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Multistep phosphorylation systems: tunable components of biological signaling circuits.

Valk E, Venta R, Ord M, Faustova I, Kõivomägi M, Loog M - Mol. Biol. Cell (2014)

Bottom Line: According to our model, linear patterns of phosphorylation along disordered protein segments determine the signal-response function of a multisite phosphorylation switch.Here we discuss the general advantages and engineering principles of multisite phosphorylation networks as processors of kinase signals.We also address the idea of using the mechanistic logic of linear multisite phosphorylation networks to design circuits for synthetic biology applications.

View Article: PubMed Central - PubMed

Affiliation: Institute of Technology, University of Tartu, 50411 Tartu, Estonia.

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Multisite phosphorylation of Cdk1 targets. (A) A schematic model of a cyclin-Cdk1-Cks1 complex containing the catalytic Cdk1 subunit, the positive regulatory subunit (cyclin), and an accessory phosphate-binding subunit (Cks1). A disordered substrate is shown together with the docking motifs that interact with each of the subunits. These are as follows: the phosphorylated consensus site (S/T-P) interacts with the Cdk1 active site (red marks with -OH), a cyclin docking motif interacts with a hydrophobic pocket on the cyclin (blue boxes), and a phosphorylated threonine on the substrate interacts with Cks1 (red marks with –P). (B) Examples of multisite phosphorylation networks in selected Cdk1 targets of Saccharomyces cerevisiae. There are at least two types of cyclin-docking motifs in Cdk1 substrates. L-P rich motifs are specific docking sites for the G1-specific Cln2-Cdk1 complex (Bhaduri and Pryciak, 2011; Koivomagi et al., 2011b). RXL motifs are docking motifs specific for the S-phase complex Clb5-Cdk1. Phosphodegrons are sequence motifs composed of two phosphorylated sites positioned 2–3 amino acids apart and recognized by the ubiquitin ligase SCF-Cdc4. Networks are mostly located in disordered regions of the proteins. (C) Graph depicting the ultrasensitive signal-response curves created by multisite phosphorylation. For the model simulation, we created a system of ordinary differential equations describing the phosphorylation of a set of kinase substrates carrying different multisite phosphorylation networks. The differently colored lines indicate simulated phosphorylation output responses at different thresholds of the increasing input signal of the kinase (the diagonal dotted line). The number of phosphorylation sites was varied in each substrate to gain temporal resolution of the response curves (1, 4, 6, 9, and 12 phosphorylation sites in substrates represented by curves from left to right, respectively). In addition, the phosphorylation rate constants of selected steps were varied in some substrates to imitate the effect of different patterning of the sites on the phosphorylation rates on individual steps. Weak phosphatase activity was included to counteract the kinase activity. The dotted horizontal lines represent the kinase thresholds at which half of the substrate has reached the fully phosphorylated state.
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Figure 1: Multisite phosphorylation of Cdk1 targets. (A) A schematic model of a cyclin-Cdk1-Cks1 complex containing the catalytic Cdk1 subunit, the positive regulatory subunit (cyclin), and an accessory phosphate-binding subunit (Cks1). A disordered substrate is shown together with the docking motifs that interact with each of the subunits. These are as follows: the phosphorylated consensus site (S/T-P) interacts with the Cdk1 active site (red marks with -OH), a cyclin docking motif interacts with a hydrophobic pocket on the cyclin (blue boxes), and a phosphorylated threonine on the substrate interacts with Cks1 (red marks with –P). (B) Examples of multisite phosphorylation networks in selected Cdk1 targets of Saccharomyces cerevisiae. There are at least two types of cyclin-docking motifs in Cdk1 substrates. L-P rich motifs are specific docking sites for the G1-specific Cln2-Cdk1 complex (Bhaduri and Pryciak, 2011; Koivomagi et al., 2011b). RXL motifs are docking motifs specific for the S-phase complex Clb5-Cdk1. Phosphodegrons are sequence motifs composed of two phosphorylated sites positioned 2–3 amino acids apart and recognized by the ubiquitin ligase SCF-Cdc4. Networks are mostly located in disordered regions of the proteins. (C) Graph depicting the ultrasensitive signal-response curves created by multisite phosphorylation. For the model simulation, we created a system of ordinary differential equations describing the phosphorylation of a set of kinase substrates carrying different multisite phosphorylation networks. The differently colored lines indicate simulated phosphorylation output responses at different thresholds of the increasing input signal of the kinase (the diagonal dotted line). The number of phosphorylation sites was varied in each substrate to gain temporal resolution of the response curves (1, 4, 6, 9, and 12 phosphorylation sites in substrates represented by curves from left to right, respectively). In addition, the phosphorylation rate constants of selected steps were varied in some substrates to imitate the effect of different patterning of the sites on the phosphorylation rates on individual steps. Weak phosphatase activity was included to counteract the kinase activity. The dotted horizontal lines represent the kinase thresholds at which half of the substrate has reached the fully phosphorylated state.

Mentions: Cdk1-dependent phosphorylation events often lead to the generation of phosphorylated sequence motifs (phosphodegrons) that are recognized by the ubiquitination machinery and thereby marked for destruction (Hao et al., 2007; Koivomagi et al., 2011a; Landry et al., 2012). For example, phosphorylation-dependent destruction of a Cdk1 inhibitor protein called Sic1 helps to trigger S phase in budding yeast. Cdk1, when bound to G1- and S-phase cyclins, phosphorylates Sic1 in an ordered sequence at multiple sites, leading to the formation of phosphodegrons that are recognized by the ubiquitin ligase SCF-Cdc4. The sequential phosphorylation of Sic1 and other substrates depends on three important interactions between Cdk1 complexes and the disordered substrate chain (Figure 1, A and B): the active site of Cdk1 interacts with the consensus phosphorylation site, typically S/T-P or S/T-PxK/R (Khoury et al., 2011); specific sites on the cyclin interact with docking motifs on the substrate (Holt et al., 2009); and the small adaptor protein Cks1 interacts with specific phosphorylated threonines on the substrate (Tyanova et al., 2013).


Multistep phosphorylation systems: tunable components of biological signaling circuits.

Valk E, Venta R, Ord M, Faustova I, Kõivomägi M, Loog M - Mol. Biol. Cell (2014)

Multisite phosphorylation of Cdk1 targets. (A) A schematic model of a cyclin-Cdk1-Cks1 complex containing the catalytic Cdk1 subunit, the positive regulatory subunit (cyclin), and an accessory phosphate-binding subunit (Cks1). A disordered substrate is shown together with the docking motifs that interact with each of the subunits. These are as follows: the phosphorylated consensus site (S/T-P) interacts with the Cdk1 active site (red marks with -OH), a cyclin docking motif interacts with a hydrophobic pocket on the cyclin (blue boxes), and a phosphorylated threonine on the substrate interacts with Cks1 (red marks with –P). (B) Examples of multisite phosphorylation networks in selected Cdk1 targets of Saccharomyces cerevisiae. There are at least two types of cyclin-docking motifs in Cdk1 substrates. L-P rich motifs are specific docking sites for the G1-specific Cln2-Cdk1 complex (Bhaduri and Pryciak, 2011; Koivomagi et al., 2011b). RXL motifs are docking motifs specific for the S-phase complex Clb5-Cdk1. Phosphodegrons are sequence motifs composed of two phosphorylated sites positioned 2–3 amino acids apart and recognized by the ubiquitin ligase SCF-Cdc4. Networks are mostly located in disordered regions of the proteins. (C) Graph depicting the ultrasensitive signal-response curves created by multisite phosphorylation. For the model simulation, we created a system of ordinary differential equations describing the phosphorylation of a set of kinase substrates carrying different multisite phosphorylation networks. The differently colored lines indicate simulated phosphorylation output responses at different thresholds of the increasing input signal of the kinase (the diagonal dotted line). The number of phosphorylation sites was varied in each substrate to gain temporal resolution of the response curves (1, 4, 6, 9, and 12 phosphorylation sites in substrates represented by curves from left to right, respectively). In addition, the phosphorylation rate constants of selected steps were varied in some substrates to imitate the effect of different patterning of the sites on the phosphorylation rates on individual steps. Weak phosphatase activity was included to counteract the kinase activity. The dotted horizontal lines represent the kinase thresholds at which half of the substrate has reached the fully phosphorylated state.
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Figure 1: Multisite phosphorylation of Cdk1 targets. (A) A schematic model of a cyclin-Cdk1-Cks1 complex containing the catalytic Cdk1 subunit, the positive regulatory subunit (cyclin), and an accessory phosphate-binding subunit (Cks1). A disordered substrate is shown together with the docking motifs that interact with each of the subunits. These are as follows: the phosphorylated consensus site (S/T-P) interacts with the Cdk1 active site (red marks with -OH), a cyclin docking motif interacts with a hydrophobic pocket on the cyclin (blue boxes), and a phosphorylated threonine on the substrate interacts with Cks1 (red marks with –P). (B) Examples of multisite phosphorylation networks in selected Cdk1 targets of Saccharomyces cerevisiae. There are at least two types of cyclin-docking motifs in Cdk1 substrates. L-P rich motifs are specific docking sites for the G1-specific Cln2-Cdk1 complex (Bhaduri and Pryciak, 2011; Koivomagi et al., 2011b). RXL motifs are docking motifs specific for the S-phase complex Clb5-Cdk1. Phosphodegrons are sequence motifs composed of two phosphorylated sites positioned 2–3 amino acids apart and recognized by the ubiquitin ligase SCF-Cdc4. Networks are mostly located in disordered regions of the proteins. (C) Graph depicting the ultrasensitive signal-response curves created by multisite phosphorylation. For the model simulation, we created a system of ordinary differential equations describing the phosphorylation of a set of kinase substrates carrying different multisite phosphorylation networks. The differently colored lines indicate simulated phosphorylation output responses at different thresholds of the increasing input signal of the kinase (the diagonal dotted line). The number of phosphorylation sites was varied in each substrate to gain temporal resolution of the response curves (1, 4, 6, 9, and 12 phosphorylation sites in substrates represented by curves from left to right, respectively). In addition, the phosphorylation rate constants of selected steps were varied in some substrates to imitate the effect of different patterning of the sites on the phosphorylation rates on individual steps. Weak phosphatase activity was included to counteract the kinase activity. The dotted horizontal lines represent the kinase thresholds at which half of the substrate has reached the fully phosphorylated state.
Mentions: Cdk1-dependent phosphorylation events often lead to the generation of phosphorylated sequence motifs (phosphodegrons) that are recognized by the ubiquitination machinery and thereby marked for destruction (Hao et al., 2007; Koivomagi et al., 2011a; Landry et al., 2012). For example, phosphorylation-dependent destruction of a Cdk1 inhibitor protein called Sic1 helps to trigger S phase in budding yeast. Cdk1, when bound to G1- and S-phase cyclins, phosphorylates Sic1 in an ordered sequence at multiple sites, leading to the formation of phosphodegrons that are recognized by the ubiquitin ligase SCF-Cdc4. The sequential phosphorylation of Sic1 and other substrates depends on three important interactions between Cdk1 complexes and the disordered substrate chain (Figure 1, A and B): the active site of Cdk1 interacts with the consensus phosphorylation site, typically S/T-P or S/T-PxK/R (Khoury et al., 2011); specific sites on the cyclin interact with docking motifs on the substrate (Holt et al., 2009); and the small adaptor protein Cks1 interacts with specific phosphorylated threonines on the substrate (Tyanova et al., 2013).

Bottom Line: According to our model, linear patterns of phosphorylation along disordered protein segments determine the signal-response function of a multisite phosphorylation switch.Here we discuss the general advantages and engineering principles of multisite phosphorylation networks as processors of kinase signals.We also address the idea of using the mechanistic logic of linear multisite phosphorylation networks to design circuits for synthetic biology applications.

View Article: PubMed Central - PubMed

Affiliation: Institute of Technology, University of Tartu, 50411 Tartu, Estonia.

Show MeSH
Related in: MedlinePlus