Short prokaryotic Argonaute systems trigger cell death upon detection of invading DNA.

Argonaute proteins use single-stranded RNA or DNA guides to target complementary nucleic acids. This allows eukaryotic Argonaute proteins to mediate RNA interference and long prokaryotic Argonaute proteins to interfere with invading nucleic acids. The function and mechanisms of the phylogenetically distinct short prokaryotic Argonaute proteins remain poorly understood. We demonstrate that short prokaryotic Argonaute and the associated TIR-APAZ (SPARTA) proteins form heterodimeric complexes. Upon guide RNA-mediated target DNA binding, four SPARTA heterodimers form oligomers in which TIR domain-mediated NAD(P)ase activity is unleashed. When expressed in Escherichia coli, SPARTA is activated in the presence of highly transcribed multicopy plasmid DNA, which causes cell death through NAD(P)+ depletion. This results in the removal of plasmid-invaded cells from bacterial cultures. Furthermore, we show that SPARTA can be repurposed for the programmable detection of DNA sequences. In conclusion, our work identifies SPARTA as a prokaryotic immune system that reduces cell viability upon RNA-guided detection of invading DNA.

Advanced Understanding of Prokaryotic Biofilm Formation through Use of a Cost-Effective and Versatile Multipanel Adhesion (mPAD) Mount.

Most microorganisms exist in biofilms, which comprise aggregates of cells surrounded by an extracellular matrix that provides protection from external stresses. Based on the conditions under which they form, biofilm structures vary in significant ways. For instance, biofilms that develop when microbes are incubated under static conditions differ from those formed when microbes encounter the shear forces of a flowing liquid. Moreover, biofilms develop dynamically over time. Here, we describe a cost-effective coverslip holder, printed with a three-dimensional (3D) printer, that facilitates surface adhesion assays under a broad range of standing and shaking culture conditions. This multipanel adhesion (mPAD) mount further allows cultures to be sampled at multiple time points, ensuring consistency and comparability between samples and enabling analyses of the dynamics of biofilm formation. As a proof of principle, using the mPAD mount for shaking, oxic cultures, we confirm previous flow chamber experiments showing that the Pseudomonas aeruginosa wild-type strain and a phenazine deletion mutant (Δphz) strain form biofilms with similar structure but reduced density in the mutant strain. Extending this analysis to anoxic conditions, we reveal that microcolony formation and biofilm formation can only be observed under shaking conditions and are decreased in the Δphz mutant compared to wild-type cultures, indicating that phenazines are crucial for the formation of biofilms if oxygen as an electron acceptor is unavailable. Furthermore, while the model archaeon Haloferax volcanii does not require archaella for surface attachment under static conditions, we demonstrate that an H. volcanii mutant that lacks archaella is impaired in early stages of biofilm formation under shaking conditions. IMPORTANCE Due to the versatility of the mPAD mount, we anticipate that it will aid the analysis of biofilm formation in a broad range of bacteria and archaea. Thereby, it contributes to answering critical biological questions about the regulatory and structural components of biofilm formation and understanding this process in a wide array of environmental, biotechnological, and medical contexts.

The structure of the Aquifex aeolicus MATE family multidrug resistance transporter and sequence comparisons suggest the existence of a new subfamily.

Multidrug and toxic compound extrusion (MATE) transporters are widespread in all domains of life. Bacterial MATE transporters confer multidrug resistance by utilizing an electrochemical gradient of H+ or Na+ to export xenobiotics across the membrane. Despite the availability of X-ray structures of several MATE transporters, a detailed understanding of the transport mechanism has remained elusive. Here we report the crystal structure of a MATE transporter from Aquifex aeolicus at 2.0-Å resolution. In light of its phylogenetic placement outside of the diversity of hitherto-described MATE transporters and the lack of conserved acidic residues, this protein may represent a subfamily of prokaryotic MATE transporters, which was proven by phylogenetic analysis. Furthermore, the crystal structure and substrate docking results indicate that the substrate binding site is located in the N bundle. The importance of residues surrounding this binding site was demonstrated by structure-based site-directed mutagenesis. We suggest that Aq_128 is functionally similar but structurally diverse from DinF subfamily transporters. Our results provide structural insights into the MATE transporter, which further advances our global understanding of this important transporter family.

Membrane nanotubes are ancient machinery for cell-to-cell communication and transport. Their interference with the immune system.

Nanotubular connections between mammalian cell types came into the focus only two decades ago, when "live cell super-resolution imaging" was introduced. Observations of these long-time overlooked structures led to understanding mechanisms of their growth/withdrawal and exploring some key genetic and signaling factors behind their formation. Unbelievable level of multiple supportive collaboration between tumor cells undergoing cytotoxic chemotherapy, cross-feeding" between independent bacterial strains or "cross-dressing" collaboration of immune cells promoting cellular immune response, all via nanotubes, have been explored recently. Key factors and "calling signals" determining the spatial directionality of their growth and their overall in vivo significance, however, still remained debated. Interestingly, prokaryotes, including even ancient archaebacteria, also seem to use such NT connections for intercellular communication. Herein, we will give a brief overview of current knowledge of membrane nanotubes and depict a simple model about their possible "historical role".

Prokaryotic rRNA-mRNA interactions are involved in all translation steps and shape bacterial transcripts.

The well-established Shine-Dalgarno model suggests that translation initiation in bacteria is regulated via base-pairing between ribosomal RNA (rRNA) and mRNA. We used novel computational analyses and modelling of 823 bacterial genomes coupled with experiments to demonstrate that rRNA-mRNA interactions are diverse and regulate all translation steps from pre-initiation to termination. Previous research has reported the significant influence of rRNA-mRNA interactions, mainly in the initiation phase of translation. The results reported in this paper suggest that, in addition to the rRNA-mRNA interactions near the start codon that trigger initiation in bacteria, rRNA-mRNA interactions affect all sub-stages of the translation process (pre-initiation, initiation, elongation, termination). As these interactions dictate translation efficiency, they serve as an evolutionary driving force for shaping transcripts in bacteria while considering trade-offs between the effects of different interactions across different transcript regions on translation efficacy and efficiency. We observed selection for strong interactions in regions where such interactions are likely to enhance initiation, regulate early elongation, and ensure translation termination fidelity. We discovered selection against strong interactions and for intermediate interactions in coding regions and presented evidence that these patterns maximize elongation efficiency while also enhancing initiation. These finding are relevant to all biomedical disciplines due to the centrality of the translation process and the effect of rRNA-mRNA interactions on transcript evolution.

Tandem Repeats in Bacillus: Unique Features and Taxonomic Distribution.

Little is known about DNA tandem repeats across prokaryotes. We have recently described an enigmatic group of tandem repeats in bacterial genomes with a constant repeat size but variable sequence. These findings strongly suggest that tandem repeat size in some bacteria is under strong selective constraints. Here, we extend these studies and describe tandem repeats in a large set of Bacillus. Some species have very few repeats, while other species have a large number. Most tandem repeats have repeats with a constant size (either 52 or 20-21 nt), but a variable sequence. We characterize in detail these intriguing tandem repeats. Individual species have several families of tandem repeats with the same repeat length and different sequence. This result is in strong contrast with eukaryotes, where tandem repeats of many sizes are found in any species. We discuss the possibility that they are transcribed as small RNA molecules. They may also be involved in the stabilization of the nucleoid through interaction with proteins. We also show that the distribution of tandem repeats in different species has a taxonomic significance. The data we present for all tandem repeats and their families in these bacterial species will be useful for further genomic studies.

Community composition and functional prediction of prokaryotes associated with sympatric sponge species of southwestern Atlantic coast.

Prokaryotes contribute to the health of marine sponges. However, there is lack of data on the assembly rules of sponge-associated prokaryotic communities, especially for those inhabiting biodiversity hotspots, such as ecoregions between tropical and warm temperate southwestern Atlantic waters. The sympatric species Aplysina caissara, Axinella corrugata, and Dragmacidon reticulatum were collected along with environmental samples from the north coast of São Paulo (Brazil). Overall, 64 prokaryotic phyla were detected; 51 were associated with sponge species, and the dominant were Proteobacteria, Bacteria (unclassified), Cyanobacteria, Crenarchaeota, and Chloroflexi. Around 64% and 89% of the unclassified operational taxonomical units (OTUs) associated with Brazilian sponge species showed a sequence similarity below 97%, with sequences in the Silva and NCBI Type Strain databases, respectively, indicating the presence of a large number of unidentified taxa. The prokaryotic communities were species-specific, ranging 56%-80% of the OTUs and distinct from the environmental samples. Fifty-four lineages were responsible for the differences detected among the categories. Functional prediction demonstrated that Ap. caissara was enriched for energy metabolism and biosynthesis of secondary metabolites, whereas D. reticulatum was enhanced for metabolism of terpenoids and polyketides, as well as xenobiotics' biodegradation and metabolism. This survey revealed a high level of novelty associated with Brazilian sponge species and that distinct members responsible from the differences among Brazilian sponge species could be correlated to the predicted functions.

Estimating maximal microbial growth rates from cultures, metagenomes, and single cells via codon usage patterns.

Maximal growth rate is a basic parameter of microbial lifestyle that varies over several orders of magnitude, with doubling times ranging from a matter of minutes to multiple days. Growth rates are typically measured using laboratory culture experiments. Yet, we lack sufficient understanding of the physiology of most microbes to design appropriate culture conditions for them, severely limiting our ability to assess the global diversity of microbial growth rates. Genomic estimators of maximal growth rate provide a practical solution to survey the distribution of microbial growth potential, regardless of cultivation status. We developed an improved maximal growth rate estimator and predicted maximal growth rates from over 200,000 genomes, metagenome-assembled genomes, and single-cell amplified genomes to survey growth potential across the range of prokaryotic diversity; extensions allow estimates from 16S rRNA sequences alone as well as weighted community estimates from metagenomes. We compared the growth rates of cultivated and uncultivated organisms to illustrate how culture collections are strongly biased toward organisms capable of rapid growth. Finally, we found that organisms naturally group into two growth classes and observed a bias in growth predictions for extremely slow-growing organisms. These observations ultimately led us to suggest evolutionary definitions of oligotrophy and copiotrophy based on the selective regime an organism occupies. We found that these growth classes are associated with distinct selective regimes and genomic functional potentials.

Learning in single cell organisms.

The survival of all species requires appropriate behavioral responses to environmental challenges. Learning is one of the key processes to acquire information about the environment and adapt to changing and uncertain conditions. Learning has long been acknowledged in animals from invertebrates to vertebrates but remains a subject of debate in non-animal systems such a plants and single cell organisms. In this review I will attempt to answer the following question: are single cell organisms capable of learning? I will first briefly discuss the concept of learning and argue that the ability to acquire and store information through learning is pervasive and may be found in single cell organisms. Second, by focusing on habituation, the simplest form of learning, I will review a series of experiments showing that single cell organisms such as slime molds and ciliates display habituation and follow most of the criteria adopted by neuroscientists to define habituation. Then I will discuss disputed evidence suggesting that single cell organisms might also undergo more sophisticated forms of learning such as associative learning. Finally, I will stress out that the challenge for the future is less about whether or not to single cell organisms fulfill the definition of learning established from extensive studies in animal systems and more about acknowledging and understanding the range of behavioral plasticity exhibited by such fascinating organisms.

Reframing cognition: getting down to biological basics.

The premise of this two-part theme issue is simple: the cognitive sciences should join the rest of the life sciences in how they approach the quarry within their research domain. Specifically, understanding how organisms on the lower branches of the phylogenetic tree become familiar with, value and exploit elements of an ecological niche while avoiding harm can be expected to aid understanding of how organisms that evolved later (including Homo sapiens) do the same or similar things. We call this approach basal cognition. In this introductory essay, we explain what the approach involves. Because no definition of cognition exists that reflects its biological basis, we advance a working definition that can be operationalized; introduce a behaviour-generating toolkit of capacities that comprise the function (e.g. sensing/perception, memory, valence, learning, decision making, communication), each element of which can be studied relatively independently; and identify a (necessarily incomplete) suite of common biophysical mechanisms found throughout the domains of life involved in implementing the toolkit. The articles in this collection illuminate different aspects of basal cognition across different forms of biological organization, from prokaryotes and single-celled eukaryotes-the focus of Part 1-to plants and finally to animals, without and with nervous systems, the focus of Part 2. By showcasing work in diverse, currently disconnected fields, we hope to sketch the outline of a new multidisciplinary approach for comprehending cognition, arguably the most fascinating and hard-to-fathom evolved function on this planet. Doing so has the potential to shed light on problems in a wide variety of research domains, including microbiology, immunology, zoology, biophysics, botany, developmental biology, neurobiology/science, regenerative medicine, computational biology, artificial life and synthetic bioengineering. This article is part of the theme issue 'Basal cognition: conceptual tools and the view from the single cell'.

Spontaneous electrical low-frequency oscillations: a possible role in Hydra and all living systems.

As one of the first model systems in biology, the basal metazoan Hydra has been revealing fundamental features of living systems since it was first discovered by Antonie van Leeuwenhoek in the early eighteenth century. While it has become well-established within cell and developmental biology, this tiny freshwater polyp is only now being re-introduced to modern neuroscience where it has already produced a curious finding: the presence of low-frequency spontaneous neural oscillations at the same frequency as those found in the default mode network in the human brain. Surprisingly, increasing evidence suggests such spontaneous electrical low-frequency oscillations (SELFOs) are found across the wide diversity of life on Earth, from bacteria to humans. This paper reviews the evidence for SELFOs in diverse phyla, beginning with the importance of their discovery in Hydra, and hypothesizes a potential role as electrical organism organizers, which supports a growing literature on the role of bioelectricity as a 'template' for developmental memory in organism regeneration. This article is part of the theme issue 'Basal cognition: conceptual tools and the view from the single cell'.

Valuing what happens: a biogenic approach to valence and (potentially) affect.

Valence is half of the pair of properties that constitute core affect, the foundation of emotion. But what is valence, and where is it found in the natural world? Currently, this question cannot be answered. The idea that emotion is the body's way of driving the organism to secure its survival, thriving and reproduction runs like a leitmotif from the pathfinding work of Antonio Damasio through four book-length neuroscientific accounts of emotion recently published by the field's leading practitioners. Yet while Damasio concluded 20 years ago that the homeostasis-affect linkage is rooted in unicellular life, no agreement exists about whether even non-human animals with brains experience emotions. Simple neural animals-those less brainy than bees, fruit flies and other charismatic invertebrates-are not even on the radar of contemporary affective research, to say nothing of aneural organisms. This near-sightedness has effectively denied the most productive method available for getting a grip on highly complex biological processes to a scientific domain whose importance for understanding biological decision-making cannot be underestimated. Valence arguably is the fulcrum around which the dance of life revolves. Without the ability to discriminate advantage from harm, life very quickly comes to an end. In this paper, we review the concept of valence, where it came from, the work it does in current leading theories of emotion, and some of the odd features revealed via experiment. We present a biologically grounded framework for investigating valence in any organism and sketch a preliminary pathway to a computational model. This article is part of the theme issue 'Basal cognition: conceptual tools and the view from the single cell'.

Grounding cognition: heterarchical control mechanisms in biology.

We advance an account that grounds cognition, specifically decision-making, in an activity all organisms as autonomous systems must perform to keep themselves viable-controlling their production mechanisms. Production mechanisms, as we characterize them, perform activities such as procuring resources from their environment, putting these resources to use to construct and repair the organism's body and moving through the environment. Given the variable nature of the environment and the continual degradation of the organism, these production mechanisms must be regulated by control mechanisms that select when a production is required and how it should be carried out. To operate on production mechanisms, control mechanisms need to procure information through measurement processes and evaluate possible actions. They are making decisions. In all organisms, these decisions are made by multiple different control mechanisms that are organized not hierarchically but heterarchically. In many cases, they employ internal models of features of the environment with which the organism must deal. Cognition, in the form of decision-making, is thus fundamental to living systems which must control their production mechanisms. This article is part of the theme issue 'Basal cognition: conceptual tools and the view from the single cell'.

All living cells are cognitive.

All living cells sense and respond to changes in external or internal conditions. Without that cognitive capacity, they could not obtain nutrition essential for growth, survive inevitable ecological changes, or correct accidents in the complex processes of reproduction. Wherever examined, even the smallest living cells (prokaryotes) display sophisticated regulatory networks establishing appropriate adaptations to stress conditions that maximize the probability of survival. Supposedly "simple" prokaryotic organisms also display remarkable capabilities for intercellular signalling and multicellular coordination. These observations indicate that all living cells are cognitive.

Hyperbolic rules of the cooperative organization of eukaryotic and prokaryotic genomes.

The author's method of oligomer sums for analysis of oligomer compositions of eukaryotic and prokaryotic genomes is described. The use of this method revealed the existence of general rules for the cooperative oligomeric organization of a wide list of genomes. These rules are called hyperbolic because they are associated with hyperbolic sequences including the harmonic progression 1, 1/2, 1/3, .., 1/n. These rules are demonstrated by examples of quantitative analysis of many genomes from the human genome to the genomes of archaea and bacteria. The hyperbolic (harmonic) rules, speaking about the existence of algebraic invariants in full genomic sequences, are considered as candidates for the role of universal rules for the cooperative organization of genomes. The results concerns additionally the problem of the origin of life. The described phenomenological results were obtained as consequences of the previously published author's quantum-information model of long DNA sequences. The oligomer sums method was also applied to the analysis of long genes and viruses including the COVID-19 virus; this revealed, in characteristics of many of them, the phenomenon of such rhythmically repeating deviations from model hyperbolic sequences, which are associated with DNA triplets. In addition, an application of the oligomer sums method is shown to the analysis of amino acid sequences in long proteins like the protein Titin. The topics of the algebraic harmony in living bodies and of the quantum-information approach in biology are discussed.

Prokaryote autoimmunity in the context of self-targeting by CRISPR-Cas systems.

Prokaryote adaptive immunity (CRISPR-Cas systems) can be a threat to its carriers. We analyze the risks of autoimmune reactions related to adaptive immunity in prokaryotes by computational methods. We found important differences between bacteria and archaea with respect to autoimmunity potential. According to the results of our analysis, CRISPR-Cas systems in bacteria are more prone to self-targeting even though they possess fewer spacers per organism on average than archaea. The results of our study provide opportunities to use self-targeting in prokaryotes for biological and medical applications.

Femtoplankton: What's New?

Since the discovery of high abundances of virus-like particles in aquatic environment, emergence of new analytical methods in microscopy and molecular biology has allowed significant advances in the characterization of the femtoplankton, i.e., floating entities filterable on a 0.2 µm pore size filter. The successive evidences in the last decade (2010-2020) of high abundances of biomimetic mineral-organic particles, extracellular vesicles, CPR/DPANN (Candidate phyla radiation/Diapherotrites, Parvarchaeota, Aenigmarchaeota, Nanoarchaeota and Nanohaloarchaeota), and very recently of aster-like nanoparticles (ALNs), show that aquatic ecosystems form a huge reservoir of unidentified and overlooked femtoplankton entities. The purpose of this review is to highlight this unsuspected diversity. Herein, we focus on the origin, composition and the ecological potentials of organic femtoplankton entities. Particular emphasis is given to the most recently discovered ALNs. All the entities described are displayed in an evolutionary context along a continuum of complexity, from minerals to cell-like living entities.

TEMPURA: Database of Growth TEMPeratures of Usual and RAre Prokaryotes.

Growth temperature is one of the most representative biological parameters for characterizing living organisms. Prokaryotes have been isolated from various temperature environments and show wide diversity in their growth temperatures. We herein constructed a database of growth TEMPeratures of Usual and RAre prokaryotes (TEMPURA, http://togodb.org/db/tempura), which contains the minimum, optimum, and maximum growth temperatures of 8,639 prokaryotic strains. Growth temperature information is linked with taxonomy IDs, phylogenies, and genomic information. TEMPURA provides useful information to researchers working on biotechnological applications of extremophiles and their biomolecules as well as those performing fundamental studies on the physiological diversity of prokaryotes.

Structural and functional shifts of soil prokaryotic community due to Eucalyptus plantation and rotation phase.

Agriculture, forestry and other land uses are currently the second highest source of anthropogenic greenhouse gases (GHGs) emissions. In soil, these gases derive from microbial activity, during carbon (C) and nitrogen (N) cycling. To investigate how Eucalyptus land use and growth period impact the microbial community, GHG fluxes and inorganic N levels, and if there is a link among these variables, we monitored three adjacent areas for 9 months: a recently planted Eucalyptus area, fully developed Eucalyptus forest (final of rotation) and native forest. We assessed the microbial community using 16S rRNA gene sequencing and qPCR of key genes involved in C and N cycles. No considerable differences in GHG flux were evident among the areas, but logging considerably increased inorganic N levels. Eucalyptus areas displayed richer and more diverse communities, with selection for specific groups. Land use influenced communities more extensively than the time of sampling or growth phase, although all were significant modulators. Several microbial groups and genes shifted temporally, and inorganic N levels shaped several of these changes. No correlations among microbial groups or genes and GHG were found, suggesting no link among these variables in this short-rotation Eucalyptus study.