Model

The Quorum sensing module is mainly involved in receiving signals from the sender cells. The sender cells secrete some signalling molecules (inducers) which bind to the regulator molecules in the receiver cells, thus activating the transcription of certain genes. The model for this module is presented below.

Chemical Species

Reactions

For the Lux system

$$ \begin{align} &\rightarrow LuxR \\ LuxAHL+LuxR & \leftrightarrow RLux\\ RLux+RLux &\leftrightarrow DRLux\\ DRLux+P_{luxOFF} & \leftrightarrow P_{luxON}\\ P_{luxON}&\rightarrow P_{luxON}+mRNA_{Bxb1}\\ mRNA_{Bxb1}&\rightarrow Bxb1\\ AHL &\rightarrow \\ LuxR &\rightarrow \\ RLux &\rightarrow\\ DRLux &\rightarrow\\ mRNA_{Bxb1} &\rightarrow\\ Bxb1 &\rightarrow \end{align}$$

For the Las system

\begin{align} &\rightarrow LasR \\ LasAHL+LasR & \leftrightarrow RLas \\ RLas+RLas & \leftrightarrow DRLas\\ DRLas+P_{LasOFF} & \leftrightarrow P_{LasON}\\ P_{LasON}&\rightarrow P_{LasON}+mRNA_{\phi C31}\\ mRNA_{\phi C31}&\rightarrow \phi C31\\ Las-AHL &\rightarrow \\ LasR &\rightarrow \\ RLas &\rightarrow\\ DRLas &\rightarrow\\ mRNA_{\phi C31} &\rightarrow \\ \phi C31 &\rightarrow \\ \end{align}

Differential Equations

Applying mass action kinetic laws, we obtain the following set of differential equations. $$\begin{align*} \frac{d[LuxAHL]}{dt} &= k_{-RLux}[R_{Lux}]-k_{RLux}[LuxAHL][LuxR]-d_{LuxAHL}[LuxAHL]\\ \frac{d[LuxR]}{dt} &= \alpha_{LuxR} -k_{RLux}[LuxAHL][LuxR] + k_{-RLux}[RLux] - d_{LuxR}[LuxR] \\ \frac{d[RLux]}{dt} &= k_{RLux}[LuxAHL][LuxR] - k_{-RLux}[RLux] - 2 k_{DRLux} [RLux]^2 + 2 k_{-DRLux} [DRLux] - d_{RLux} [RLux] \\ \frac{d[DRLux]}{dt} &= k_{DRLux} [RLux]^2 - k_{-DRLux} [DRLux] - d_{DRLux} [DRLux] \\ \frac{d[P_{LuxON}]}{dt} &= k_{P_{LuxON}} [P_{LuxOFF}][DRLux] - k_{-P_{LuxON}} [P_{LuxON}]\\ \frac{d[mRNA_{Bxb1}]}{dt} &= L_{P_{Lux}} + k_{mRNA_{Bxb1}} [P_{LuxON}] - d_{mRNA_{Bxb1}} [mRNA_{Bxb1}]\\ \frac{d[Bxb1]}{dt} &= k_{Bxb1} [mRNA_{Bxb1}] - d_{Bxb1}[Bxb1]\\ \end{align*}$$

The same holds true for the Las system.

From the original set of reactions, we reduce the rate of production of mRNA_Bxb1 as a Hill function of RLux instead of Mass action kinetics in terms of P_LuxON and P_LuxOFF. For more information please check the characterization section.

Characterization: K_mLux and K_mLas

Data

For the Quorum sensing module we used established experimentally determined parameters for the rate of formation of RLux (reference). Since, in the literature the other parameters were estimated or fitted to their data, we decided to determine the parameters specific to our system. Hence, we used our data for the remaining parameters. Our data was mainly a transfer function of normalized GFP concentration as a function of input LuxAHL concentrations. (link to data)

Assumptions

Assumption A

We assumed that the dimerization of RLux to DRLux is quick. Quasi steady state approximation (QSSA) as follows

$$\frac{d[DRLux]}{dt} = k_{DRLux} [RLux]^2 - k_{-DRLux} [DRLux] - d_{DRLux} [DRLux] \approx 0\\$$

Assumption B

Further, from literature, we found that DRLux is specific to DNA and the dissociation constant is low (k_m = 0.1nM) {Reference}. Therefore, we using QSSA again,

$$\frac{d[P_{LuxON}]}{dt} = k_{P_{LuxON}} [P_{LuxOFF}][DRLux] - k_{-P_{LuxON}} [P_{LuxON}] \approx 0\\$$

Solving, we get the rate of production of mRNA_Bxb1 as

$$\frac{d[mRNA_{Bxb1}]}{dt} = L_{P_{Lux}} + \frac{k_{mRNA_{Bxb1}}[RLux]^2}{K_{mLux}^2 + [RLux]^2 }- d_{mRNA_{Bxb1}} [mRNA_{Bxb1}]\\$$

where

$$K_{mLux} = \sqrt{\frac {k_{-P_{LuxON}}}{k_{P_{LuxON}}}.\frac {k_{-DRLux} + d_{DRLux}}{k_{DRLux}}}$$

is a lumped parameter which we fitted to our data.

Similarly, lumped parameter K_mLas was derived for the las system and fitted to a transfer function of normalized GFP concentration as a function of input Las-AHL.

Parameter fitting

We used MEIGO Toolbox to fit the parameters to the experimental data. We used the concentrations at the end of five hours from each simulation and fit it to the experimental concentrations at the same time.

Lux QS Module fitted to experimental data from riboregulated Lux system.

Using the 'DHC' local-solver (Direct search method) in MEIGO, we found the lumped parameters $$K_{mLux} = 0.0124 nM$$

and

$$K_{mLas} = 0.3818 nM$$

respectively.

Las QS Module fitted to experimental data from riboregulated Las system.

Range of validity of the assumptions

These assumptions hold true for all input LuxAHL and LasAHL concentrations.

Leakiness

Cross-talk

Alternative Design