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A key feature that differentiates prokaryotic cells from eukaryotes is the absence of an intracellular membrane surrounding the chromosomal DNA. Here, we isolate a member of the ubiquitous, yet-to-be-cultivated phylum ‘ Candidatus Atribacteria’ (also known as OP9) that has an intracytoplasmic membrane apparently surrounding the nucleoid. The isolate, RT761, is a subsurface-derived anaerobic bacterium that appears to have three lipid membrane-like layers, as shown by cryo-electron tomography. Our observations are consistent with a classical gram-negative structure with an additional intracytoplasmic membrane. However, further studies are needed to provide conclusive evidence for this unique intracellular structure. The RT761 genome encodes proteins with features that might be related to the complex cellular structure, including: N-terminal extensions in proteins involved in important processes (such as cell-division protein FtsZ); one of the highest percentages of transmembrane proteins among gram-negative bacteria; and predicted Sec-secreted proteins with unique signal peptides. Physiologically, RT761 primarily produces hydrogen for electron disposal during sugar degradation, and co-cultivation with a hydrogen-scavenging methanogen improves growth. We propose RT761 as a new species, Atribacter laminatus gen. nov. sp. nov. and a new phylum, Atribacterota phy. nov. A key feature that differentiates prokaryotic cells from eukaryotes is the absence of an intracellular membrane surrounding the chromosomal DNA. Here, the authors isolate a member of the ubiquitous, yet-to-be-cultivated bacterial phylum ‘ Candidatus Atribacteria’ that has an intracytoplasmic membrane apparently surrounding the nucleoid.

Cholesterol is an integral component of eukaryotic cell membranes and a key molecule in controlling membrane fluidity, organization, and other physicochemical parameters. It also plays a regulatory function in antibiotic drug resistance and the immune response of cells against viruses, by stabilizing the membrane against structural damage. While it iswell understood that, structurally, cholesterol exhibits a densification effect on fluid lipid membranes, its effects on membrane bending rigidity are assumed to be nonuniversal; i.e., cholesterol stiffens saturated lipid membranes, but has no stiffening effect on membranes populated by unsaturated lipids, such as 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC). This observation presents a clear challenge to structure–property relationships and to our understanding of cholesterol-mediated biological functions. Here, using a comprehensive approach—combining neutron spin-echo (NSE) spectroscopy, solid-state deuterium NMR (²H NMR) spectroscopy, and molecular dynamics (MD) simulations—we report that cholesterol locally increases the bending rigidity of DOPC membranes, similar to saturated membranes, by increasing the bilayer’s packing density. All three techniques, inherently sensitive to mesoscale bending fluctuations, show up to a threefold increase in effective bending rigidity with increasing cholesterol content approaching a mole fraction of 50%. Our observations are in good agreement with the known effects of cholesterol on the area-compressibility modulus and membrane structure, reaffirming membrane structure–property relationships. The current findings point to a scale-dependent manifestation of membrane properties, highlighting the need to reassess cholesterol’s role in controlling membrane bending rigidity over mesoscopic length and time scales of important biological functions, such as viral budding and lipid–protein interactions.

Extracellular vesicle production is a ubiquitous process in Gram-negative bacteria, but little is known about such process in Gram-positive bacteria. We report the isolation of extracellular vesicles from the supernatants of Bacillus anthracis, a Gram-positive bacillus that is a powerful agent for biological warfare. B. anthracis vesicles formed at the outer layer of the bacterial cell had double-membrane spheres and ranged from 50 to 150 nm in diameter. Immunoelectron microscopy with mAbs to protective antigen, lethal factor, edema toxin, and anthrolysin revealed toxin components and anthrolysin in vesicles, with some vesicles containing more than one toxin component. Toxin-containing vesicles were also visualized inside B. anthracis-infected macrophages. ELISA and immunoblot analysis of vesicle preparations confirmed the presence of B. anthracis toxin components. A mAb to protective antigen protected macrophages against vesicles from an anthrolysin-deficient strain, but not against vesicles from Sterne 34F2 and Sterne δT strains, consistent with the notion that vesicles delivered both toxin and anthrolysin to host cells. Vesicles were immunogenic in BALB/c mice, which produced a robust IgM response to toxin components. Furthermore, vesicle-immunized mice lived significantly longer than controls after B. anthracis challenge. Our results indicate that toxin secretion in B. anthracis is, at least, partially vesicle-associated, thus allowing concentrated delivery of toxin components to target host cells, a mechanism that may increase toxin potency. Our observations may have important implications for the design of vaccines, for passive antibody strategies, and provide a previously unexplored system for studying secretory pathways in Gram-positive bacteria.

Cells maintain membrane fluidity by regulating lipid saturation, but the molecular mechanisms of this homeoviscous adaptation remain poorly understood. We have reconstituted the core machinery for regulating lipid saturation in baker’s yeast to study its molecular mechanism. By combining molecular dynamics simulations with experiments, we uncover a remarkable sensitivity of the transcriptional regulator Mga2 to the abundance, position, and configuration of double bonds in lipid acyl chains, and provide insights into the molecular rules of membrane adaptation. Our data challenge the prevailing hypothesis that membrane fluidity serves as the measured variable for regulating lipid saturation. Rather, we show that Mga2 senses the molecular lipid-packing density in a defined region of the membrane. Our findings suggest that membrane property sensors have evolved remarkable sensitivities to highly specific aspects of membrane structure and dynamics, thus paving the way toward the development of genetically encoded reporters for such properties in the future. Cells maintain membrane fluidity by regulating lipid saturation, but the molecular mechanisms of this homeoviscous adaptation remain poorly understood. Here authors reconstituted the core machinery for regulating lipid saturation in baker’s yeast to directly characterize its response to defined membrane environments and uncover its mode-of-action.

Microplastics (MPs) cannot be completely removed from water/wastewater in conventional wastewater treatment plants (WWTPs) and drinking water treatment plants (DWTPs). According to the literature analysis, membrane technologies, one of the advanced treatment technologies, are the most effective and promising technologies for MP removal from water and wastewater. In this paper, firstly, the properties of MPs commonly present in WWTPs/DWTPs and the MP removal efficiency of WWTPs/DWTPs are briefly reviewed. In addition, research studies on MP removal from water/wastewater by microfiltration (MF), ultrafiltration (UF), nanofiltration (NF), reverse osmosis (RO), and membrane bioreactors (MBRs) are reviewed. In the next section, membrane filtration is compared with other methods used for MP removal from water/wastewater, and the advantages/disadvantages of the removal methods are discussed. Moreover, the problem of membrane fouling with MPs during filtration and the potential for MP release from polymeric membrane structure to water/wastewater are discussed. Finally, based on the studies in the literature, the current status and research deficiencies of MP removal by membrane technologies are identified, and recommendations are made for further studies.

Examining the fundamental structure and processes of living cells at the nanoscale poses a unique analytical challenge, as cells are dynamic, chemically diverse, and fragile. A case in point is the cell membrane, which is too small to be seen directly with optical microscopy and provides little observational contrast for other methods. As a consequence, nanoscale characterization of the membrane has been performed ex vivo or in the presence of exogenous labels used to enhance contrast and impart specificity. Here, we introduce an isotopic labeling strategy in the gram-positive bacterium Bacillus subtilis to investigate the nanoscale structure and organization of its plasma membrane in vivo. Through genetic and chemical manipulation of the organism, we labeled the cell and its membrane independently with specific amounts of hydrogen (H) and deuterium (D). These isotopes have different neutron scattering properties without altering the chemical composition of the cells. From neutron scattering spectra, we confirmed that the B. subtilis cell membrane is lamellar and determined that its average hydrophobic thickness is 24.3 ± 0.9 Ångstroms (Å). Furthermore, by creating neutron contrast within the plane of the membrane using a mixture of H- and D-fatty acids, we detected lateral features smaller than 40 nm that are consistent with the notion of lipid rafts. These experiments-performed under biologically relevant conditions-answer long-standing questions in membrane biology and illustrate a fundamentally new approach for systematic in vivo investigations of cell membrane structure.

Phospholipids are asymmetrically distributed in the plasma membrane (PM), with phosphatidylcholine and sphingomyelin abundant in the outer leaflet. However, the mechanisms by which their distribution is regulated remain unclear. Here, we show that transmembrane protein 63B (TMEM63B) functions as a membrane structure-responsive lipid scramblase localized at the PM and lysosomes, activating bidirectional lipid translocation upon changes in membrane curvature and thickness. TMEM63B contains two intracellular loops with palmitoylated cysteine residue clusters essential for its scrambling function. TMEM63B deficiency alters phosphatidylcholine and sphingomyelin distributions in the PM. Persons with heterozygous mutations in TMEM63B are known to develop neurodevelopmental disorders. We show that V44M, the most frequent substitution, confers constitutive scramblase activity on TMEM63B, disrupting PM phospholipid asymmetry. We determined the cryo-electron microscopy structures of TMEM63B in its open and closed conformations, uncovering a lipid translocation pathway formed in response to changes in the membrane environment. Together, our results identify TMEM63B as a membrane structure-responsive scramblase that controls PM lipid distribution and we reveal the molecular basis for lipid scrambling and its biological importance. By combining genome-wide clustered regularly interspaced short palindromic repeats with Cas9 screening and cryo-electron microscopy structure analysis, the authors identified transmembrane protein 63B as a lipid scramblase that detects structural changes in the lipid bilayer and scrambles lipids to regulate membrane lipid distributions.

Implant-associated infections due to the formation of bacterial biofilms pose a serious threat in medical healthcare, which needs effective therapeutic methods. Here, we propose a multifunctional nanoreactor by spatiotemporal ultrasound-driven tandem catalysis to amplify the efficacy of sonodynamic and chemodynamic therapy. By combining piezoelectric barium titanate with polydopamine and copper, the ultrasound-activated piezo-hot carriers transfer easily to copper by polydopamine. It boosts reactive oxygen species production by piezoelectrics, and facilitates the interconversion between Cu2 + and Cu + to promote hydroxyl radical generation via Cu +  -catalyzed chemodynamic reactions. Finally, the elevated reactive oxygen species cause bacterial membrane structure loosening and DNA damage. Transcriptomics and metabolomics analysis reveal that intracellular copper overload restricts the tricarboxylic acid cycle, promoting bacterial cuproptosis-like death. Therefore, the polyetherketoneketone scaffold engineered with the designed nanoreactor shows excellent antibacterial performance with ultrasound stimulation and promotes angiogenesis and osteogenesis on-demand in vivo. Implantation-associated infections often lead to infections. Here, the authors propose a piezo-based nanoreactor to achieve US-excited tandem catalysis, endowing the polyetherketoneketone bone scaffold with on-demand antibacterial and osteogenic capacities.

Autophagosome biogenesis is an essential feature of autophagy. Lipidation of Atg8 plays a critical role in this process. Previous in vitro studies identified membrane tethering and hemi-fusion/fusion activities of Atg8, yet definitive roles in autophagosome biogenesis remained controversial. Here, we studied the effect of Atg8 lipidation on membrane structure. Lipidation of Saccharomyces cerevisiae Atg8 on nonspherical giant vesicles induced dramatic vesicle deformation into a sphere with an out-bud. Solution NMR spectroscopy of Atg8 lipidated on nanodiscs identified two aromatic membrane-facing residues that mediate membrane-area expansion and fragmentation of giant vesicles in vitro. These residues also contribute to the in vivo maintenance of fragmented vacuolar morphology under stress in fission yeast, a moonlighting function of Atg8. Furthermore, these aromatic residues are crucial for the formation of a sufficient number of autophagosomes and regulate autophagosome size. Together, these data demonstrate that Atg8 can cause membrane perturbations that underlie efficient autophagosome biogenesis. Lipidated Atg8 affects membrane morphology via two aromatic membrane-facing residues that are important for autophagy in budding yeast and mammalian cells.

RNase E, which is the central component of the multienzyme RNA degradosome, serves as a scaffold for interaction with other enzymes involved in mRNA degradation including the DEAD-box RNA helicase RhlB. Epifluorescence microscopy under live cell conditions shows that RNase E and RhlB are membrane associated, but neither protein forms cytoskeletal-like structures as reported earlier by Taghbalout and Rothfield. We show that association of RhlB with the membrane depends on a direct protein interaction with RNase E, which is anchored to the inner cytoplasmic membrane through an MTS (Membrane Targeting Sequence). Molecular dynamics simulations show that the MTS interacts with the phospholipid bilayer by forming a stabilized amphipathic α-helix with the helical axis oriented parallel to the plane of the bilayer and hydrophobic side chains buried deep in the acyl core of the membrane. Based on the molecular dynamics simulations, we propose that the MTS freely diffuses in the membrane by a novel mechanism in which a large number of weak contacts are rapidly broken and reformed. TIRFm (Total Internal Reflection microscopy) shows that RNase E in live cells rapidly diffuses over the entire inner membrane forming short-lived foci. Diffusion could be part of a scanning mechanism facilitating substrate recognition and cooperativity. Remarkably, RNase E foci disappear and the rate of RNase E diffusion increases with rifampicin treatment. Control experiments show that the effect of rifampicin is specific to RNase E and that the effect is not a secondary consequence of the shut off of E. coli transcription. We therefore interpret the effect of rifampicin as being due to the depletion of RNA substrates for degradation. We propose a model in which formation of foci and constraints on diffusion arise from the transient clustering of RNase E into cooperative degradation bodies.