Part:BBa_I1051

Lux cassette right promoter

The lux cassette of V. fischeri contains a left and a right promoter. The right promoter gives weak constitutive expression of downstream genes.This expression is up-regulated by the action of the Lux repressor, LuxR. Two molecules of LuxR protein form a complex with two molecules the signalling compound homoserine lactone (HSL). This complex binds to a palindromic site on the promoter, increasing the rate of transcription.In thisversion, a binding site for cI dimer has been added: binding to this site dominantly represses transcription.

Usage and Biology

Unspecified.

Sequence and Features

Assembly Compatibility:

• 10
  COMPATIBLE WITH RFC[10]
• 12
  COMPATIBLE WITH RFC[12]
• 21
  COMPATIBLE WITH RFC[21]
• 23
  COMPATIBLE WITH RFC[23]
• 25
  COMPATIBLE WITH RFC[25]
• 1000
  COMPATIBLE WITH RFC[1000]

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Contribution by the NYU Abu Dhabi iGEM Team

Written by Zerina Rahic

Introduction

Lux cassette is a unique member of the TetR superfamily of transcription factors [1]. Due to their ability to activate and repress large regulons of genes, bacterial Lux system is used as gene expression reporter and regulators for quorum sensing [1]. Benefits of using this system is its speed, sensitivity and non-destructivity [2]. However, due to its nonlinear molecular and enzyme dynamics, Lux cassette’s bioluminescence is different from gene expression [2]. This causes limitations in bioluminescent data interpretation.

Approach

According to research done by Mudassar Iqbal in 2017, computational approaches could enable researchers to infer gene transcription levels of Lux cassette right promoters. Previously, bioluminescent profiles resulted in transient peaks of light output being different in experimental data sets compared to gene expression [2]. Therefore, using Bayesian inference schemes, researchers are able to reverse engineer promoter activity from the bioluminescence [2].

Mathematical Model

Figure 1. Mathematical model for LUX system

Using this mathematical approach, researchers were able infer LUX right promoter activity from light readout. The Lux right promoter input function is modeled as a series of K heights at fixed positions. Using a model proposed by Green in1995, researchers were able to measure the distribution for each height at point n with mean value equal to the current height at n-1. Points are chosen at random and new heights are proposed accordingly [2]. Using the Gaussian error model, a likelihood function is proposed [2]. This approach is crucial as it enables normalization of light values for the Lux system including the Lux right promoter [2].

Discovery

Data from this research provides evidence that decrease in bioluminescence can be interpreted as a switching off of the promoter, while an increase in bioluminescence would be better interpreted as a longer period of gene expression [2].

References

1. Van Kessel, J. C., Ulrich, L. E., Zhulin, I. B., & Bassler, B. L. (2013). Analysis of activator and repressor functions reveals the requirements for transcriptional control by LuxR, the master regulator of quorum sensing in Vibrio harveyi. MBio, 4(4), e00378-13.
2. Iqbal, M., Doherty, N., Page, A. M., Qazi, S. N., Ajmera, I., Lund, P. A., ... & Stekel, D. J. (2017). Reconstructing promoter activity from Lux bioluminescent reporters. PLoS computational biology, 13(9), e1005731.