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An Extended, Boolean Model of the Septation Initiation Network in S.Pombe Provides Insights into Its Regulation.

Chasapi A, Wachowicz P, Niknejad A, Collin P, Krapp A, Cano E, Simanis V, Xenarios I - PLoS ONE (2015)

Bottom Line: In order to describe the qualitative behavior of the system and predict unknown mutant behaviors we decided to adopt a Boolean modeling approach.The model was optimized using single knock-out experiments of known phenotypic effect as a training set, and was able to correctly predict a double knock-out test set.Moreover, the model has made in silico predictions that have been validated in vivo, providing new insights into the regulation and hierarchical organization of the SIN.

View Article: PubMed Central - PubMed

Affiliation: Vital-IT Group, Swiss Institute of Bioinformatics (SIB), Lausanne, Switzerland.

ABSTRACT
Cytokinesis in fission yeast is controlled by the Septation Initiation Network (SIN), a protein kinase signaling network using the spindle pole body as scaffold. In order to describe the qualitative behavior of the system and predict unknown mutant behaviors we decided to adopt a Boolean modeling approach. In this paper, we report the construction of an extended, Boolean model of the SIN, comprising most SIN components and regulators as individual, experimentally testable nodes. The model uses CDK activity levels as control nodes for the simulation of SIN related events in different stages of the cell cycle. The model was optimized using single knock-out experiments of known phenotypic effect as a training set, and was able to correctly predict a double knock-out test set. Moreover, the model has made in silico predictions that have been validated in vivo, providing new insights into the regulation and hierarchical organization of the SIN.

No MeSH data available.


Related in: MedlinePlus

Cdc7p over-expression in a sid4 mutant will result in septation.(A) Steady states of in silico, double mutation experiments. The model predicts that in the absence of SIN scaffold proteins (Cdc11p or Sid4p) and over-expression of Cdc7p, the cell will septate. (B) sid4-SA1 leu1-32 was transformed with a REP1-based plasmids [80] expressing cdc7; empty vector served as a control. Cells were grown to exponential phase in EMM2 medium at 25°C containing 2mM thiamine. Expression was induced by washing with EMM2 and growth for 16h at 25°C; cells were then shifted to 36°C for 5h, fixed, and stained with DAPI and Calcofluor as described [81]. Note that the cells carrying empty vector have become elongated and multinucleated, while 75% of cells expressing cdc7 have one or more septa. The scale bar represents 10 μm. (C) The strain leu1::pADH1-cdc7 was grown to exponential phase in YE medium at 19°C. A sample was taken and cells were fixed and stained with DAPI and Calcofluor. The remainder of the culture was incubated for 5h at 36°C before fixation. Note the elevated percentage of septated cells. The scale bar represents 10 μm. (D) The indicated strains were grown to exponential phase in YE medium, counted, and diluted to 106 ml-1. 10 μl of serial 5-fold dilutions were spotted on plates, allowed to dry and then incubated at the indicated temperature until the wild-type control had formed colonies.
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pone.0134214.g004: Cdc7p over-expression in a sid4 mutant will result in septation.(A) Steady states of in silico, double mutation experiments. The model predicts that in the absence of SIN scaffold proteins (Cdc11p or Sid4p) and over-expression of Cdc7p, the cell will septate. (B) sid4-SA1 leu1-32 was transformed with a REP1-based plasmids [80] expressing cdc7; empty vector served as a control. Cells were grown to exponential phase in EMM2 medium at 25°C containing 2mM thiamine. Expression was induced by washing with EMM2 and growth for 16h at 25°C; cells were then shifted to 36°C for 5h, fixed, and stained with DAPI and Calcofluor as described [81]. Note that the cells carrying empty vector have become elongated and multinucleated, while 75% of cells expressing cdc7 have one or more septa. The scale bar represents 10 μm. (C) The strain leu1::pADH1-cdc7 was grown to exponential phase in YE medium at 19°C. A sample was taken and cells were fixed and stained with DAPI and Calcofluor. The remainder of the culture was incubated for 5h at 36°C before fixation. Note the elevated percentage of septated cells. The scale bar represents 10 μm. (D) The indicated strains were grown to exponential phase in YE medium, counted, and diluted to 106 ml-1. 10 μl of serial 5-fold dilutions were spotted on plates, allowed to dry and then incubated at the indicated temperature until the wild-type control had formed colonies.

Mentions: Apart from the experiments that were used as training set for the model refinement, we performed double mutant experiments towards which the model had not been optimized (test set). These experiments assess the predictive value of the model, as the in silico predictions are in accordance with the expected results. Specifically, the double deletion of cdc11 and cdc16 simulation predicts that cells should not septate, as shown in Fig 3, with supporting evidence from the literature [78]. A Cdc7p over-expression in an spg1 deletion mutant will septate, in agreement with in vivo studies [13]. Moreover, Cdc7p over-expression will produce septation in the absence of Cdc11p (Fig 4A), as confirmed by the literature [79]. In this project, setting a node to 1 throughout the simulation has been used to simulate over-expression in silico, except in cases where it is known that the over-expression phenotype results from an indirect effect, such as the titration of another protein.


An Extended, Boolean Model of the Septation Initiation Network in S.Pombe Provides Insights into Its Regulation.

Chasapi A, Wachowicz P, Niknejad A, Collin P, Krapp A, Cano E, Simanis V, Xenarios I - PLoS ONE (2015)

Cdc7p over-expression in a sid4 mutant will result in septation.(A) Steady states of in silico, double mutation experiments. The model predicts that in the absence of SIN scaffold proteins (Cdc11p or Sid4p) and over-expression of Cdc7p, the cell will septate. (B) sid4-SA1 leu1-32 was transformed with a REP1-based plasmids [80] expressing cdc7; empty vector served as a control. Cells were grown to exponential phase in EMM2 medium at 25°C containing 2mM thiamine. Expression was induced by washing with EMM2 and growth for 16h at 25°C; cells were then shifted to 36°C for 5h, fixed, and stained with DAPI and Calcofluor as described [81]. Note that the cells carrying empty vector have become elongated and multinucleated, while 75% of cells expressing cdc7 have one or more septa. The scale bar represents 10 μm. (C) The strain leu1::pADH1-cdc7 was grown to exponential phase in YE medium at 19°C. A sample was taken and cells were fixed and stained with DAPI and Calcofluor. The remainder of the culture was incubated for 5h at 36°C before fixation. Note the elevated percentage of septated cells. The scale bar represents 10 μm. (D) The indicated strains were grown to exponential phase in YE medium, counted, and diluted to 106 ml-1. 10 μl of serial 5-fold dilutions were spotted on plates, allowed to dry and then incubated at the indicated temperature until the wild-type control had formed colonies.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4526654&req=5

pone.0134214.g004: Cdc7p over-expression in a sid4 mutant will result in septation.(A) Steady states of in silico, double mutation experiments. The model predicts that in the absence of SIN scaffold proteins (Cdc11p or Sid4p) and over-expression of Cdc7p, the cell will septate. (B) sid4-SA1 leu1-32 was transformed with a REP1-based plasmids [80] expressing cdc7; empty vector served as a control. Cells were grown to exponential phase in EMM2 medium at 25°C containing 2mM thiamine. Expression was induced by washing with EMM2 and growth for 16h at 25°C; cells were then shifted to 36°C for 5h, fixed, and stained with DAPI and Calcofluor as described [81]. Note that the cells carrying empty vector have become elongated and multinucleated, while 75% of cells expressing cdc7 have one or more septa. The scale bar represents 10 μm. (C) The strain leu1::pADH1-cdc7 was grown to exponential phase in YE medium at 19°C. A sample was taken and cells were fixed and stained with DAPI and Calcofluor. The remainder of the culture was incubated for 5h at 36°C before fixation. Note the elevated percentage of septated cells. The scale bar represents 10 μm. (D) The indicated strains were grown to exponential phase in YE medium, counted, and diluted to 106 ml-1. 10 μl of serial 5-fold dilutions were spotted on plates, allowed to dry and then incubated at the indicated temperature until the wild-type control had formed colonies.
Mentions: Apart from the experiments that were used as training set for the model refinement, we performed double mutant experiments towards which the model had not been optimized (test set). These experiments assess the predictive value of the model, as the in silico predictions are in accordance with the expected results. Specifically, the double deletion of cdc11 and cdc16 simulation predicts that cells should not septate, as shown in Fig 3, with supporting evidence from the literature [78]. A Cdc7p over-expression in an spg1 deletion mutant will septate, in agreement with in vivo studies [13]. Moreover, Cdc7p over-expression will produce septation in the absence of Cdc11p (Fig 4A), as confirmed by the literature [79]. In this project, setting a node to 1 throughout the simulation has been used to simulate over-expression in silico, except in cases where it is known that the over-expression phenotype results from an indirect effect, such as the titration of another protein.

Bottom Line: In order to describe the qualitative behavior of the system and predict unknown mutant behaviors we decided to adopt a Boolean modeling approach.The model was optimized using single knock-out experiments of known phenotypic effect as a training set, and was able to correctly predict a double knock-out test set.Moreover, the model has made in silico predictions that have been validated in vivo, providing new insights into the regulation and hierarchical organization of the SIN.

View Article: PubMed Central - PubMed

Affiliation: Vital-IT Group, Swiss Institute of Bioinformatics (SIB), Lausanne, Switzerland.

ABSTRACT
Cytokinesis in fission yeast is controlled by the Septation Initiation Network (SIN), a protein kinase signaling network using the spindle pole body as scaffold. In order to describe the qualitative behavior of the system and predict unknown mutant behaviors we decided to adopt a Boolean modeling approach. In this paper, we report the construction of an extended, Boolean model of the SIN, comprising most SIN components and regulators as individual, experimentally testable nodes. The model uses CDK activity levels as control nodes for the simulation of SIN related events in different stages of the cell cycle. The model was optimized using single knock-out experiments of known phenotypic effect as a training set, and was able to correctly predict a double knock-out test set. Moreover, the model has made in silico predictions that have been validated in vivo, providing new insights into the regulation and hierarchical organization of the SIN.

No MeSH data available.


Related in: MedlinePlus