerase II (pol II) Elongator complex prevents sensitive strains from cell cycle progression. Studying Elongator subunit communications, Tot1p (Elp1p), the yeast homologue of human IKK-associated protein, was found to be essentially involved in maintaining the structural integrity of Elongator. Thus, the ability of Tot2p (Elp2p) to interact with the HAT subunit Tot3p (Elp3p) of Elongator and with subunit Tot5p (Elp5p) is dependent on Tot1p (Elp1p). Also, the association of core-Elongator (Tot1-3p/Elp1-3p) with HAP (Elp4 -6p/Tot5-7p), the second three-subunit subcomplex of Elongator, was found to be sensitive to loss of TOT1 (ELP1) gene function. Structural integrity of the HAP complex itself requires the ELP4/TOT7, ELP5/ TOT5, and ELP6/TOT6 genes, and elp6⌬/tot6⌬ as well as elp4⌬/tot7⌬ cells can no longer promote interaction between Tot5p (Elp5p) and Tot2p (Elp2p). The association between Elongator and Tot4p (Kti12p), a factor that may modulate the TOT activity of Elongator, requires Tot1-3p (Elp1-3p) and Tot5p (Elp5p), indicating that this contact requires a preassembled holo-Elongator complex. Tot4p also binds pol II hyperphosphorylated at its C-terminal domain Ser 5 raising the possibility that Tot4p bridges the contact between Elongator and pol II.Microbial rivalry between Kluyveromyces lactis killer strains and sensitive Saccharomyces cerevisiae cells relies on secretion of zymocin, a heterotrimeric (␣␤␥) protein toxin complex that acts as a cell cycle blocker in G 1 (1-3). Zymocin docking involves interaction of its ␣-subunit, an exo-chitinase, with S. cerevisiae cell wall chitin (4, 5), and anti-zymotic activity resides within the ␥-subunit, also termed ␥-toxin (6, 7). In an effort to identify the intracellular ␥-toxin target (TOT) 1 process, seven TOT genes were found to abrogate toxicity when mutated (8, 9). TOT1-3 and TOT5-7 are identical with ELP1-6 coding for Elongator, an RNA polymerase II (pol II)-associated histone acetyltransferase (HAT) complex (8 -15). In addition, loss of KTI11, KTI13, and SIT4 results in tot phenotypes characteristic for TOT mutants, implying that these genes may also be linked to Elongator function (16 -18). Tot4p (Kti12p) can be found promoter-associated, and it contacts Elongator and pol II (8,19,20). Since both removal and overproduction of Tot4p induce zymocin resistance, Tot4p is likely to influence Elongator by modulating its TOT activity (16,20). Holo-Elongator contains the core complex, Elp1-3p (Tot1-3p), and HAP, a second heterotrimer composed of Elp4 -6p (HapI-3p/Tot5-7p) (8, 9, 13-15). Elp1p is the largest subunit within core-Elongator and homologous to human IKK-associated protein, an IB kinase scaffold protein and a member of a five-subunit protein complex (21, 22). Elp2p is a WD40 protein homologous to murine STAT3-interacting protein StIP1 (12,23,24), and Elp3p is the HAT subunit (11, 25) whose activity requires the HAP complex (26). Consis...

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Saccharomyces cerevisiae RNA Polymerase II Is Affected by Kluyveromyces lactis Zymocin

Cited by 16 publications

References 47 publications

Physical and Functional Interaction between Elongator and the Chromatin-associated Kti12 Protein

Physical and Functional Interaction between Elongator and the Chromatin-associated Kti12 Protein

Defects in yeast RNA polymerase II transcription elicit hypersensitivity to G1 arrest induced by Kluyveromyces lactis zymocin

Subunit Communications Crucial for the Functional Integrity of the Yeast RNA Polymerase II Elongator (γ-Toxin Target (TOT)) Complex

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