GENE REGULATION AND EXTRACYTOPLASMIC FUNCTION SIGMA FACTORS
Figure 1: General mechanism of ECF mode of action
Bacteria are constantly faced with fluctuating environmental conditions, which necessitate adequate and fast responses for survival. Such responses are mediated by one of four different signaling strategies:
(i) one-component systems (classical transcription factors);
(ii) two-component systems;
(iii) riboswitches and small regulatory RNAs;
(iv) alternative σ factors.
In addition to essential primary σ factors, which allow expression of housekeeping genes, most bacteria harbor tightly regulated and non-essential σ factors. Under inducing conditions, they are activated and replace the primary σ factor, thereby altering the promoter specificity of the RNA polymerase towards cognate alternative promoter sequences. Extracytoplasmic Function σ factors (ECFs) form the largest, functionally most diverse, yet structurally most simplistic, group of alternative σ factors.
While under regular growth conditions ECFs are kept inactive, they come into play once conditions change and they are activated. When active, these alternative σ factors shift the RNA polymerase holoenzyme pool towards specific promoters, ultimately activating the expression of gene products that mediate the adequate response to the trigger condition.
Our interest in this group of alternative σ factors falls into three aspects:
(i) ECF protein family classification
(ii) ECF activation mechanisms
(iii) ECFs as tools for the development of synthetic switches and circuits
ECF Projects
The genomic era has provided ample evidence that even simple prokaryotes are equipped with large numbers of signal transducing proteins that allow them to raise adequate responses to ever changing environmental conditions. After one- and two-component systems, ECFs represent the third fundamental mechanism of bacterial signal transduction. Comprehensive analyses of ECFs encoded in almost 500 genomes allowed the identification of 94 distinct groups The functional relevance of this classification is supported by sequence similarity and domain architecture of the cognate anti-σ factors, genomic context conservation and conservation of target promoter motifs.
Such analyses demonstrated the wide distribution, modular design and combinatorial complexity of ECF-dependent signal transduction and revealed unique features indicating novel and group specific mechanisms of ECF-mediated signal transduction. With this approach we aim at not only expand our knowledge of the diversity of ECFs but also provide clear and testable hypotheses about their mechanisms, directly feeding into both fundamental and applied research on ECF σ factors.
People
Franziska Dürr
Philipp Popp
Key publications
Pinto D., Mascher T. (2016) The ECF classification: a phylogenetic reflection of the regulatory diversity in the extracytoplasmic function sigma factor protein family. In: Stress and Environmental Regulation of Gene Expression and Adaptation in Bacteria, de Bruijn F.J. (ed.), John Wily & Sons, Inc., Chapter 2.6.
Pinto D., Mascher T. (2016) (Actino)Bacterial “intelligence”: using comparative genomics to unravel the information processing capacities of microbes. Curr Genet. [Pubmed]
Huang X., Pinto D., Fritz G., Mascher T. (2015) Environmental sensing in Actinobacteria: a comprehensive survey on the signaling capacity of this phylum. J Bacteriol 197:2517-35. [Pubmed]
Mascher T. (2013) Signaling diversity and evolution of extracytoplasmic function (ECF) σ factors. Curr Opin Microbiol 16:148-155. [Pubmed]
Jogler C., Waldmann J., Huang X., Jogler M., Glöckner F.O., Mascher T., Kolter R. (2012) Planctomycetes Comparative Genomics: Identification of Proteins Likely Involved in Morphogenesis, Cell Division and Signal Transduction. J. Bacteriol 194:6419-30. [Pubmed]
Staroń A, Sofia H.J., Dietrich S., Ulrich L.E., Liesegang H., Mascher T. (2009) The third pillar of bacterial signal transduction: classification of the extracytoplasmic function (ECF) sigma factor protein family. Mol Microbiol 74:557-81. [Pubmed]
Our work in accessing the diversity of extracytoplasmic function σ factors (ECFs) not only unraveled new ECF groups but also raised new hypotheses towards their signal transduction mechanisms. Thus far, extensive experimental evidence has accumulated about the proteolysis of the cognate anti-σ factors (ASFs), conformational changes of the ASFs, protein-protein interaction cascades, partner-switching and transcriptional activation by two-component systems. However, from the 94 groups described so far, only very few have been subject to detailed experimental investigations.
Our classification identified new ECF groups with a conserved C-terminal extension which lack obvious ASFs, indicating a regulatory role of these extensions and further suggesting a novel ECF σ factor activation mechanism. It is our goal to understand this new way of ECF-dependent signaling to ultimately exploit these regulatory network for more applied purposes. Our model organisms of choice are:
(i) Bacillus subtilis for the heterologous investigations of these ECFs due to its readily accessible genetics and of biotechnological relevance;
(ii) Streptomyces venezuelae for the analyses of these ECFs in their native environment because its genome encodes a very large number of ECFs suggesting that in this organisms a significant part of gene expression control is ECF-dependent.
People
Franziska Dürr
Key publications
Mascher T. (2013) Signaling diversity and evolution of extracytoplasmic function (ECF) σ factors. Curr Opin Microbiol 16:148-155 [Pubmed]
Wecke T., Halang P., Staroń A., Dufour Y. S., Donohue T. J., Mascher T. (2012). Extracytoplasmic function σ factors of the widely distributed group ECF41 contain a fused regulatory domain. MicrobiologyOpen 1:194-213. [Pubmed]
One of the major goals of Synthetic Biology is the functional implementation of novel metabolic pathways into new organisms. Engineered microorganisms are already used in the production of complex pharmaceutical compounds that are beyond the reach of existing approaches in (bio)chemical synthesis. Moreover, other Synthetic Biology applications are the development of biological delivery systems for therapy, the environmentally friendly production of chemicals as well as bioremediation and energy production.
An essential prerequisite for complex manipulations of microorganisms is the availability of large sets of well-evaluated expression parts and devices to orchestrate the underlying in vivo expression programs. The currently available toolbox is far from being comprehensive and is often based on modifications of the same few regulatory principles. ECFs have barely been explored so far despite the fact that their features – diversity, conservation, ubiquity, modularity and orthogonality – make them ideal candidates for designing switches and expression devices.
Our goal is to make use of the wealth of available information on ECFs to implement novel ECF-based switches and circuits and greatly improve the available toolbox for manipulation of B. subtilis.
People
Franziska Dürr