Computational tools and Interoperability in Comparative ... - CBS

Chapter 3
rRNA operons and promoter analysis
rRNA operons and promoter
analysis
3.1 Introduction
This chapter covers two papers (VI and VII), dealing with rRNA localization within the
genome, and analysis of the promoter region upstream of rRNA operons. The RNAmmer
tool (Lagesen et al., 2007) presented in paper VI was motivated by the lack of a software
tools that was able to accurately and consistently annotate ribosomal RNA (rRNA) genes
in prokaryotes. BLAST strategies are widely used for this as the rRNA genes are highly
conserved. However, homology search methods produces often less accurate gene boundaries
as they fail to account for the observed variation in some regions. Hidden Markov
Model (HMM) strategies, such as RNAmmer, can take into account conserved stem loop
structures, greatly improving the accuracy of prediction of the full length rRNA genes.
Particular detail will be given to the E. coli rRNA operons in terms of promoter predictions,
since much experimental information is known about this system. An application
of the gwBrowser as a tool for visualization of promoter regions upstream of the rRNA
operons in E. coli concludes the chapter. The gwBrowser effort is currently being published
in the Standards In Genomic Sciences journal. The P1 and P2 prediction tools are
still developmental, and have not been published.
Encoding the central structure of the ribosome, the 5S, 16S, and 23S rRNA genes are
essential for protein synthesis and are transcribed at high levels. In E. coli the rrn operons
are regulated by a tandem promotor system. With abundant transcription, the system is
favorable for studying the mechanisms of highly expressed genes and establish connection
to the physical properties of the DNA. In this work, the SIDD energy (Wang et al., 2004;
Wang & Benham, 2008) was used to measure the energy requirement to melt the DNA
helix near the promotor region. The work was carried out during my visit to Professor
Craig Benhams lab at UC Davis, fall 2007.
3.2 P1 and P2 promoters in E. coli
The seven rRNA operons of E. coli are regulated by the two promotors P1 and P2,
where P1 is active predominately during exponential growth whereas P2 is active during
stationay phase (Hirvonen et al., 2001; Murray & Gourse, 2004). Apart from the –10 and
–35 hexamers, the P1 site contains between 3 and 5 FIS (Factor for Inversion Stimulation)
binding sites and an UP element. FIS has been reported to increase the transcription in
vivo by 4-10 fold in this system (Bokal et al., 1995).
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