Clb6/Cdc28 and Cdc14 Regulate Phosphorylation Status and Cellular Localization of Swi6

Geymonat, Marco; Spanos, Ad; Wells, Glenn; Smerdon, Stephen J.; Sedgwick, Steven G.

doi:10.1128/mcb.24.6.2277-2285.2004

Cited by 65 publications

(82 citation statements)

References 62 publications

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“…The Clb6/Cdc28 S-phase cell cycle kinase is responsible for phosphorylation of Swi6 on Ser160 (Geymonat et al 2004). Swi6 resides predominantly in the cytoplasm from late G1 until late M phase, at which time it relocalizes to the nucleus in response to dephosphorylation at Ser160, where it remains for most of G1.…”

Section: Control Of Swi6 Nucleocytoplasmic Shuttling By Mpk1mentioning

confidence: 99%

Regulation of Cell Wall Biogenesis in Saccharomyces cerevisiae: The Cell Wall Integrity Signaling Pathway

2011

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The yeast cell wall is a strong, but elastic, structure that is essential not only for the maintenance of cell shape and integrity, but also for progression through the cell cycle. During growth and morphogenesis, and in response to environmental challenges, the cell wall is remodeled in a highly regulated and polarized manner, a process that is principally under the control of the cell wall integrity (CWI) signaling pathway. This pathway transmits wall stress signals from the cell surface to the Rho1 GTPase, which mobilizes a physiologic response through a variety of effectors. Activation of CWI signaling regulates the production of various carbohydrate polymers of the cell wall, as well as their polarized delivery to the site of cell wall remodeling. This review article centers on CWI signaling in Saccharomyces cerevisiae through the cell cycle and in response to cell wall stress. The interface of this signaling pathway with other pathways that contribute to the maintenance of cell wall integrity is also discussed.

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Section: Control Of Swi6 Nucleocytoplasmic Shuttling By Mpk1mentioning

confidence: 99%

Regulation of Cell Wall Biogenesis in Saccharomyces cerevisiae: The Cell Wall Integrity Signaling Pathway

2011

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“…The pathway(s) responsible for cdc14 replication insufficiency can be potentially revealed through the identification of Cdc14-dependent events in the previous cell cycle. Incidentally, dephosphorylation of Swi6p (Ser-160) by Cdc14 is needed for its nuclear import in telophase (33) and formation of the SBF and MBF transciption complexes (34), which activate the G1 expression of a wide range of genes essential for S phase. Indeed, we found that several key SWI6-controlled genes involved in DNA replication and other processes showed reproducibly lower transcript levels in cdc14 in G1 (at 23°C), as well as 10 and 20 min into S phase (at 37°C) (Fig.…”

Section: G1 Transcription and Nuclear Import Defects In Cdc14 Impairmentioning

confidence: 99%

Essential global role of CDC14 in DNA synthesis revealed by chromosome underreplication unrecognized by checkpoints in cdc14 mutants

Dulev

Renty

Mehta

et al. 2009

Proc. Natl. Acad. Sci. U.S.A.

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The CDC14 family of multifunctional evolutionarily conserved phosphatases includes major regulators of mitosis in eukaryotes and of DNA damage response in humans. The CDC14 function is also crucial for accurate chromosome segregation, which is exemplified by its absolute requirement in yeast for the anaphase segregation of nucleolar organizers; however the nature of this essential pathway is not understood. Upon investigation of the rDNA nondisjunction phenomenon, it was found that cdc14 mutants fail to complete replication of this locus. Moreover, other late-replicating genomic regions (10% of the genome) are also underreplicated in cdc14 mutants undergoing anaphase. This selective genome-wide replication defect is due to dosage insufficiency of replication factors in the nucleus, which stems from two defects, both contingent on the reduced CDC14 function in the preceding mitosis. First, a constitutive nuclear import defect results in a drastic dosage decrease for those replication proteins that are regulated by nuclear transport. Particularly, essential RPA subunits display both lower mRNA and protein levels, as well as abnormal cytoplasmic localization. Second, the reduced transcription of MBF and SBF-controlled genes in G1 leads to the reduction in protein levels of many proteins involved in DNA replication. The failure to complete replication of late replicons is the primary reason for chromosome nondisjunction upon CDC14 dysfunction. As the genome-wide slow-down of DNA replication does not trigger checkpoints [Lengronne A, Schwob E (2002) Mol Cell 9:1067-1078], CDC14 mutations pose an overwhelming challenge to genome stability, both generating chromosome damage and undermining the checkpoint control mechanisms.S uccessful chromosome segregation in mitosis requires the coordinated work of cellular regulatory circuits, which transmit cell cycle signals to chromatin and spindle proteins. One of such factors is the Cdc14 protein, an evolutionary conserved protein phosphatase (1-3). While in human cells a CDC14 ortholog has been recently shown to play a key role in DNA damage response (4), studies on S. cerevisiae were mostly focused on Cdc14p roles in anaphase regulation and in the exit from mitosis. The scope of Cdc14p activity in budding yeast is believed to be limited to anaphase, because Cdc14p is sequestered in the nucleolus (5) in apparently inactive form (6) at other cell cycle stages. Therefore, while Cdc14 can potentially dephosphorylate many substrates (7,8), the most studied physiological pathways are the anaphase pathways (FEAR and MEN), which are both dependent on the two sequential bursts of Cdc14 release (1, 9). The cdc14 mutations cause a mitotic exit block, but also display defects in nucleolar (10) and telomeric (11) segregation. The mechanisms of chromosome segregation defects (11-15) in cdc14 mutants are generally poorly understood. While condensin mutations phenocopy the cdc14 rDNA nondisjunction (11, 16) and Cdc14p is required for condensin loading to rDNA (14), it is unlikely that conden...

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“…In the case of the early eluting peptide, we were unable to distinguish between phosphorylation at any of the first three residues. Because it is less likely that trypsin would efficiently cleave between Arg 3 and Thr 4 , we conclude that this sequence is most likely phosphorylated at Thr 5 . Phosphorylation at Ser 243 was found in several peptides due to the presence of multiple lysine and arginine residues at either end of the sequence surrounding it.…”

Section: Phosphate-dependent Phosphorylation Profile Of Pho4mentioning

confidence: 99%

“…It can serve as a recognition element, targeting a substrate for ubiquitin-dependent proteolysis (2), or serve as a docking site to recruit other proteins into multiprotein complexes (3). Phosphorylation can trigger a change in the three-dimensional structure of a protein or initiate translocation of the protein to another compartment of the cell (4). Disruption of normal cellular phosphorylation events is responsible for a number of human diseases (5).…”

mentioning

confidence: 99%

A Quantitative Results-driven Approach to Analyzing Multisite Protein Phosphorylation

Zappacosta

Collingwood

Huddleston

et al. 2006

Molecular & Cellular Proteomics

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Multisite protein phosphorylation appears to be quite common. Nevertheless our understanding of how multiple phosphorylation events regulate the function of a protein is limited in many cases. The ability to measure temporal changes in the site-specific phosphorylation profile of a protein in response to a given stimulus or cellular activity would provide an immediate indication of the functional significance of any phosphorylation site to a given process. Here we describe a mass spectrometry-based method to identify functionally relevant phosphorylation sites on a protein. It combines stable isotope labeling with a highly selective mass spectrometry analysis to detect and quantitate phosphorylation sites in response to a cellular signal. This approach requires no a priori knowledge of the phosphorylation state of the protein, does not require purification of phosphopeptides, and reliably detects substoichiometric levels of phosphorylation. Following a review of the quantitative results, only those phosphorylation sites that show a change in relative abundance are selected for identification and further study. We used this results-driven approach to study phosphorylation of the budding yeast transcription factor Pho4 in response to phosphate starvation. Phosphorylation of Pho4 on five cyclin-dependent kinase (Cdk) consensus sites has been shown to regulate the transcriptional activity of Pho4 in response to changes in environmental phosphate levels. Here we show that in phosphate-rich medium Pho4 is phosphorylated on at least 15 distinct sites including the five Cdk sites described previously. In excellent agreement with the known mechanism for regulation of Pho4 we found that phosphorylation at all five of the Cdk sites was repressed in phosphate-depleted medium. In addition to these five sites, we identified four novel phosphorylation sites that were also responsive to changes in phosphate availability. Selecting a limited number of Pho4 phosphorylation sites, we performed a more detailed kinetic analysis using an isotope-free strategy. We used LC-MS with selected reaction monitoring to greatly improve the accuracy, sensitivity, and dynamic range of the subsequent experiments. A detailed analysis of the cell-based phosphorylation at the selected Pho4 sites confirmed an apparent site preference for the Pho80-Pho85 cyclin-cyclindependent kinase complex.

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Clb6/Cdc28 and Cdc14 Regulate Phosphorylation Status and Cellular Localization of Swi6

Cited by 65 publications

References 62 publications

Regulation of Cell Wall Biogenesis in Saccharomyces cerevisiae: The Cell Wall Integrity Signaling Pathway

Regulation of Cell Wall Biogenesis in Saccharomyces cerevisiae: The Cell Wall Integrity Signaling Pathway

Essential global role of CDC14 in DNA synthesis revealed by chromosome underreplication unrecognized by checkpoints in cdc14 mutants

A Quantitative Results-driven Approach to Analyzing Multisite Protein Phosphorylation

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