Comprehensive phylogenetic study of ECF sigma factors
Extracytoplasmic function (ECF) σ factors are the most minimalistic member of the σ70 family. ECFs and their activity regulators are one of the main signal transduction mechanisms that allow bacteria to respond to extracellular changes. Aside from their natural role in bacterial homeostasis, ECFs ar...
|Online Access:||PDF Full Text|
No Tags, Be the first to tag this record!
|Summary:||Extracytoplasmic function (ECF) σ factors are the most minimalistic member of the σ70 family. ECFs and their activity regulators are one of the main signal transduction mechanisms that allow bacteria to respond to extracellular changes. Aside from their natural role in bacterial homeostasis, ECFs are generally host independent and functionally orthogonal, which makes them especially attractive for constructing bacterial synthetic circuits. In silico identification of sets of ECFs, their target promoters and their regulators is particularly simple since ECFs and their regulators are typically encoded in the same genetic neighborhood and usually in the same operon, and ECFs usually target their own promoter. Earlier works on the phylogenetic classification of ECFs revealed that there is a correlation between ECF groups, which harbor proteins with a similar sequence, regulator type and target promoter motif elements. This showed that the phylogenetic classification of ECFs is essential to understand their modes of regulation. The large number of sequenced bacterial genomes currently deposited in databases suggests that an ECF reclassification would expand our knowledge on ECF regulation. This thesis addresses the analysis of the main modes of regulation found in the comprehensive classification of ECF σ factor subfamily.
For this study, I first extracted ECFs from all bacterial genomes deposited in NCBI. I identified more than 170,000 unique protein sequences that are likely to function as ECFs. This resulted in a 50-fold expansion over the original ECF library. Then, I classified the conserved σ domains of these proteins into more than 150 phylogenetic groups, each associated to a conserved type of regulator. I systematically described each ECF group in terms of its putative regulator, putative target promoter, taxonomic distribution and putative function. I confirmed these predictions for groups with described members. Anti-σ factors are the main type of ECF regulator across groups, followed by C-terminal extensions of their protein and serine/threonine kinases, which have been suggested to phosphorylate ECFs. I hypothesized new alternative types of regulators for some ECF groups.
Using a combination of bioinformatic tools and collaborating with different experimental research groups, I focused on the most important regulatory elements of ECFs to shed light into their mechanism of regulation. In the case of anti-σ factors, I focused on their most common type, class I anti-σ factors, to reveal two shared binding interfaces between ECFs and these inhibitors. Then, I focused on the three largest ECF groups associated to C-terminal extensions, showing a different role of this additional region in the control of ECF activity in the different groups. Lastly, I focused on serine/threonine kinases to find that phosphorylation compensates for the lack of negative charges in one of the main RNA polymerase binding surfaces of ECF σ factors.
In summary, this thesis provides the scientific community with a comprehensive overview of ECF σ factor regulation, target promoter and function across phylogenetic groups, and sheds light into some of their most important regulatory mechanisms.|
|Physical Description:||217 Pages|