microLife最新文献

Second messengers transfer signals from changing intra- and extracellular conditions to a cellular response. Over the last few decades, several nucleotide-based second messengers have been identified and characterized in especially bacteria and eukaryotes. Also in archaea, several nucleotide-based second messengers have been identified. This review will summarize our understanding of nucleotide-based second messengers in archaea. For some of the nucleotide-based second messengers, like cyclic di-AMP and cyclic oligoadenylates, their roles in archaea have become clear. Cyclic di-AMP plays a similar role in osmoregulation in euryarchaea as in bacteria, and cyclic oligoadenylates are important in the Type III CRISPR-Cas response to activate CRISPR ancillary proteins involved in antiviral defense. Other putative nucleotide-based second messengers, like 3',5'- and 2',3'-cyclic mononucleotides and adenine dinucleotides, have been identified in archaea, but their synthesis and degradation pathways, as well as their functions as secondary messengers, still remain to be demonstrated. In contrast, 3'-3'-cGAMP has not yet been identified in archaea, but the enzymes required to synthesize 3'-3'-cGAMP have been found in several euryarchaeotes. Finally, the widely distributed bacterial second messengers, cyclic diguanosine monophosphate and guanosine (penta-)/tetraphosphate, do not appear to be present in archaea.

The stringent response and its signalling nucleotides, pppGpp and ppGpp, have been the subject of intense research since the discovery of (p)ppGpp in 1969. Recent studies have revealed that the downstream events that follow (p)ppGpp accumulation vary among species. Consequently, the stringent response as initially characterized in Escherichia coli largely differs from the response in Firmicutes (Bacillota), wherein synthesis and degradation of the messengers (p)ppGpp are orchestrated by the bifunctional Rel enzyme with synthetase and hydrolase activity and the two synthetases SasA/RelP and SasB/RelQ. Here we will summarize recent studies supporting the role of (p)ppGpp in the development of antibiotic resistance and tolerance as well as survival under adverse environmental conditions in Firmicutes. We will also discuss the impact of elevated (p)ppGpp levels on the development of persister cells and the establishment of persistent infections. (p)ppGpp levels are usually tightly controlled to allow optimal growth under non-stressed conditions. Upon the onset of certain 'stringent conditions' the sudden increase in (p)ppGpp levels limits growth while exerting protective effects. In Firmicutes, the (p)ppGpp-mediated restriction of GTP accumulation is one major mechanism of protection and survival under stresses such as antibiotic exposure.

The soil-dwelling plant symbiont Sinorhizobium meliloti is a major model organism of Alphaproteobacteria. Despite numerous detailed OMICS studies, information about small open reading frame (sORF)-encoded proteins (SEPs) is largely missing, because sORFs are poorly annotated and SEPs are hard to detect experimentally. However, given that SEPs can fulfill important functions, identification of translated sORFs is critical for analyzing their roles in bacterial physiology. Ribosome profiling (Ribo-seq) can detect translated sORFs with high sensitivity, but is not yet routinely applied to bacteria because it must be adapted for each species. Here, we established a Ribo-seq procedure for S. meliloti 2011 based on RNase I digestion and detected translation for 60% of the annotated coding sequences during growth in minimal medium. Using ORF prediction tools based on Ribo-seq data, subsequent filtering, and manual curation, the translation of 37 non-annotated sORFs with ≤ 70 amino acids was predicted with confidence. The Ribo-seq data were supplemented by mass spectrometry (MS) analyses from three sample preparation approaches and two integrated proteogenomic search database (iPtgxDB) types. Searches against standard and 20-fold smaller Ribo-seq data-informed custom iPtgxDBs confirmed 47 annotated SEPs and identified 11 additional novel SEPs. Epitope tagging and Western blot analysis confirmed the translation of 15 out of 20 SEPs selected from the translatome map. Overall, by combining MS and Ribo-seq approaches, the small proteome of S. meliloti was substantially expanded by 48 novel SEPs. Several of them are part of predicted operons and/or are conserved from Rhizobiaceae to Bacteria, suggesting important physiological functions.

In this short piece, I connect the dots between the pervasive influence of microbial activities on our health and that of the planet, including their positive and negative roles in current polycrises, our ability to influence microbes to promote their positive influences and mitigate their negative impacts, the roles of everyone as stewards and stakeholders in personal, family, community, national, and global wellbeing, the need for stewards and stakeholders to possess relevant information in order to fulfil their roles and obligations, and the compelling case for microbiology literacy and introduction of a societally relevant microbiology curriculum in school.

In contrast to extensively studied prokaryotic 'small' transcriptomes (encompassing all small noncoding RNAs), small proteomes (here defined as including proteins ≤70 aa) are only now entering the limelight. The absence of a complete small protein catalogue in most prokaryotes precludes our understanding of how these molecules affect physiology. So far, archaeal genomes have not yet been analyzed broadly with a dedicated focus on small proteins. Here, we present a combinatorial approach, integrating experimental data from small protein-optimized mass spectrometry (MS) and ribosome profiling (Ribo-seq), to generate a high confidence inventory of small proteins in the model archaeon Haloferax volcanii. We demonstrate by MS and Ribo-seq that 67% of the 317 annotated small open reading frames (sORFs) are translated under standard growth conditions. Furthermore, annotation-independent analysis of Ribo-seq data showed ribosomal engagement for 47 novel sORFs in intergenic regions. A total of seven of these were also detected by proteomics, in addition to an eighth novel small protein solely identified by MS. We also provide independent experimental evidence in vivo for the translation of 12 sORFs (annotated and novel) using epitope tagging and western blotting, underlining the validity of our identification scheme. Several novel sORFs are conserved in Haloferax species and might have important functions. Based on our findings, we conclude that the small proteome of H. volcanii is larger than previously appreciated, and that combining MS with Ribo-seq is a powerful approach for the discovery of novel small protein coding genes in archaea.

Microbial taxonomy is critical for describing ecosystem composition, yet the link between taxonomy and properties of microbes, such as their cellular architecture, remains poorly defined. We hypothesized that the cellular architecture represents microbial niche adaptation. We used cryo-electron microscopy and tomography to analyze microbial morphology in order to associate cellular architecture with phylogeny and genomic contents. As a model system, we chose the core rumen microbiome and imaged a large isolate collection covering 90% of its richness at the order level. Based on quantifications of several morphological features, we found that the visual similarity of microbiota is significantly related to their phylogenetic distance. Up to the Family level, closely related microbes have similar cellular architectures, which are highly correlated with genome similarity. However, in more distantly related bacteria, the correlation both with taxonomy and genome similarity is lost. This is the first comprehensive study of microbial cellular architecture and our results highlight that structure remains an important parameter in classification of microorganisms, along with functional parameters such as metabolomics. Furthermore, the high-quality images presented in this study represent a reference database for the identification of bacteria in anaerobic ecosystems.

The formation of plaques represents the hallmark of phage infection visualizing the clearance of the bacterial lawn in structured environments. In this study, we have addressed the impact of cellular development on phage infection in Streptomyces undergoing a complex developmental life cycle. Analysis of plaque dynamics revealed, after a period of plaque size enlargement, a significant regrowth of transiently phage-resistant Streptomyces mycelium into the lysis zone. Analysis of Streptomyces venezuelae mutant strains defective at different stages of cellular development indicated that this regrowth was dependent on the onset of the formation of aerial hyphae and spores at the infection interface. Mutants restricted to vegetative growth (ΔbldN) featured no significant constriction of plaque area. Fluorescence microscopy further confirmed the emergence of a distinct zone of cells/spores with reduced cell permeability towards propidium iodide staining at the plaque periphery. Mature mycelium was further shown to be significantly less susceptible to phage infection, which is less pronounced in strains defective in cellular development. Transcriptome analysis revealed the repression of cellular development at the early stages of phage infection probably facilitating efficient phage propagation. We further observed an induction of the chloramphenicol biosynthetic gene cluster highlighting phage infection as a trigger of cryptic metabolism in Streptomyces. Altogether, our study emphasizes cellular development and the emergence of transient phage resistance as an important layer of Streptomyces antiviral immunity.

Adult humans harbor at least as many microbial cells as eukaryotic ones. The largest compartment of this diverse microbial population, the gut microbiota, encompasses the collection of bacteria, archaea, viruses, and eukaryotic organisms that populate the gastrointestinal tract, and represents a complex and dynamic ecosystem that has been increasingly implicated in health and disease. The gut microbiota carries ∼100-to-150-times more genes than the human genome and is intimately involved in development, homeostasis, and disease. Of the several microbial metabolites that have been studied, short-chain fatty acids emerge as a group of molecules that shape gene expression in several types of eukaryotic cells by multiple mechanisms, which include DNA methylation changes, histone post-translational modifications, and microRNA-mediated gene silencing. Butyric acid, one of the most extensively studied short-chain fatty acids, reaches higher concentrations in the colonic lumen, where it provides a source of energy for healthy colonocytes, and its concentrations decrease towards the bottom of the colonic crypts, where stem cells reside. The lower butyric acid concentration in the colonic crypts allows undifferentiated cells, such as stem cells, to progress through the cell cycle, pointing towards the importance of the crypts in providing them with a protective niche. In cancerous colonocytes, which metabolize relatively little butyric acid and mostly rely on glycolysis, butyric acid preferentially acts as a histone deacetylase inhibitor, leading to decreased cell proliferation and increased apoptosis. A better understanding of the interface between the gut microbiota metabolites and epigenetic changes in eukaryotic cells promises to unravel in more detail processes that occur physiologically and as part of disease, help develop novel biomarkers, and identify new therapeutic modalities.

Bacterial membrane vesicles (MVs) are abundant in the oceans, but their potential functional roles remain unclear. In this study we characterized MV production and protein content of six strains of Alteromonas macleodii, a cosmopolitan marine bacterium. Alteromonas macleodii strains varied in their MV production rates, with some releasing up to 30 MVs per cell per generation. Microscopy imaging revealed heterogenous MV morphologies, including some MVs aggregated within larger membrane structures. Proteomic characterization revealed that A. macleodii MVs are rich in membrane proteins related to iron and phosphate uptake, as well as proteins with potential functions in biofilm formation. Furthermore, MVs harbored ectoenzymes, such as aminopeptidases and alkaline phosphatases, which comprised up to 20% of the total extracellular enzymatic activity. Our results suggest that A. macleodii MVs may support its growth through generation of extracellular 'hotspots' that facilitate access to essential substrates. This study provides an important basis to decipher the ecological relevance of MVs in heterotrophic marine bacteria.