Experimental and computational validation of models of fluorescent and luminescent reporter genes in bacteria.
BACKGROUND
Fluorescent and luminescent reporter genes have become popular tools for the real-time monitoring of gene expression in living cells. However, mathematical models are necessary for extracting biologically meaningful quantities from the primary data.


RESULTS
We present a rigorous method for deriving relative protein synthesis rates (mRNA concentrations) and protein concentrations by means of kinetic models of gene expression. We experimentally and computationally validate this approach in the case of the protein Fis, a global regulator of transcription in Escherichia coli. We show that the mRNA and protein concentration profiles predicted from the models agree quite well with direct measurements obtained by Northern and Western blots, respectively. Moreover, we present computational procedures for taking into account systematic biases like the folding time of the fluorescent reporter protein and differences in the half-lives of reporter and host gene products. The results show that large differences in protein half-lives, more than mRNA half-lives, may be critical for the interpretation of reporter gene data in the analysis of the dynamics of regulatory systems.


CONCLUSIONS
The paper contributes to the development of sound methods for the interpretation of reporter gene data, notably in the context of the reconstruction and validation of models of regulatory networks. The results have wide applicability for the analysis of gene expression in bacteria and may be extended to higher organisms.
de Jong
Hidde
H
Institut Jean Roget, LAPM, UMR5163, Campus Santé, Université Joseph Fourier, Domaine de la Merci, 38700 La Tronche, France.
Ranquet
Caroline
C
Ropers
Delphine
D
Pinel
Corinne
C
Geiselmann
Johannes
J
D000818:Animals; D019295:Computational Biology / Q000379:methods; D004926:Escherichia coli / Q000378:metabolism*; D020869:Gene Expression Profiling; D015964:Gene Expression Regulation, Bacterial; D017930:Genes, Reporter; D049452:Green Fluorescent Proteins / Q000378:metabolism; D007700:Kinetics; D049449:Luminescence; D008856:Microscopy, Fluorescence / Q000379:methods; D008957:Models, Genetic; D010957:Plasmids / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D014158:Transcription, Genetic
D012333:RNA, Messenger; D049452:Green Fluorescent Proteins