Multisite phosphorylation networks as signal processors for Cdk1

Kõivomägi, Mardo; Örd, Mihkel; Iofik, Anna; Valk, Ervin; Venta, Rainis; Faustova, Ilona; Kivi, Rait; Balog, Eva Rose M.; Rubin, Seth M.; Loog, Mart

doi:10.1038/nsmb.2706

Cited by 121 publications

(191 citation statements)

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“…In addition, this mechanism allows for greater control by opposing phosphatases and fine tuning of the signal process based on the number of phosphor- ylated sites involved. Cdk1-dependent multisite phosphorylation has been observed in several yeast species and Xenopus laevis suggesting that this mechanism could be conserved in human cells (57,(65)(66)(67). Further investigation of Cdk1-dependent multisite phosphorylation will be important to determine if the underlying mechanisms are also conserved.…”

Section: Discussionmentioning

confidence: 99%

“…3C), we found that 71% contained two or more regulated phosphopeptides and 46% contained three or more, suggesting that Cdk1 regulation of protein function occurs through cumulative multisite phosphorylation events. Multisite phosphorylation of Cdk substrates was previously reported in several yeast species and Xenopus laevis, and proposed to allow for differential regulation of substrate phosphorylation and function (57,(65)(66)(67).…”

Section: Quantitative Analysis Of the Phosphoproteome Of Flavopiridolmentioning

confidence: 99%

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Identification of Candidate Cyclin-dependent kinase 1 (Cdk1) Substrates in Mitosis by Quantitative Phosphoproteomics

Petrone

Adamo

Cheng

et al. 2016

Molecular & Cellular Proteomics

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An important objective of a cell is to accurately replicate its genetic material and evenly divide along with its subcellular components into two identical daughter cells. To do so, a cell reduces or halts growth, transcription, cap-dependent translation, and undergoes dramatic changes in cellular structure and organization. These changes include chromosome condensation, nuclear envelope breakdown, disassembly of the endoplasmic reticulum and Golgi apparatus, reorganization of the actin cortex, and the formation of the mitotic spindle. Deregulation and errors in these processes can produce nonidentical daughter cells with aberrant chromosome numbers, a state known as aneuploidy and a hallmark of human cancer and the origin of many birth defects (1-5). Therefore, it is imperative that mitosis proceeds in a highly accurate and controlled manner. This is achieved by a sophisticated network of proteins that engage in a multitude of protein-protein interactions regulated by post-translational modifications, including dynamic protein phosphorylation by protein kinases and phosphatases (summarized in (6 -8)).One of the master regulators of mitosis that is conserved from yeast to human is the cyclin-dependent kinase 1 (Cdk1) 1 (9). Cdk1 expression is constant across the cell cycle and the regulation of its activity relies on its association with cyclin A and B, as well as on post-translational modifications including phosphorylation. Specifically, mRNA and protein abundance of cyclin A and B oscillate during the cell cycle due to temporally regulated transcription, translation, and degradation cycles that restrict Cdk1 activity from S-phase to mitosis

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Section: Discussionmentioning

confidence: 99%

Section: Quantitative Analysis Of the Phosphoproteome Of Flavopiridolmentioning

confidence: 99%

Identification of Candidate Cyclin-dependent kinase 1 (Cdk1) Substrates in Mitosis by Quantitative Phosphoproteomics

Petrone

Adamo

Cheng

et al. 2016

Molecular & Cellular Proteomics

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“…The metaphase phosphorylation peak is followed by rapid dephosphorylation resulting in Cnn1 enrichment to kinetochores at anaphase onset (Bock et al 2012). A Cdc28-dependent threshold triggers the Skp1-Cul1-F box (SCF)-mediated destruction of Sic1 at S phase entry (Koivomagi et al 2011(Koivomagi et al , 2013 and the Cnn1 metaphase phospho threshold may initiate phosphatase activity on S74 and surrounding residues. S74 from Cnn1 likely needs surrounding sites to be phosphorylated because the Cnn1 60-84 sequence can replace a similar Ndc80 binding motif in the Dsn1 protein, indicating S74 is not phosphorylated in that context (Malvezzi et al 2013).…”

Section: Discussionmentioning

confidence: 99%

The Mps1 Kinase Modulates the Recruitment and Activity of Cnn1CENP-T at Saccharomyces cerevisiae Kinetochores

et al. 2015

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Kinetochores are conserved protein complexes that bind the replicated chromosomes to the mitotic spindle and then direct their segregation. To better comprehend Saccharomyces cerevisiae kinetochore function, we dissected the phospho-regulated dynamic interaction between conserved kinetochore protein Cnn1 CENP-T , the centromere region, and the Ndc80 complex through the cell cycle. Cnn1 localizes to kinetochores at basal levels from G1 through metaphase but accumulates abruptly at anaphase onset. How Cnn1 is recruited and which activities regulate its dynamic localization are unclear. We show that Cnn1 harbors two kinetochore-localization activities: a C-terminal histone-fold domain (HFD) that associates with the centromere region and a N-terminal Spc24/Spc25 interaction sequence that mediates linkage to the microtubule-binding Ndc80 complex. We demonstrate that the established Ndc80 binding site in the N terminus of Cnn1, Cnn1 , should be extended with flanking residues, Cnn1 , to allow near maximal binding affinity to Ndc80. Cnn1 localization was proposed to depend on Mps1 kinase activity at Cnn1-S74, based on in vitro experiments demonstrating the Cnn1-Ndc80 complex interaction. We demonstrate that from G1 through metaphase, Cnn1 localizes via both its HFD and N-terminal Spc24/Spc25 interaction sequence, and deletion or mutation of either region results in anomalous Cnn1 kinetochore levels. At anaphase onset (when Mps1 activity decreases) Cnn1 becomes enriched mainly via the N-terminal Spc24/Spc25 interaction sequence. In sum, we provide the first in vivo evidence of Cnn1 preanaphase linkages with the kinetochore and enrichment of the linkages during anaphase. KEYWORDS Cnn1; Mps1; kinetochore; centromere; CENP-T K INETOCHORES are large protein structures that assemble hierarchically on the centromeres (CEN) of replicated chromosomes (sister chromatids). They biorient each sister chromatid pair to the microtubules (MTs) of the mitotic spindle and orchestrate chromatid segregation into the daughter cells (Cheeseman 2014;Malvezzi and Westermann 2014). At the core of each kinetochore lies a protein network named KMN (KLN1/Spc105, MIS12/Mtw1, and NDC80/Ndc80 complexes) that bridges the CEN and MTs (Cheeseman et al. 2006;Westermann et al. 2007). The Ndc80 complex attaches kinetochores to the MTs via its outer Ndc80/Nuf2 dimer (Wei et al. 2007;Ciferri et al. 2008;Alushin et al. 2010), while its CEN-proximal Spc24/Spc25 dimer interacts with a putatively CEN-associated protein, known as Cnn1 in budding yeast (CENP-T in metazoans). The N-terminal domain of Cnn1 and CENP-T hook onto the interface of the Spc24/Spc25 dimer (Malvezzi et al. 2013;Nishino et al. 2013). In its C terminus, Cnn1 harbors a histonefold domain (HFD) (Schleiffer et al. 2012), which may associate with CEN DNA, as does CENP-T (Hori et al. 2008;Nishino et al. 2012).Cnn1 levels at kinetochores are low from G1 through metaphase but increase two-to threefold at anaphase entry and drop back to base level at anaphase exit. Cnn1 interacts with the...

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“…For assignment, approximately 0.7 mM 15 N, 13 C Ash1 sample was used to acquire standard triple-resonance backbone assignment experiments and carbon-detect triple resonance experiments. Standard assignment experiments were based on sensitivity enhanced 1 H- 15 65 Additionally, CCCON-IPAP, a 3D 13 C TOCSY, was used to distinguish trans versus cis proline (32 scans, 1024 ( 13 C) × 48 ( 15 N) × 160 ( 13 C) complex data points, with 18 ppm, 36 ppm and 72 ppm as 13 C, 15 N and 13 C (TOCSY) sweep widths). 66 15 N NMR relaxation experiments acquired on a Bruker Avance 800 MHz spectrometer at 278 K using standard pulse programs (16 scans, 2048 ( 1 H) × 150 ( 15 N) complex data points).…”

Section: Saxs Data Analysismentioning

confidence: 99%

“…1 Multisite phosphorylation is dynamic 2 and it enables rapid signal integration [3][4] . Many nonlinear downstream responses [5][6][7][8][9][10] such as transcriptional regulation [11][12][13] , cell cycle control [4][5][14][15] , and cell proliferation 16 are coordinated by multisite phosphorylation of IDRs. Archetypal IDRs that undergo multisite phosphorylation include the C-terminal domain of RNA polymerase II 13 , the C-terminal tail of the epidermal growth factor receptor 13 , and sidearms of intermediate filaments in neurons 17 .…”

Section: Introductionmentioning

confidence: 99%

Sequence Determinants of the Conformational Properties of an Intrinsically Disordered Protein Prior to and Upon Multisite Phosphorylation

Martin

Holehouse

Pappu

et al. 2017

Biophysical Journal

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Many cell signaling events are coordinated by intrinsically disordered protein regions (IDRs) that undergo multisite Serine/Threonine phosphorylation. The conformational properties of these IDRs prior to and following multi-site phosphorylation are directly relevant to understanding their functions. Here, we present results from biophysical studies and molecular simulations that quantify the conformational properties of an 81-residue IDR from the S. cerevisiae transcription factor Ash1. We show that the unphosphorylated Ash1 IDR adopts coil-like conformations that are expanded and well-solvated. This result contradicts inferences regarding global compaction that are derived from heuristics based on amino acid compositions for IDRs with low proline contents. Upon phosphorylation at ten distinct sites, the global conformational properties of pAsh1 are indistinguishable from those of unphosphorylated Ash1. This insensitivity derives from compensatory changes to the pattern of local and long-range intra-chain contacts. We show that the conformational properties of Ash1 and pAsh1 can be explained in terms of the linear sequence patterning of proline and charged residues vis-à-vis all other residues. The sequence features of the Ash1 IDR are shared by many other IDRs that undergo multisite phosphorylation. Accordingly, we propose that our findings might be generalizable to other IDRs involved in cell signaling. TOC image

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Multisite phosphorylation networks as signal processors for Cdk1

Cited by 121 publications

References 58 publications

Identification of Candidate Cyclin-dependent kinase 1 (Cdk1) Substrates in Mitosis by Quantitative Phosphoproteomics

Identification of Candidate Cyclin-dependent kinase 1 (Cdk1) Substrates in Mitosis by Quantitative Phosphoproteomics

The Mps1 Kinase Modulates the Recruitment and Activity of Cnn1CENP-T at Saccharomyces cerevisiae Kinetochores

Sequence Determinants of the Conformational Properties of an Intrinsically Disordered Protein Prior to and Upon Multisite Phosphorylation

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