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Pierre P. Roger - One of the best experts on this subject based on the ideXlab platform.
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cdk4 t172 phosphorylation is central in a cdk7 dependent bidirectional cdk4 cdk2 interplay mediated by p21 phosphorylation at the restriction point
PLOS Genetics, 2013Co-Authors: Xavier Bisteau, Sabine Paternot, Katia Coulonval, Bianca Colleoni, Karin Ecker, Philippe De Groote, Wim Declercq, Ludger Hengst, Pierre P. RogerAbstract:Cell Cycle progression, including genome duplication, is orchestrated by cyclin-dependent kinases (CDKs). CDK activation depends on phosphorylation of their T-loop by a CDK-activating kinase (CAK). In animals, the only known CAK for CDK2 and CDK1 is cyclin H-CDK7, which is constitutively active. Therefore, the critical activation step is dephosphorylation of inhibitory sites by Cdc25 phosphatases rather than unrestricted T-loop phosphorylation. Homologous CDK4 and CDK6 bound to cyclins D are master integrators of mitogenic/oncogenic signaling cascades by initiating the inactivation of the central oncosuppressor pRb and cell Cycle Commitment at the restriction point. Unlike the situation in CDK1 and CDK2 cyclin complexes, and in contrast to the weak but constitutive T177 phosphorylation of CDK6, we have identified the T-loop phosphorylation at T172 as the highly regulated step determining CDK4 activity. Whether both CDK4 and CDK6 phosphorylations are catalyzed by CDK7 remains unclear. To answer this question, we took a chemical-genetics approach by using analogue-sensitive CDK7(as/as) mutant HCT116 cells, in which CDK7 can be specifically inhibited by bulky adenine analogs. Intriguingly, CDK7 inhibition prevented activating phosphorylations of CDK4/6, but for CDK4 this was at least partly dependent on its binding to p21 (cip1) . In response to CDK7 inhibition, p21-binding to CDK4 increased concomitantly with disappearance of the most abundant phosphorylation of p21, which we localized at S130 and found to be catalyzed by both CDK4 and CDK2. The S130A mutation of p21 prevented the activating CDK4 phosphorylation, and inhibition of CDK4/6 and CDK2 impaired phosphorylations of both p21 and p21-bound CDK4. Therefore, specific CDK7 inhibition revealed the following: a crucial but partly indirect CDK7 involvement in phosphorylation/activation of CDK4 and CDK6; existence of CDK4-activating kinase(s) other than CDK7; and novel CDK7-dependent positive feedbacks mediated by p21 phosphorylation by CDK4 and CDK2 to sustain CDK4 activation, pRb inactivation, and restriction point passage.
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Cyclic AMP-dependent Phosphorylation of Cyclin D3-bound CDK4 Determines the Passage through the Cell Cycle Restriction Point in Thyroid Epithelial Cells
The Journal of biological chemistry, 2003Co-Authors: Sabine Paternot, Katia Coulonval, Jacques Emile Dumont, Pierre P. RogerAbstract:According to current concepts, the cell Cycle Commitment after restriction (R) point passage requires the sustained stimulation by mitogens of the synthesis of labile d-type cyclins, which associate with cyclin-dependent kinase (CDK) 4/6 to phosphorylate pRb family proteins and sequester the CDK inhibitor p27kip1. In primary cultures of dog thyroid epithelial cells, the cAMP-dependent cell Cycle induced by a sustained stimulation by thyrotropin or forskolin differs from growth factor mitogenic pathways, as cAMP does not upregulate d-type cyclins but increases p27 levels. Instead, cAMP induces the assembly of required cyclin D3-CDK4 complexes, which associate with nuclear p27. In this study, the arrest of forskolin stimulation rapidly slowed down the entry of dog thyrocytes into S phase and the phosphorylation of pRb family proteins. The pRb kinase activity, but not the formation, of the cyclin D3-CDK4-p27 complex was strongly reduced. Using two-dimensional gel electrophoresis, a phosphorylated form of CDK4 was separated. It appeared in response to forskolin and was bound to both cyclin D3 and p27, presumably reflecting the activating Thr-172 phosphorylation of CDK4. Upon forskolin withdrawal or after cycloheximide addition, this CDK4 phosphoform unexpectedly persisted in p27 complexes devoid of cyclin D3 but it disappeared from the more labile cyclin D3 complexes. These data demonstrate that the assembly of the cyclin D3-CDK4-p27 holoenzyme and the subsequent phosphorylation and activation of CDK4 depend on distinct cAMP actions. This provides a first example of a crucial regulation of CDK4 phosphorylation by a mitogenic cascade and a novel mechanism of cell Cycle control at the R point.
Sabine Paternot - One of the best experts on this subject based on the ideXlab platform.
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cdk4 t172 phosphorylation is central in a cdk7 dependent bidirectional cdk4 cdk2 interplay mediated by p21 phosphorylation at the restriction point
PLOS Genetics, 2013Co-Authors: Xavier Bisteau, Sabine Paternot, Katia Coulonval, Bianca Colleoni, Karin Ecker, Philippe De Groote, Wim Declercq, Ludger Hengst, Pierre P. RogerAbstract:Cell Cycle progression, including genome duplication, is orchestrated by cyclin-dependent kinases (CDKs). CDK activation depends on phosphorylation of their T-loop by a CDK-activating kinase (CAK). In animals, the only known CAK for CDK2 and CDK1 is cyclin H-CDK7, which is constitutively active. Therefore, the critical activation step is dephosphorylation of inhibitory sites by Cdc25 phosphatases rather than unrestricted T-loop phosphorylation. Homologous CDK4 and CDK6 bound to cyclins D are master integrators of mitogenic/oncogenic signaling cascades by initiating the inactivation of the central oncosuppressor pRb and cell Cycle Commitment at the restriction point. Unlike the situation in CDK1 and CDK2 cyclin complexes, and in contrast to the weak but constitutive T177 phosphorylation of CDK6, we have identified the T-loop phosphorylation at T172 as the highly regulated step determining CDK4 activity. Whether both CDK4 and CDK6 phosphorylations are catalyzed by CDK7 remains unclear. To answer this question, we took a chemical-genetics approach by using analogue-sensitive CDK7(as/as) mutant HCT116 cells, in which CDK7 can be specifically inhibited by bulky adenine analogs. Intriguingly, CDK7 inhibition prevented activating phosphorylations of CDK4/6, but for CDK4 this was at least partly dependent on its binding to p21 (cip1) . In response to CDK7 inhibition, p21-binding to CDK4 increased concomitantly with disappearance of the most abundant phosphorylation of p21, which we localized at S130 and found to be catalyzed by both CDK4 and CDK2. The S130A mutation of p21 prevented the activating CDK4 phosphorylation, and inhibition of CDK4/6 and CDK2 impaired phosphorylations of both p21 and p21-bound CDK4. Therefore, specific CDK7 inhibition revealed the following: a crucial but partly indirect CDK7 involvement in phosphorylation/activation of CDK4 and CDK6; existence of CDK4-activating kinase(s) other than CDK7; and novel CDK7-dependent positive feedbacks mediated by p21 phosphorylation by CDK4 and CDK2 to sustain CDK4 activation, pRb inactivation, and restriction point passage.
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Cyclic AMP-dependent Phosphorylation of Cyclin D3-bound CDK4 Determines the Passage through the Cell Cycle Restriction Point in Thyroid Epithelial Cells
The Journal of biological chemistry, 2003Co-Authors: Sabine Paternot, Katia Coulonval, Jacques Emile Dumont, Pierre P. RogerAbstract:According to current concepts, the cell Cycle Commitment after restriction (R) point passage requires the sustained stimulation by mitogens of the synthesis of labile d-type cyclins, which associate with cyclin-dependent kinase (CDK) 4/6 to phosphorylate pRb family proteins and sequester the CDK inhibitor p27kip1. In primary cultures of dog thyroid epithelial cells, the cAMP-dependent cell Cycle induced by a sustained stimulation by thyrotropin or forskolin differs from growth factor mitogenic pathways, as cAMP does not upregulate d-type cyclins but increases p27 levels. Instead, cAMP induces the assembly of required cyclin D3-CDK4 complexes, which associate with nuclear p27. In this study, the arrest of forskolin stimulation rapidly slowed down the entry of dog thyrocytes into S phase and the phosphorylation of pRb family proteins. The pRb kinase activity, but not the formation, of the cyclin D3-CDK4-p27 complex was strongly reduced. Using two-dimensional gel electrophoresis, a phosphorylated form of CDK4 was separated. It appeared in response to forskolin and was bound to both cyclin D3 and p27, presumably reflecting the activating Thr-172 phosphorylation of CDK4. Upon forskolin withdrawal or after cycloheximide addition, this CDK4 phosphoform unexpectedly persisted in p27 complexes devoid of cyclin D3 but it disappeared from the more labile cyclin D3 complexes. These data demonstrate that the assembly of the cyclin D3-CDK4-p27 holoenzyme and the subsequent phosphorylation and activation of CDK4 depend on distinct cAMP actions. This provides a first example of a crucial regulation of CDK4 phosphorylation by a mitogenic cascade and a novel mechanism of cell Cycle control at the R point.
Bela Novak - One of the best experts on this subject based on the ideXlab platform.
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Cell Cycle Commitment in budding yeast emerges from the cooperation of multiple bistable switches
Open biology, 2011Co-Authors: Tongli Zhang, Bernhard Schmierer, Bela NovakAbstract:The start-transition (START) in the G1 phase marks the point in the cell Cycle at which a yeast cell initiates a new round of cell division. Once made, this decision is irreversible and the cell is committed to progressing through the entire cell Cycle, irrespective of arrest signals such as pheromone. How Commitment emerges from the underlying molecular interaction network is poorly understood. Here, we perform a dynamical systems analysis of an established cell Cycle model, which has never been analysed from a Commitment perspective. We show that the irreversibility of the START transition and subsequent Commitment can be consistently explained in terms of the interplay of multiple bistable molecular switches. By applying an existing mathematical model to a novel problem and by expanding the model in a self-consistent manner, we achieve several goals: we bring together a large number of experimental findings into a coherent theoretical framework; we increase the scope and the applicability of the original model; we give a systems level explanation of how the START transition and the cell Cycle Commitment arise from the dynamical features of the underlying molecular interaction network; and we make clear, experimentally testable predictions.
Chun Liang - One of the best experts on this subject based on the ideXlab platform.
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Cdc48p is required for the cell Cycle Commitment point at Start via degradation of the G1-CDK inhibitor Far1p.
The Journal of cell biology, 2003Co-Authors: Daorong Feng, Chun LiangAbstract:The budding yeast Cdc48p and its mammalian homologue p97 are involved in many important cellular activities. Because previous cdc48 mutants have exclusive G2/M arrest, Cdc48p was thought to play an essential role only during mitosis. We found that Cdc48p is required for the execution of Start (a yeast cell Cycle Commitment point equivalent to the restriction point in mammalian cells) in both a normal mitotic cell Cycle and cell Cycle reentry after mating pheromone withdrawal through degradation of the G1-cyclin-dependent kinase inhibitor Far1p. Our work is the first to uncover novel roles of Cdc48p as a critical cell Cycle regulator in G1, and to shed new light on cell Cycle regulation of Far1p, which is the first cyclin-dependent kinase inhibitor shown to be a substrate of an essential proteolysis event mediated by Cdc48p.
Katia Coulonval - One of the best experts on this subject based on the ideXlab platform.
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cdk4 t172 phosphorylation is central in a cdk7 dependent bidirectional cdk4 cdk2 interplay mediated by p21 phosphorylation at the restriction point
PLOS Genetics, 2013Co-Authors: Xavier Bisteau, Sabine Paternot, Katia Coulonval, Bianca Colleoni, Karin Ecker, Philippe De Groote, Wim Declercq, Ludger Hengst, Pierre P. RogerAbstract:Cell Cycle progression, including genome duplication, is orchestrated by cyclin-dependent kinases (CDKs). CDK activation depends on phosphorylation of their T-loop by a CDK-activating kinase (CAK). In animals, the only known CAK for CDK2 and CDK1 is cyclin H-CDK7, which is constitutively active. Therefore, the critical activation step is dephosphorylation of inhibitory sites by Cdc25 phosphatases rather than unrestricted T-loop phosphorylation. Homologous CDK4 and CDK6 bound to cyclins D are master integrators of mitogenic/oncogenic signaling cascades by initiating the inactivation of the central oncosuppressor pRb and cell Cycle Commitment at the restriction point. Unlike the situation in CDK1 and CDK2 cyclin complexes, and in contrast to the weak but constitutive T177 phosphorylation of CDK6, we have identified the T-loop phosphorylation at T172 as the highly regulated step determining CDK4 activity. Whether both CDK4 and CDK6 phosphorylations are catalyzed by CDK7 remains unclear. To answer this question, we took a chemical-genetics approach by using analogue-sensitive CDK7(as/as) mutant HCT116 cells, in which CDK7 can be specifically inhibited by bulky adenine analogs. Intriguingly, CDK7 inhibition prevented activating phosphorylations of CDK4/6, but for CDK4 this was at least partly dependent on its binding to p21 (cip1) . In response to CDK7 inhibition, p21-binding to CDK4 increased concomitantly with disappearance of the most abundant phosphorylation of p21, which we localized at S130 and found to be catalyzed by both CDK4 and CDK2. The S130A mutation of p21 prevented the activating CDK4 phosphorylation, and inhibition of CDK4/6 and CDK2 impaired phosphorylations of both p21 and p21-bound CDK4. Therefore, specific CDK7 inhibition revealed the following: a crucial but partly indirect CDK7 involvement in phosphorylation/activation of CDK4 and CDK6; existence of CDK4-activating kinase(s) other than CDK7; and novel CDK7-dependent positive feedbacks mediated by p21 phosphorylation by CDK4 and CDK2 to sustain CDK4 activation, pRb inactivation, and restriction point passage.
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Cyclic AMP-dependent Phosphorylation of Cyclin D3-bound CDK4 Determines the Passage through the Cell Cycle Restriction Point in Thyroid Epithelial Cells
The Journal of biological chemistry, 2003Co-Authors: Sabine Paternot, Katia Coulonval, Jacques Emile Dumont, Pierre P. RogerAbstract:According to current concepts, the cell Cycle Commitment after restriction (R) point passage requires the sustained stimulation by mitogens of the synthesis of labile d-type cyclins, which associate with cyclin-dependent kinase (CDK) 4/6 to phosphorylate pRb family proteins and sequester the CDK inhibitor p27kip1. In primary cultures of dog thyroid epithelial cells, the cAMP-dependent cell Cycle induced by a sustained stimulation by thyrotropin or forskolin differs from growth factor mitogenic pathways, as cAMP does not upregulate d-type cyclins but increases p27 levels. Instead, cAMP induces the assembly of required cyclin D3-CDK4 complexes, which associate with nuclear p27. In this study, the arrest of forskolin stimulation rapidly slowed down the entry of dog thyrocytes into S phase and the phosphorylation of pRb family proteins. The pRb kinase activity, but not the formation, of the cyclin D3-CDK4-p27 complex was strongly reduced. Using two-dimensional gel electrophoresis, a phosphorylated form of CDK4 was separated. It appeared in response to forskolin and was bound to both cyclin D3 and p27, presumably reflecting the activating Thr-172 phosphorylation of CDK4. Upon forskolin withdrawal or after cycloheximide addition, this CDK4 phosphoform unexpectedly persisted in p27 complexes devoid of cyclin D3 but it disappeared from the more labile cyclin D3 complexes. These data demonstrate that the assembly of the cyclin D3-CDK4-p27 holoenzyme and the subsequent phosphorylation and activation of CDK4 depend on distinct cAMP actions. This provides a first example of a crucial regulation of CDK4 phosphorylation by a mitogenic cascade and a novel mechanism of cell Cycle control at the R point.
