A MoClo-Compatible Toolbox of ECF Sigma Factor-Based Regulatory Switches for Proteobacterial Chassis.

The construction of complex synthetic gene circuits with predetermined and reliable output depends on orthogonal regulatory parts that do not inadvertently interfere with the host machinery or with other circuit components. Previously, extracytoplasmic function sigma factors (ECFs), a diverse group of alternative sigma factors with distinct promoter specificities, were shown to have great potential as context-independent regulators, but so far, they have only been used in a few model species. Here, we show that the alphaproteobacterium , which has been proposed as a plant-associated bacterial chassis for synthetic biology, has a similar phylogenetic ECF acceptance range as the gammaproteobacterium .

T-bet B cells are activated by and control endogenous retroviruses through TLR-dependent mechanisms.

Endogenous retroviruses (ERVs) are an integral part of the mammalian genome. The role of immune control of ERVs in general is poorly defined as is their function as anti-cancer immune targets or drivers of autoimmune disease. Here, we generate mouse-strains where Moloney-Murine Leukemia Virus tagged with GFP (ERV-GFP) infected the mouse germline.

Proteinortho6: pseudo-reciprocal best alignment heuristic for graph-based detection of (co-)orthologs.

Proteinortho is a widely used tool to predict (co)-orthologous groups of genes for any set of species. It finds application in comparative and functional genomics, phylogenomics, and evolutionary reconstructions. With a rapidly increasing number of available genomes, the demand for large-scale predictions is also growing.

Processing and decay of 6S-1 and 6S-2 RNAs in .

Noncoding 6S RNAs regulate transcription by binding to the active site of bacterial RNA polymerase holoenzymes. Processing and decay of 6S-1 and 6S-2 RNA were investigated in by northern blot and RNA-seq analyses using different RNase knockout strains, as well as by in vitro processing assays. For both 6S RNA paralogs, we identified a key-but mechanistically different-role of RNase J1.

Evolutionary reconstruction, nomenclature and functional meta-analysis of the Kiwellin protein family.

Crop diseases caused by pathogens critically affect global food security and plant ecology. Pathogens are well adapted to their host plants and have developed sophisticated mechanisms allowing successful colonization. Plants in turn have taken measures to counteract pathogen attacks resulting in an evolutionary arms race.

The Vibrio vulnificus stressosome is an oxygen-sensor involved in regulating iron metabolism.

Stressosomes are stress-sensing protein complexes widely conserved among bacteria. Although a role in the regulation of the general stress response is well documented in Gram-positive bacteria, the activating signals are still unclear, and little is known about the physiological function of stressosomes in the Gram-negative bacteria. Here we investigated the stressosome of the Gram-negative marine pathogen Vibrio vulnificus.

Structural and Functional Insight into the Mechanism of 6S-1 RNA Release from RNA Polymerase.

Here we investigated the refolding of 6S-1 RNA and its release from σ-RNA polymerase (σ-RNAP) in vitro using truncated and mutated 6S-1 RNA variants. Truncated 6S-1 RNAs, only consisting of the central bubble (CB) flanked by two short helical arms, can still traverse the mechanistic 6S RNA cycle in vitro despite ~10-fold reduced σ-RNAP affinity. This indicates that the RNA's extended helical arms including the '-35'-like region are not required for basic 6S-1 RNA functionality.

Identification and characterization of short leader and trailer RNAs synthesized by the Ebola virus RNA polymerase.

Transcription of non-segmented negative sense (NNS) RNA viruses follows a stop-start mechanism and is thought to be initiated at the genome's very 3'-end. The synthesis of short abortive leader transcripts (leaderRNAs) has been linked to transcription initiation for some NNS viruses. Here, we identified the synthesis of abortive leaderRNAs (as well as trailer RNAs) that are specifically initiated opposite to (anti)genome nt 2; leaderRNAs are predominantly terminated in the region of nt ~ 60-80.

Insights into 6S RNA in lactic acid bacteria (LAB).

Background: 6S RNA is a regulator of cellular transcription that tunes the metabolism of cells. This small non-coding RNA is found in nearly all bacteria and among the most abundant transcripts. Lactic acid bacteria (LAB) constitute a group of microorganisms with strong biotechnological relevance, often exploited as starter cultures for industrial products through fermentation.

iCLIP analysis of RNA substrates of the archaeal exosome.

Background: The archaeal exosome is an exoribonucleolytic multiprotein complex, which degrades single-stranded RNA in 3' to 5' direction phosphorolytically. In a reverse reaction, it can add A-rich tails to the 3'-end of RNA. The catalytic center of the exosome is in the aRrp41 subunit of its hexameric core.

6S-2 RNA deletion in the undomesticated strain NCIB 3610 causes a biofilm derepression phenotype.

Bacterial 6S RNA regulates transcription via binding to the active site of RNA polymerase holoenzymes. 6S RNA has been identified in the majority of bacteria, in most cases encoded by a single gene. Firmicutes including encode two 6S RNA paralogs, 6S-1 and 6S-2 RNA.

Tol-Pal System and Rgs Proteins Interact to Promote Unipolar Growth and Cell Division in Sinorhizobium meliloti.

is an alphaproteobacterium belonging to the Bacteria from this order elongate their cell wall at the new cell pole, generated by cell division. Screening for protein interaction partners of the previously characterized polar growth factors RgsP and RgsM, we identified the inner membrane components of the Tol-Pal system (TolQ and TolR) and novel Rgs (rhizobial growth and septation) proteins with unknown functions. TolQ, Pal, and all Rgs proteins, except for RgsE, were indispensable for cell growth.

The two paralogous kiwellin proteins KWL1 and KWL1-b from maize are structurally related and have overlapping functions in plant defense.

Many plant-pathogenic bacteria and fungi deploy effector proteins that down-regulate plant defense responses and reprogram plant metabolism for colonization and survival Kiwellin (KWL) proteins are a widespread family of plant-defense proteins that target these microbial effectors. The KWL1 protein from maize (corn, ) specifically inhibits the enzymatic activity of the secreted chorismate mutase Cmu1, a virulence-promoting effector of the smut fungus In addition to KWL1, 19 additional KWL paralogs have been identified in maize. Here, we investigated the structure and mechanism of the closest KWL1 homolog, KWL1-b (ZEAMA_GRMZM2G305329).

Noncoding RNA is an integral component of the TLR4-TRIF pathway.

RNA has been proposed as an important scaffolding factor in the nucleus, aiding protein complex assembly in the dense intracellular milieu. Architectural contributions of RNA to cytosolic signaling pathways, however, remain largely unknown. Here, we devised a multidimensional gradient approach, which systematically locates RNA components within cellular protein networks.

Sequence and structural properties of circular RNAs in the brain of nurse and forager honeybees (Apis mellifera).

Background: The honeybee (Apis mellifera) represents a model organism for social insects displaying behavioral plasticity. This is reflected by an age-dependent task allocation. The most protruding tasks are performed by young nurse bees and older forager bees that take care of the brood inside the hive and collect food from outside the hive, respectively.

Minimal and RNA-free RNase P in .

RNase P is an essential tRNA-processing enzyme in all domains of life. We identified an unknown type of protein-only RNase P in the hyperthermophilic bacterium : Without an RNA subunit and the smallest of its kind, the 23-kDa polypeptide comprises a metallonuclease domain only. The protein has RNase P activity in vitro and rescued the growth of and strains with inactivations of their more complex and larger endogenous ribonucleoprotein RNase P.

6S RNA in Rhodobacter sphaeroides: 6S RNA and pRNA transcript levels peak in late exponential phase and gene deletion causes a high salt stress phenotype.

The function of 6S RNA, a global regulator of transcription, was studied in the photosynthetic α-proteobacterium Rhodobacter sphaeroides. The cellular levels of R. sphaeroides 6S RNA peak toward the transition to stationary phase and strongly decrease during extended stationary phase.

Protein-only RNase P function in Escherichia coli: viability, processing defects and differences between PRORP isoenzymes.

The RNase P family comprises structurally diverse endoribonucleases ranging from complex ribonucleoproteins to single polypeptides. We show that the organellar (AtPRORP1) and the two nuclear (AtPRORP2,3) single-polypeptide RNase P isoenzymes from Arabidopsis thaliana confer viability to Escherichia coli cells with a lethal knockdown of its endogenous RNA-based RNase P. RNA-Seq revealed that AtPRORP1, compared with bacterial RNase P or AtPRORP3, cleaves several precursor tRNAs (pre-tRNAs) aberrantly in E.

The Conserved Dcw Gene Cluster of R. sphaeroides Is Preceded by an Uncommonly Extended 5' Leader Featuring the sRNA UpsM.

Cell division and cell wall synthesis mechanisms are similarly conserved among bacteria. Consequently some bacterial species have comparable sets of genes organized in the dcw (division and cell wall) gene cluster. Dcw genes, their regulation and their relative order within the cluster are outstandingly conserved among rod shaped and gram negative bacteria to ensure an efficient coordination of growth and division.

A genomic view of food-related and probiotic Enterococcus strains.

The study of enterococcal genomes has grown considerably in recent years. While special attention is paid to comparative genomic analysis among clinical relevant isolates, in this study we performed an exhaustive comparative analysis of enterococcal genomes of food origin and/or with potential to be used as probiotics. Beyond common genetic features, we especially aimed to identify those that are specific to enterococcal strains isolated from a certain food-related source as well as features present in a species-specific manner.

Identification of specific residues in avian influenza A virus NS1 that enhance viral replication and pathogenicity in mammalian systems.

Reassortment of their segmented genomes allows influenza A viruses (IAV) to gain new characteristics, which potentially enable them to cross the species barrier and infect new hosts. Improved replication was observed for reassortants of the strictly avian IAV A/FPV/Rostock/34 (FPV, H7N1) containing the NS segment from A/Goose/Guangdong/1/1996 (GD, H5N1), but not for reassortants containing the NS segment of A/Mallard/NL/12/2000 (MA, H7N3). The NS1 of GD and MA differ only in 8 aa positions.

Bacillus subtilis 6S-2 RNA serves as a template for short transcripts in vivo.

The global transcriptional regulator 6S RNA is abundant in a broad range of bacteria. The RNA competes with DNA promoters for binding to the housekeeping RNA polymerase (RNAP) holoenzyme. When bound to RNAP, 6S RNA serves as a transcription template for RNAP in an RNA-dependent RNA polymerization reaction.

Positive Selection or Free to Vary? Assessing the Functional Significance of Sequence Change Using Molecular Dynamics.

Evolutionary arms races between pathogens and their hosts may be manifested as selection for rapid evolutionary change of key genes, and are sometimes detectable through sequence-level analyses. In the case of protein-coding genes, such analyses frequently predict that specific codons are under positive selection. However, detecting positive selection can be non-trivial, and false positive predictions are a common concern in such analyses.

Distribution of Ribonucleoprotein and Protein-Only RNase P in Eukarya.

RNase P is the endonuclease that removes 5' leader sequences from tRNA precursors. In Eukarya, separate RNase P activities exist in the nucleus and mitochondria/plastids. Although all RNase P enzymes catalyze the same reaction, the different architectures found in Eukarya range from ribonucleoprotein (RNP) enzymes with a catalytic RNA and up to 10 protein subunits to single-subunit protein-only RNase P (PRORP) enzymes.