Team:ETHZ Basel/Modeling/Light Switch
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= Modeling of the light switch = | = Modeling of the light switch = | ||
+ | [[Image:ETHZ_Basel_molecular_comb.png|thumb|400px|'''Schematical overview of the devices and change upon light pulse induction.''' LSP refers to light switch protein, AP to anchor protein, anchor to the plasmid anchor and Che to the attacked protein of the chemotaxis pathway.]] | ||
- | == | + | == Relocation of the proteins by red and far-red light == |
- | + | Relocation of the Che proteins was achieved by fusing them to a light-sensitive protein LSP1 (either to PhyB or to PIF3), which dimerizes by red light induction with the corresponding LSP2 (PIF3 or PhyB), fused to a spatially dislocated anchor. To model the relocation, we decided to adapt a model recently published by Sorokina et al [[Team:ETHZ Basel/Modeling/Light_Switch#References|[1]]]. | |
- | + | ||
- | | | + | == General remarks == |
+ | In [[Team:ETHZ Basel/Modeling/Light_Switch#References|[1]]] Sorokina et al developed (''in vivo'') and modeled (''in silico'') the reversible activation of different phytochromes by red light and their deactivation by far-red light in yeast. Their main model is based on the proteins PhyA and FHL, which have similar properties as PhyB and PIF3 used in E. lemming , as described by the underlying [[Team:ETHZ_Basel/Biology/Molecular_Mechanism|molecular mechanism]]. | ||
+ | In this system, PhyA is fused to the Gal4 bindig domain (GBD) and FHL to the Gal4 transcriptional activator domain. Upon activation with red light, both chimeric proteins bind to each other and the complex can activate the transcription of a lacZ reporter which is under the control of a GAL4-responsive artificial promoter. Upon deactivation with far-red light, the complex dissociates and transcription of the lacZ gene is significantly reduced. | ||
- | + | In this model, Sorokina et al assume that PhyA protein sequesteres and only free PhyA can interact with FHL. They state that this interaction is necessary to explain the experimental data presented in the paper, consisting of the strength of fluorescence induced by lacZ over time for different experimental settings. It should be noticed that this signal constantly changes with time in the range of hours. | |
- | == | + | == Implementation == |
- | + | One of the models of system described by Sorokina et al [[Team:ETHZ Basel/Modeling/Light_Switch#References|[1]]] is available in the SBML format [[Team:ETHZ Basel/Modeling/Light_Switch#References|[2]]]. The model was exported to Matlab using SBToolbox2 [[Team:ETHZ Basel/Modeling/Light_Switch#References|[3]]]. However, only the system without input light signals was possible to be reproduced as presented in [[Team:ETHZ Basel/Modeling/Light_Switch#References|[1]]], since the parameters necessary to define these signals were missing. | |
- | + | Furthermore, the model available in SBML format was slightly different from the model as described in the article: it was not possible to set the strength of the red and far-red light signals directly, but only their derivatives could be adjusted. This modeling decision might have been made to circumvent the problem that some integrators fail to solve a system of differential equations correctly when facing a step change too high in one of the derivatives. To avoid this problem and test the model, two measures were necessary: | |
+ | * the light inputs were not changed by steps, but by ramps with a length of 1s, which is significantly smaller than the other time constants relevant for this part of the model. | ||
+ | * the strengths of the light signals were decreased by approximately 40 fold. | ||
+ | |||
+ | == Evaluation of the model == | ||
+ | {| border="0" align="center" | ||
+ | |- valign="top" | ||
+ | |[[Image:Sorokina1.jpg|thumb|440px|'''Figure 1.''' Reaction of the model to no light input (dashed black curve), red light for 10 min at t=18h (black curve), red light for 10 min at t=18h followed by a 10min far-red pulse after 0h (blue), 0.5h (cyan), 1h (red), 3h (green), 9h (blue). The values for RLU were set so that the data reproduced the measured data as described by Sorokina et al. This figure corresponds to figure 6A in [[Team:ETHZ Basel/Modeling/Light_Switch#References|[1]]].]] | ||
+ | |[[Image:Sorokina2.jpg|thumb|440px|'''Figure 2.''' Same as Figure 1, only with the same value for RLU for every experiment (set to a value to reproduce Figure 8A in [[Team:ETHZ Basel/Modeling/Light_Switch#References|[1]).]]]] | ||
+ | |} | ||
+ | |||
+ | With the above changes, it was possible to reproduce the data as presented in the paper. Two of the most significant figures (see Figure 1 and 2) were reproduced, which show the response of the system to different light input signals: no light input (dark), a single red light pulse at t=18h lasting for 10 min, or a 10 min red light pulse at t=18h followed by a 10 min far-red light pulse after 0.5h, 1h, 3h, or 9h. All experiments qualitatively and quantitaively reproduced the simulation results obtained by Sorokina et al (see Figure 6B and Figure 8A in [[Team:ETHZ Basel/Modeling/Light_Switch#References|[1]]]). | ||
+ | |||
+ | == References == | ||
+ | [1] [http://www.jbioleng.org/content/3/1/15 Sorokina et al: A switchable light-input, light-output system modelled and constructed in yeast. J Biol Eng. 2009 Sep 17;3:15.] | ||
+ | |||
+ | [2] [http://www.biomedcentral.com/content/supplementary/1754-1611-3-15-S3.XML 1754-1611-3-15-S3.XML] | ||
+ | |||
+ | [3] [http://www.sbtoolbox2.org/ SBToolbox2] |
Revision as of 14:55, 25 October 2010
Modeling of the light switch
Relocation of the proteins by red and far-red light
Relocation of the Che proteins was achieved by fusing them to a light-sensitive protein LSP1 (either to PhyB or to PIF3), which dimerizes by red light induction with the corresponding LSP2 (PIF3 or PhyB), fused to a spatially dislocated anchor. To model the relocation, we decided to adapt a model recently published by Sorokina et al [1].
General remarks
In [1] Sorokina et al developed (in vivo) and modeled (in silico) the reversible activation of different phytochromes by red light and their deactivation by far-red light in yeast. Their main model is based on the proteins PhyA and FHL, which have similar properties as PhyB and PIF3 used in E. lemming , as described by the underlying molecular mechanism. In this system, PhyA is fused to the Gal4 bindig domain (GBD) and FHL to the Gal4 transcriptional activator domain. Upon activation with red light, both chimeric proteins bind to each other and the complex can activate the transcription of a lacZ reporter which is under the control of a GAL4-responsive artificial promoter. Upon deactivation with far-red light, the complex dissociates and transcription of the lacZ gene is significantly reduced.
In this model, Sorokina et al assume that PhyA protein sequesteres and only free PhyA can interact with FHL. They state that this interaction is necessary to explain the experimental data presented in the paper, consisting of the strength of fluorescence induced by lacZ over time for different experimental settings. It should be noticed that this signal constantly changes with time in the range of hours.
Implementation
One of the models of system described by Sorokina et al [1] is available in the SBML format [2]. The model was exported to Matlab using SBToolbox2 [3]. However, only the system without input light signals was possible to be reproduced as presented in [1], since the parameters necessary to define these signals were missing.
Furthermore, the model available in SBML format was slightly different from the model as described in the article: it was not possible to set the strength of the red and far-red light signals directly, but only their derivatives could be adjusted. This modeling decision might have been made to circumvent the problem that some integrators fail to solve a system of differential equations correctly when facing a step change too high in one of the derivatives. To avoid this problem and test the model, two measures were necessary:
- the light inputs were not changed by steps, but by ramps with a length of 1s, which is significantly smaller than the other time constants relevant for this part of the model.
- the strengths of the light signals were decreased by approximately 40 fold.
Evaluation of the model
![]() Figure 1. Reaction of the model to no light input (dashed black curve), red light for 10 min at t=18h (black curve), red light for 10 min at t=18h followed by a 10min far-red pulse after 0h (blue), 0.5h (cyan), 1h (red), 3h (green), 9h (blue). The values for RLU were set so that the data reproduced the measured data as described by Sorokina et al. This figure corresponds to figure 6A in [1]. | ![]() Figure 2. Same as Figure 1, only with the same value for RLU for every experiment (set to a value to reproduce Figure 8A in [1]). |
With the above changes, it was possible to reproduce the data as presented in the paper. Two of the most significant figures (see Figure 1 and 2) were reproduced, which show the response of the system to different light input signals: no light input (dark), a single red light pulse at t=18h lasting for 10 min, or a 10 min red light pulse at t=18h followed by a 10 min far-red light pulse after 0.5h, 1h, 3h, or 9h. All experiments qualitatively and quantitaively reproduced the simulation results obtained by Sorokina et al (see Figure 6B and Figure 8A in [1]).
References
[1] [http://www.jbioleng.org/content/3/1/15 Sorokina et al: A switchable light-input, light-output system modelled and constructed in yeast. J Biol Eng. 2009 Sep 17;3:15.]
[2] [http://www.biomedcentral.com/content/supplementary/1754-1611-3-15-S3.XML 1754-1611-3-15-S3.XML]
[3] [http://www.sbtoolbox2.org/ SBToolbox2]