E-CELL2 User's Manual Chapter 2 Tutorial [Chapter 1:Chapter 3:Chapter 4:Chapter 5:Chapter 6][Top]

# 1 Features of E-CELL2

In this chapter, the features and operation methods of E-CELL2.26 will be outlined. For details regarding the theoretical background of E-CELL, please refer to the tutorial available at E-CELL.Org(http://www.e-cell.org).

To model a cell, the abstraction of its components is necessary. In E-CELL, the "Substance-Reactor Model" has been adopted for describing the structures of a cell and the chemical reactions inside a cell. This model has the following features.

• The state of a cell is regarded as a set of "quantity of substances (Substance)".
• The activity of a cell is represented by a "change in the quantity of a Substance", using a chemical reaction equation (Reactor).
• Nodes of Substance connected by a Reactor constitute a directed graph.

Figure 1. Substance-Reactor Model

• Cellular organelles are described via the location (System) of substances.

Figure 2. Substance-Reactor Model inside a System

Please note that the concept of a substance (Substance) includes not only chemical molecules, ions, and radicals, but also voluntary physical properties such as osmotic pressure and cell volume. Negative values however cannot be described.

In E-CELL, it is assumed that the intracellular concentration of a substance is uniform throughout the cell. To handle diffusion, it is therefore necessary to prepare multiple systems each with varying substance quantities.

To simulate various intracellular phenomena in E-CELL, both rate(flux)-based reactions such as enzymatic reactions, and rapid reactions such as chemical equilibrium can be implemented.

Flux-based (i.e. can be solved as differential equations) chemical reactions are handled inside E-CELL as follows.

• Changes the rate of chemical reaction equations using a standard (Regular) Reactor.
• Solves the ordinary differential equations using the Runge-Kutta method.
• Michaelis-Menten reactions are examples of rate equations.
Chemical equibrium-based (i.e. can be solved as algebraic equations) chemical reactions are handled as follows.
• Manipulates the quantity of Substances directly using a back door (Postern) Reactor.
• Phenomena such as rapid equilibrium and changes in osmotic pressure are examples of this type of reaction.

From the standpoint of software engineering, E-CELL2 has the following features. The part written in C++ will be described first. This part, the so-called heart of E-Cell2, corresponds to the view of the world (Ontology) that shows how E-CELL2 describes a cell. Such components are called "classes" in the field of object-oriented modeling. The computation mechanisms necessary for simulation are implemented by the interactions between the classes.

Figure 3. Internal structure of E-CELL2

The differences between E-CELL2 and the Linux-based E-CELL1 are listed below.

• Multiple class inheritance has been abolished in E-CELL2.
• Friend functions have also been abolished in E-CELL2.
• The "Integrator" and "Accumulator" classes have been integrated into the "Substance" class.
• The " Stepper"class has been integrated into the "System" class.
• A modeling launcher has been implemented

These changes were made to improve the portability of E-CELL1, which only works on Linux. The GUI (Graphic User Interface) and the modules for interpreting the script files for automating the operation of E-CELL2 are written in Java.

In E-CELL2, there is a GUI mode that allows for interactive operation, and a batch mode that supports command-line execution. This chapter provides explanations for only the former mode(GUI mode). Please refer to Chapter 3 Section 2.9 "Batch mode and Logger", for explanations on the latter mode(batch mode). Please note that the accuracy of the GUI mode and that of the batch mode is 64 bits and 80 bits, respectively. The inferior accuracy of the GUI mode is due to the limitations in numerical computation that exist in the Java Native Interface (JNI).

# 2 Demonstration of E-CELL2.26

E-CELL2 will be operated here using an actual model. In this section, a metabolic model of the Human erythrocyte will be simulated using E-CELL2. Why is an erythrocyte suitable as a target of simulation? The answers to this question are as follows.

• Mature erythrocyte cells do not undergo transcription, translation, or replication, and are therefore simple.
• Because erythrocyte cells are relatively easy to handle as experimental materials, there is a large accumulation of biochemical experimental data available.

The main metabolic pathways of an erythrocyte consist of glycolysis, pentose phosphate cycle, and nucleotide synthesis. In addition, large amounts of oxygen-carrying hemoglobin exist in an erythrocyte. An erythrocyte cell can therefore be likened to a bag full of hemoglobin that has self-regulating functions based on these metabolic pathways. Although this model aims at the reconstruction of only the metabolic pathways, it is expected that hemoglobin transport will be included in a future model.

Figure 4. Model diagram of the Human erythrocyte metabolic pathway

This metabolic model of the Human erythrocyte cell was constructed by assistant professor Nakayama, professor Tomita, and their colleagues at Keio University. Although only the metabolic model of a normal human erythrocyte is introduced in this demonstration, models of anemia caused by genetic disorders are also currently being simulated. Please contact assistant professor Nakayama (ynakayam@sfc.keio.ac.jp) for further information.

To observe an actual simulation of the erythrocyte cell, please select [Program]-[E-CELL2]-[erythrocyte] from [Start], or click on the icon in your desktop.

E-CELL2, loaded with the erythrocyte model, will start. If E-CELL2 finds the file "default.ecs" in the same directory as ECELL2.BAT, it reads the file(default.ecs) and attempts to start simulation. Because a default.ecs file has already been prepared for the erythrocyte model shown in Fig. 4, the demonstration will automatically start. The file "Erythrocyte_v236.eri" describes the parameters and initial values for this simulation.

Files with the ".ecs" extension usually are scripts for automating E-CELL2, and include instructions for operating E-CELL2. For details, please refer to Section 2.3 and Section 3.4. The simulation of the erythrocyte model shown in Fig. 4 will start by executing this script. Using this script, simulation would stop after 500 seconds.

Figure 5. Simulation of the erythrocyte model

To terminate E-CELL2 before the script stops the simulation, please press the "Stop" button in the Control Panel, and then select [Quit] from the [File] menu.

In the next section, the operating procedure for E-CELL2 will be briefly explained. After understanding the basic operation procedure, please reload E-CELL2 and experiment with it.

# 3 Creating Rules for E-CELL2

Before going into the details, a simple simulation model will be introduced. The figure below illustrates the so-called "Toy" model, which includes a simple feedback system.

Figure 6. Toy model

5 "S"ubstrates, 4 "E"nzymes, and 1 "C"omplex constitute the model. The Substrates are interconnected via 4 Reactors (chemical reaction equations). Reactors embody chemical reaction equations, such as those based on the Michaelis-Menten scheme. In the case of a cultured cell, these reactions would exist inside a Cell, surrounded by culture medium called Environment. A Membrane compartmentalizes the external world and the cell interior, which is filled with Cytoplasm.

The network of chemical reactions has now been depicted as static graph.

However, to actually start simulation, the rules that define the parameters for each reaction(e.g. initial values, rates),and how the reactions proceed(i.e. which chemical reaction equations to choose), have to be decided.

When creating models for E-CELL, simulation rules called Rule Files, which are distinguished by the ".er" extension, are described using text editors. Then, using tools included with E-CELL, files that can be directly read by E-CELL called Rule Intermediate Files are created. Rule Intermediate Files are usually distinguished by the ".eri" extension.

Because it is difficult for beginners of E-CELL to describe Rule Files from scratch, tools for creating Rule Files from spreadsheet files are also provided to support the Rule File creation process.

Here, the methods for creating spreadsheets for simulation, and converting them to Rule Intermediate Files for E-CELL2 will be outlined.

The minimum operating procedures necessary for creating Rule Files using the modeling launcher, which is included with E-CELL2, will be explained.

To start the modeling launcher, please select [Program]-[E-CELL2]-[ModelingLauncher] from [Start], or click on the icon in your desktop.

Figure 7. Window of the modeling launcher(1)

Figure 8. Window of the modeling launcher(2)

After finishing editing, please save the file by choosing [Save] or [Save As] from the [File] menu.

## 3.2 Creating Rule Files and Rule Intermediate Files

The procedures for converting spreadsheets into Rule Files and Rule Files into Rule Intermediate Files will be explained. Please refer to Chapter 4 for details regarding Rule Files.

Please start the modeling launcher and load "sample.txt " and "toy.txt ". The file "toy.txt " is an incomplete spreadsheet file for the Toy model. These files are located in the tutorial-related "standard" directory.

It has already been explained that "Systems", "Substances", and "Reactors" exist in the Substance-Reactor model of E-CELL. The leftmost column in the spreadsheet is called Type. Each field will include one of the above Types. Systems are used to show the structures of a cell or the localization of substances. Systems usually include "Environment", "Cell", "Membrane", and "Cytoplasm". Please refer to Section 4.2.2 for details regarding Systems. Please keep in mind here that embedded structures such as cells can be described by distinguishing "Inside" from "Outside".

A Substance implies a broad definition of a substance. The spreadsheet includes a description similar to the table shown below. In the "Toy" model, Substances are the materials undergoing chemical reactions and the enzymes catalyzing the reactions.

Table 2.1 Example descriptions of Substances used as materials for chemical reactions

TypepathIDNameQTY
Substance/CELL/CYTOPLASMSASubstance A1000
FIX
Substance/CELL/CYTOPLASMSBSubstance B0
Substance/CELL/CYTOPLASMSCSubstance C0

Path is the location of the Substance. It is described in a fashion similar to the directory path in UNIX. ID is the ID of the specified Substance, and is an essential item. Name is the name of the specified Substance. Either the Qty or the Conc for a Substance has to described. Qty is the initial "Quantity" value of the Substance, whereas Conc is the initial "Concentration" value of the Substance. The unit of Conc is mol/l. If the Quantity and Conc should not change during simulation, the values may be fixed by adding a "Fix" tag to the row right beneath. Please refer to Section 4.2.3 for details regarding Substances.

The following are descriptions regarding enzymes.

Table 2.2 Example descriptions of Substances that catalyze reactions as enzymes

TypepathIDNameCONC
Substance/CELL/CYTOPLASME.bcEnzyme B0.02
Substance/CELL/CYTOPLASME.cdEnzyme C0.01
Substance/CELL/CYTOPLASMC.E.bc-SDComplex of E.bc and SD0

<Problems>
1. In "toy.txt", a few descriptions for Substance D and Substance E are missing. Please fill in the gaps by referring to "sample.txt".
2. Similarly, please complete the descriptions for Isomerase of A and Isomerase of D.

Reactors on the other hand are implemented chemical reactions. An example is defined below.

Table 2.3 Example descriptions of Reactors

TypeClasspathIDNameS_IDS_pathS_coeffP_IDP_pathP_coeffC_IDC_pathArg_tagArg_coeff
ReactorMichaelisUniUniReactor/CELL/CYTOPLASME.ab-0A->BSA/CELL/CYTOPLASM1SB/CELL/CYTOPLASM1E.ab/CELL/CYTOPLASMKmS10
KcF5
ReactorMichaelisUniUniReactor/CELL/CYTOPLASME.bc-0B->CSB/CELL/CYTOPLASM1SC/CELL/CYTOPLASM1E.bc/CELL/CYTOPLASMKmS0.1
KcF3
ReactorMichaelisUniUniReactor/CELL/CYTOPLASME.cd-0C->DSC/CELL/CYTOPLASM1SD/CELL/CYTOPLASM1E.cd/CELL/CYTOPLASMKmS0.1
KcF2

Path is the location of the Reactor. It is described in a fashion similar to the directory path in UNIX. S_ID is the "Substrate", or the starting substance of the chemical reaction. S_Path is its location. P_ID is the "Product", or the resulting substance of the chemical reaction. P_Path is its location. C_ID is the "Catalyst", or the chemical substance that promotes or represses the reaction. C_Path is its location. In the Arg_tag column, the constant names that are defined differently according to the type of Reactor are inputted. In the Arg_coeff column, the constant values that correspond to the Arg_tag are inputted.

Reactors whose IDs begin with "!" are back door Reactors, used for describing reactions that cannot be described by differential equations. In the "Toy" model, "!EQ-Ebc-D" is the algebraic equation-based back door Reactor. Ebc+SD < - > C.Ebc-D is a reversible reaction that consists of a reaction that produces C.Ebc-D from Ebc and SD, and a reaction that decomposes C.Ebc-D. Therefore, it cannot be solved by differential equations. Thus, "RapidEquibriumPReactor" for rapid equilibrium is used here.

<Problems>
1. Please complete the descriptions for the reaction in Reactor D->E.


Please refer to Section 4.2.4 for details regarding Reactors. If all of the above operations have been finished, please save "toy.txt", and start working on creating Rule Files.

The actual operations for creating Rule Files from spreadsheet files will be explained using the model launcher.

Please click on the "File..." button in the spreadsheet file column, and then open the file selection window by selecting "Choose...". Please search and load "toy.txt".

Figure 9. Selection of toy.txt

If the Execute button is clicked, a Rule File "toy.er" and a Rule Intermediate File "toy.eri" will be created.

The creation of Rule Files and Rule Intermediate Files have been explained in this section. Please proceed to the next section on creating Reactors.

# 4 Creating User-defined Reactors for E-CELL2

To describe a chemical reaction that is not included in the standard reactors, one must design and create a Reactor for E-CELL2.

## 4.1 Outline of Reactors

• Reactors calculate the temporal changes in the quantity of Substances.
• The source code is described in C++ language.
• Writing a Reactor in E-CELL, means writing a Reactor-describing (Reactor Description, RD) file (e.g., filename.rd).
• In an RD file, the specifications of the Reactor and the contents of the processes to be executed are written.
• Using rd2ch.pl and rd2tex.pl, RD files are converted into tex files(i.e. Reactor Spec Sheets in LaTeX format) and ".dll" files(i.e. Dynamic link libraries in an E-CELL-loadable format).

RD files are created by arranging lines consisting of pairs of keywords and values.

RD files can roughly be divided into the following three parts;
• General information
• Reactor Spec Sheet
• Reactor Source Code

When creating Reactor Description files, please conform to the following rules.

• Keywords consist of capital letters or "_" , and do not contain any spaces.
• Keywords begin with "@" or "%". The contents of lines beginning with "@" are directly read. The contents of lines beginning with "%" are delimited by "," and processed as arrays.
• Lines beginning with # are treated as comments. To directly output the "#" in the beginning of a line, please type "\#" or place a space before "#". However, the "#" other than those located in the beginning of a line are directly outputted.
• Lines that do not include keywords at the beginning are interpreted as a continuation of the previous lines. (To start a new line when converting to Spec Sheets, please place a " \\ " where the new line should start)
• Keywords may be omitted if they are not necessary.

An example RD file is shown below.

1. This is the file for the MichaelisUniUniReactor. This Reactor is based on the following rate reaction.
v = KcF[E][S]
KmS + [S]

The file for the MichaelisUniUniReactor

@CLASSNAME:MichaelisUniUniReactor

@BASECLASS: FluxReactor
@AUTHOR: E-CECLL Tutorial
@EMAIL: tutorial@e-cell.org
@DATE: 2000 12/12

%VERSION: ecs-v1, 0.1

@BRIEF_DESCRIPTION:Unireactor enzyme activity of which kinetics can be described by the Henri-Michaelis-Menten equation.

@DESCRIPTION:A reactor class for unireactant enzyme activity where kinetics can be described
by the Henri-Michaelis-Menten equation derived from rapid equilibrium assumptions.
\vspace{0.2cm}

This reactor is applicable to the following reaction sequence:

\begin{center}