Expansion and re-classification of the extracytoplasmic function (ECF) σ factor family

Abstract

Extracytoplasmic function σ factors (ECFs) represent one of the major bacterial signal transduction mechanisms in terms of abundance, diversity and importance, particularly in mediating stress responses. Here, we performed a comprehensive phylogenetic analysis of this protein family by scrutinizing all proteins in the NCBI database. As a result, we identified an average of ∼10 ECFs per bacterial genome and 157 phylogenetic ECF groups that feature a conserved genetic neighborhood and a similar regulation mechanism. Our analysis expands previous classification efforts ∼50-fold, enriches many original ECF groups with previously unclassified proteins and identifies 22 entirely new ECF groups. The ECF groups are hierarchically related to each other and are further composed of subgroups with closely related sequences. This two-tiered classification allows for the accurate prediction of common promoter motifs and the inference of putative regulatory mechanisms across subgroups composing an ECF group. This comprehensive, high-resolution description of the phylogenetic distribution of the ECF family, together with the massive expansion of classified ECF sequences and an openly accessible data repository called ‘ECF Hub’ (https://www.computational.bio.uni-giessen.de/ecfhub), will serve as a powerful hypothesis-generator to guide future research in the field.

REC protein family expansion by the emergence of a new signaling pathway

“…NtrC-like RRs require sigma54 transcription factors to activate genes in their regulons, whereas NarL-like RRs interact with sigma70. It is well documented that cellular levels of small nucleotide, (p)ppGpp, in combination with the protein DksA heavily regulate RNA polymerase's (RNAP) preference for sigma70 (housekeeping or extracytoplasmic function (ECF) family) (Casas-Pastor et al, 2019;Lee et al, 2012;Lonetto et al, 1998) or sigma54 (Bernardo et al, 2006(Bernardo et al, , 2009Dalebroux and Swanson, 2012;Jurado et al, 2003;Ronneau and Hallez, 2019;Wigneshweraraj et al, 2008). While the exact environmental conditions needed for pseudomonads to reach the appropriate levels of (p)ppGpp and DksA for sigma54 occupation of RNAP are unknown (Potvin et al, 2008;Ronneau and Hallez, 2019;Shingler, 2011), it can be reasoned that the switch is lifestyle dependent.…”

Extracytoplasmic Function σ Factors as Tools for Coordinating Stress Responses

“…The advent of bacterial genome sequencing and annotation projects has led to a great increase in the number of available sequences in the databases, which has helped mitigate the former bias toward highly represented phyla. In its latest version [56], the classification has undergone an important reorganization and expansion, increasing its accuracy, mainly due to the increase in the number of analyzed genomes, and organizing the ECFs into monophyletic groups within the ECF phylogenetic group. At the time when the analysis was performed (2017), more than 180,000 bacterial genomes were available at the National Center for Biotechnology Information (NCBI) database and were used as an initial dataset.…”

“…Despite the classification being based on sequence similarity and genomic context of ECFs, nevertheless, due to their degree of conservation, target promoter motifs and potential functions can be inferred for each distinct group with a high level of accuracy based on the few experimentally studied representatives. Even in groups with no experimentally analyzed representatives, regulatory mechanisms and target motifs may be predicted based on the genomic context and the amino acid composition of the DNA-binding domains compared to other ECFs [56]. Regarding their function, although ECFs are not classified based on it, their conservation also allows inferences to be made regarding the physiological processes in which they are involved, again based on the studies of a number of representatives.…”

“…For instance, some members of the ECF41 are known to be involved in the protection against oxidative stress [57], but other representatives of this group do not participate in that response, or any other known so far [58]. Reviewing the latest ECF classification [56] and based on the manually studied examples of each group (which are also provided in the classification), we have attempted to cluster them function-wise into three broadly related classes: Class (I) are ECFs involved in responding to external or internal cues that may be directly deleterious to maintaining cell integrity and homeostasis. These include responses such as resistance to abiotic stress, to development, virulence and interaction with host organisms; Class (II) are ECFs involved in responding to metabolic and nutritional cues, including nutrient limitation, transport and biosynthesis; and Class (III), in which there are groups including those of unknown function.…”

“…Since the majority of the members of each ECF group remain unstudied, it is possible that some of their specific functions may not fit precisely with the broad functions assigned in this classification. However, it gives an idea of the importance of ECFs for stress responses and may serve as a starting point to tackle the characterization of newly studied ECFs in combination with the additional information provided in the ECF classification [56].…”