Nur1 Dephosphorylation Confers Positive Feedback to Mitotic Exit Phosphatase Activation in Budding Yeast

AbstractCells can encode information about their environment by modulating signaling dynamics and responding accordingly. Yet, the mechanisms cells use to decode these dynamics remain unknown when cells respond exclusively to transient signals. Here, we approach design principles underlying such decoding by rationally engineering a synthetic short-pulse decoder in budding yeast. A computational method for rapid prototyping, TopoDesign, allowed us to explore 4122 possible circuit architectures, design targeted experiments, and then rationally select a single circuit for implementation. This circuit demonstrates short-pulse decoding through incoherent feedforward and positive feedback. We predict incoherent feedforward to be essential for decoding transient signals, thereby complementing proposed design principles of temporal filtering, the ability to respond to sustained signals, but not to transient signals. More generally, we anticipate TopoDesign to help designing other synthetic circuits with non-intuitive dynamics, simply by assembling available biological components.

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Computational modelling of mitotic exit in budding yeast: the role of separase and Cdc14 endocycles

Journal of The Royal Society Interface ◽

10.1098/rsif.2010.0649 ◽

2011 ◽

Vol 8 (61) ◽

pp. 1128-1141 ◽

Cited By ~ 20

Author(s):

P. K. Vinod ◽

Paula Freire ◽

Ahmed Rattani ◽

Andrea Ciliberto ◽

Frank Uhlmann ◽

...

Keyword(s):

Regulatory Networks ◽

Budding Yeast ◽

Cell Cycle Control ◽

Computational Modelling ◽

Eukaryotic Cell ◽

Mitotic Exit ◽

Intuitive Reasoning ◽

Systematic Analysis ◽

Systems View

The operating principles of complex regulatory networks are best understood with the help of mathematical modelling rather than by intuitive reasoning. Hereby, we study the dynamics of the mitotic exit (ME) control system in budding yeast by further developing the Queralt's model. A comprehensive systems view of the network regulating ME is provided based on classical experiments in the literature. In this picture, Cdc20–APC is a critical node controlling both cyclin (Clb2 and Clb5) and phosphatase (Cdc14) branches of the regulatory network. On the basis of experimental situations ranging from single to quintuple mutants, the kinetic parameters of the network are estimated. Numerical analysis of the model quantifies the dependence of ME control on the proteolytic and non-proteolytic functions of separase. We show that the requirement of the non-proteolytic function of separase for ME depends on cyclin-dependent kinase activity. The model is also used for the systematic analysis of the recently discovered Cdc14 endocycles. The significance of Cdc14 endocycles in eukaryotic cell cycle control is discussed as well.

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Order of function of the budding-yeast mitotic exit-network proteins Tem1, Cdc15, Mob1, Dbf2, and Cdc5

Current Biology ◽

10.1016/s0960-9822(01)00228-7 ◽

2001 ◽

Vol 11 (10) ◽

pp. 784-788 ◽

Cited By ~ 104

Author(s):

Sarah E. Lee ◽

Lisa M. Frenz ◽

Nicholas J. Wells ◽

Anthony L. Johnson ◽

Leland H. Johnston

Keyword(s):

Budding Yeast ◽

Mitotic Exit ◽

Mitotic Exit Network

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A Defect of Kap104 Alleviates the Requirement of Mitotic Exit Network Gene Functions in Saccharomyces cerevisiae

Genetics ◽

10.1093/genetics/162.4.1545 ◽

2002 ◽

Vol 162 (4) ◽

pp. 1545-1556

Author(s):

Kazuhide Asakawa ◽

Akio Toh-e

Keyword(s):

High Temperature ◽

Cell Cycle Progression ◽

Budding Yeast ◽

Mitotic Exit ◽

Temperature Sensitive ◽

Permissive Temperature ◽

Β Protein ◽

Responsible Gene ◽

Early Anaphase ◽

Mrna Binding

Abstract A subgroup of the karyopherin β (also called importin β) protein that includes budding yeast Kap104 and human transportin/karyopherin β2 is reported to function as a receptor for the transport of mRNA-binding proteins into the nucleus. We identified KAP104 as a responsible gene for a suppressor mutation of cdc15-2. We found that the kap104-E604K mutation suppressed the temperature-sensitive growth of cdc15-2 cells by promoting the exit from mitosis and suppressed the temperature sensitivity of various mitoticexit mutations. The cytokinesis defect of these mitotic-exit mutants was not suppressed by kap104-E604K. Furthermore, the kap104-E604K mutation delays entry into DNA synthesis even at a permissive temperature. In cdc15-2 kap104-E604K cells, SWI5 and SIC1, but not CDH1, became essential at a high temperature, suggesting that the kap104-E604K mutation promotes mitotic exit via the Swi5-Sic1 pathway. Interestingly, SPO12, which is involved in the release of Cdc14 from the nucleolus during early anaphase, also became essential in cdc15-2 kap104-E604K cells at a high temperature. The kap104-E604K mutation caused a partial delocalization of Cdc14 from the nucleolus during interphase. This delocalization of Cdc14 was suppressed by the deletion of SPO12. These results suggest that a mutation in Kap104 stimulates exit from mitosis through the activation of Cdc14 and implies a novel role for Kap104 in cell-cycle progression in budding yeast.

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A Mathematical Model of Mitotic Exit in Budding Yeast: The Role of Polo Kinase

PLoS ONE ◽

10.1371/journal.pone.0030810 ◽

2012 ◽

Vol 7 (2) ◽

pp. e30810 ◽

Cited By ~ 7

Author(s):

Baris Hancioglu ◽

John J. Tyson

Keyword(s):

Mathematical Model ◽

Budding Yeast ◽

Mitotic Exit ◽

Polo Kinase

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The Differential Roles of Budding Yeast Tem1p, Cdc15p, and Bub2p Protein Dynamics in Mitotic Exit

Molecular Biology of the Cell ◽

10.1091/mbc.e03-09-0708 ◽

2004 ◽

Vol 15 (4) ◽

pp. 1519-1532 ◽

Cited By ~ 63

Author(s):

Jeffrey N. Molk ◽

Scott C. Schuyler ◽

Jenny Y. Liu ◽

James G. Evans ◽

E. D. Salmon ◽

...

Keyword(s):

Budding Yeast ◽

Protein Localization ◽

Spindle Pole Body ◽

Spindle Pole ◽

Mitotic Exit ◽

Yeast Saccharomyces Cerevisiae ◽

Late Anaphase ◽

Gfp Fluorescence ◽

Mitotic Exit Network

In the budding yeast Saccharomyces cerevisiae the mitotic spindle must be positioned along the mother-bud axis to activate the mitotic exit network (MEN) in anaphase. To examine MEN proteins during mitotic exit, we imaged the MEN activators Tem1p and Cdc15p and the MEN regulator Bub2p in vivo. Quantitative live cell fluorescence microscopy demonstrated the spindle pole body that segregated into the daughter cell (dSPB) signaled mitotic exit upon penetration into the bud. Activation of mitotic exit was associated with an increased abundance of Tem1p-GFP and the localization of Cdc15p-GFP on the dSPB. In contrast, Bub2p-GFP fluorescence intensity decreased in mid-to-late anaphase on the dSPB. Therefore, MEN protein localization fluctuates to switch from Bub2p inhibition of mitotic exit to Cdc15p activation of mitotic exit. The mechanism that elevates Tem1p-GFP abundance in anaphase is specific to dSPB penetration into the bud and Dhc1p and Lte1p promote Tem1p-GFP localization. Finally, fluorescence recovery after photobleaching (FRAP) measurements revealed Tem1p-GFP is dynamic at the dSPB in late anaphase. These data suggest spindle pole penetration into the bud activates mitotic exit, resulting in Tem1p and Cdc15p persistence at the dSPB to initiate the MEN signal cascade.

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Asymmetry of the Budding Yeast Tem1 GTPase at Spindle Poles Is Required for Spindle Positioning But Not for Mitotic Exit

PLoS Genetics ◽

10.1371/journal.pgen.1004938 ◽

2015 ◽

Vol 11 (2) ◽

pp. e1004938 ◽

Cited By ~ 18

Author(s):

Ilaria Scarfone ◽

Marianna Venturetti ◽

Manuel Hotz ◽

Jette Lengefeld ◽

Yves Barral ◽

...

Keyword(s):

Budding Yeast ◽

Mitotic Exit ◽

Spindle Positioning ◽

Spindle Poles

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Lipid droplets maintain lipid homeostasis during anaphase for efficient cell separation in budding yeast

Molecular Biology of the Cell ◽

10.1091/mbc.e16-02-0106 ◽

2016 ◽

Vol 27 (15) ◽

pp. 2368-2380 ◽

Cited By ~ 19

Author(s):

Po-Lin Yang ◽

Tzu-Han Hsu ◽

Chao-Wen Wang ◽

Rey-Huei Chen

Keyword(s):

Cell Separation ◽

Neutral Lipids ◽

Budding Yeast ◽

Acid Synthesis ◽

Mitotic Exit ◽

Lipid Homeostasis ◽

Physiological Conditions ◽

Division Site ◽

Quadruple Mutant ◽

Membrane Bound

The neutral lipids steryl ester and triacylglycerol (TAG) are stored in the membrane-bound organelle lipid droplet (LD) in essentially all eukaryotic cells. It is unclear what physiological conditions require the mobilization or storage of these lipids. Here, we study the budding yeast mutant are1Δ are2Δ dga1Δ lro1Δ, which cannot synthesize the neutral lipids and therefore lacks LDs. This quadruple mutant is delayed at cell separation upon release from mitotic arrest. The cells have abnormal septa, unstable septin assembly during cytokinesis, and prolonged exocytosis at the division site at the end of cytokinesis. Lipidomic analysis shows a marked increase of diacylglycerol (DAG) and phosphatidic acid, the precursors for TAG, in the mutant during mitotic exit. The cytokinesis and separation defects are rescued by adding phospholipid precursors or inhibiting fatty acid synthesis, which both reduce DAG levels. Our results suggest that converting excess lipids to neutral lipids for storage during mitotic exit is important for proper execution of cytokinesis and efficient cell separation.

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Inhibition of Cdc42 during mitotic exit is required for cytokinesis

The Journal of Cell Biology ◽

10.1083/jcb.201301090 ◽

2013 ◽

Vol 202 (2) ◽

pp. 231-240 ◽

Cited By ~ 55

Author(s):

Benjamin D. Atkins ◽

Satoshi Yoshida ◽

Koji Saito ◽

Chi-Fang Wu ◽

Daniel J. Lew ◽

...

Keyword(s):

Cell Cycle ◽

General Feature ◽

Budding Yeast ◽

Mitotic Exit ◽

Division Site ◽

Polo Kinase ◽

Cell Cycle Regulator ◽

P21 Activated Kinase

The role of Cdc42 and its regulation during cytokinesis is not well understood. Using biochemical and imaging approaches in budding yeast, we demonstrate that Cdc42 activation peaks during the G1/S transition and during anaphase but drops during mitotic exit and cytokinesis. Cdc5/Polo kinase is an important upstream cell cycle regulator that suppresses Cdc42 activity. Failure to down-regulate Cdc42 during mitotic exit impairs the normal localization of key cytokinesis regulators—Iqg1 and Inn1—at the division site, and results in an abnormal septum. The effects of Cdc42 hyperactivation are largely mediated by the Cdc42 effector p21-activated kinase Ste20. Inhibition of Cdc42 and related Rho guanosine triphosphatases may be a general feature of cytokinesis in eukaryotes.

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