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Past functional toxicogenomic studies have indicated that genes relevant to membrane lipid synthesis are important for tolerance to the lanthanides. Moreover, previously reported imaging of patient's brains following administration of gadolinium-based contrast agents shows gadolinium lining the vessels of the brain. Taken together, these findings suggest the disruption of cytoplasmic membrane integrity as a mechanism by which lanthanides induce cytotoxicity. In the presented work we used scanning transmission electron microscopy and spatially resolved elemental spectroscopy to image the morphology and composition of gadolinium, europium, and samarium precipitates that formed on the outside of yeast cell membranes. In no sample did we find that the lanthanide contaminant had crossed the cell membrane, even in experiments using yeast mutants with disrupted genes for sphingolipid synthesis-the primary lipids found in yeast cytoplasmic membranes. Rather, we have evidence that lanthanides are co-located with phosphorus outside the yeast cells. These results lead us to hypothesize that the lanthanides scavenge or otherwise form complexes with phosphorus from the sphingophospholipid head groups in the cellular membrane, thereby compromising the structure or function of the membrane, and gaining the ability to disrupt membrane function without entering the cell.

Signaling through the Ras-MAPK pathway can exhibit switch-like activation, which has been attributed to the underlying positive feedback and bimodality in the activation of RasGDP to RasGTP by SOS. SOS contains both catalytic and allosteric Ras binding sites, and a common assumption is that allosteric activation selectively by RasGTP provides the mechanism of positive feedback. However, recent single-molecule studies have revealed that SOS catalytic rates are independent of the nucleotide state of Ras in the allosteric binding site, raising doubt about this as a positive feedback mechanism. Here, we perform detailed kinetic analyses of receptor-mediated recruitment of full-length SOS to the membrane while simultaneously monitoring its catalytic activation of Ras. These results, along with kinetic modeling, expose the autoinhibition release step in SOS, rather than either recruitment or allosteric activation, as the underlying mechanism giving rise to positive feedback in Ras activation.

Clathrin-mediated endocytosis (CME) is a critical trafficking process that begins when an elaborate endocytic protein network is established at the plasma membrane. Interaction of early endocytic proteins with anionic phospholipids and/or cargo has been suggested to trigger CME initiation. However, the exact mechanism by which CME sites are initiated has not been fully elucidated. In the budding yeast Saccharomyces cerevisiae, higher levels of anionic phospholipids and cargo molecules exist in the newly formed daughter cell compared to the levels in the mother cell during polarized growth. Taking advantage of this asymmetry, we quantitatively compared CME proteins in S. cerevisiae mother versus daughter cells, observing differences in the dynamics and composition of key endocytic proteins. Our results show that CME site initiation occurs preferentially on regions of the plasma membrane with a relatively higher density of endocytic cargo and/or acidic phospholipids. Furthermore, our combined live cell-imaging and yeast genetics analysis provided evidence for a molecular mechanism in which CME sites are initiated when Yap1801 and Yap1802 (yeast CALM/AP180) and Syp1 (yeast FCHo1/2) coordinate with anionic phospholipids and cargo molecules to trigger Ede1 (yeast Eps15)-centric CME initiation complex assembly at the plasma membrane.

The cAMP cascade is increasingly recognized to transduce physiological effects locally through spatially limited cAMP gradients. However, little is known about how adenylyl cyclase enzymes that initiate cAMP gradients are localized. Here we address this question in physiologically relevant striatal neurons and investigate how AC localization impacts downstream signaling function. We show that the major striatal AC isoforms are differentially sorted between ciliary and extraciliary domains of the plasma membrane, and that one isoform, AC9, is uniquely concentrated in endosomes. We identify key sorting determinants in the N-terminal cytoplasmic domain responsible for isoform-specific localization. We further show that AC9-containing endosomes accumulate activated dopamine receptors and form an elaborately intertwined network with juxtanuclear PKA stores bound to Golgi membranes. Finally, we provide evidence that endosomal localization enables AC9 to selectively elevate PKA activity in the nucleus relative to the cytoplasm. Together, these results reveal a precise spatial landscape of the cAMP cascade in neurons and a key role of AC localization in directing downstream PKA signaling to the nucleus.

To facilitate our understanding of proteome dynamics during signaling events, robust workflows affording fast time resolution without confounding factors are essential. We present Surface-exposed protein Labeling using PeroxidaSe, H2O2, and Tyramide-derivative (SLAPSHOT) to label extracellularly exposed proteins in a rapid, specific, and sensitive manner. Simple and flexible SLAPSHOT utilizes recombinant soluble APEX2 protein applied to cells, thus circumventing the engineering of tools and cells, biological perturbations, and labeling biases. We applied SLAPSHOT and quantitative proteomics to examine the TMEM16F-dependent plasma membrane remodeling in WT and TMEM16F KO cells. Time-course data ranging from 1 to 30 min of calcium stimulation revealed co-regulation of known protein families, including the integrin and ICAM families, and identified proteins known to reside in intracellular organelles as occupants of the freshly deposited extracellularly exposed membrane. Our data provide the first accounts of the immediate consequences of calcium signaling on the extracellularly exposed proteome.

The M2 proton channel aids in the exit of mature influenza viral particles from the host plasma membrane through its ability to stabilize regions of high negative Gaussian curvature (NGC) that occur at the neck of budding virions. The channels are homo-tetramers that contain a cytoplasm-facing amphipathic helix (AH) that is necessary and sufficient for NGC generation; however, constructs containing the transmembrane spanning helix, which facilitates tetramerization, exhibit enhanced curvature generation. Here, we used all-atom molecular dynamics (MD) simulations to explore the conformational dynamics of M2 channels in lipid bilayers revealing that the AH is dynamic, quickly breaking the fourfold symmetry observed in most structures. Next, we carried out MD simulations with the protein restrained in four- and twofold symmetric conformations to determine the impact on the membrane shape. While each pattern was distinct, all configurations induced pronounced curvature in the outer leaflet, while conversely, the inner leaflets showed minimal curvature and significant lipid tilt around the AHs. The MD-generated profiles at the protein-membrane interface were then extracted and used as boundary conditions in a continuum elastic membrane model to calculate the membrane-bending energy of each conformation embedded in different membrane surfaces characteristic of a budding virus. The calculations show that all three M2 conformations are stabilized in inward-budding, concave spherical caps and destabilized in outward-budding, convex spherical caps, the latter reminiscent of a budding virus. One of the C2-broken symmetry conformations is stabilized by 4 kT in NGC surfaces with the minimum energy conformation occurring at a curvature corresponding to 33 nm radii. In total, our work provides atomistic insight into the curvature sensing capabilities of M2 channels and how enrichment in the nascent viral particle depends on protein shape and membrane geometry.

Saponins are plant secondary metabolites comprising glycosylated triterpenoids, steroids or steroidal alkaloids with a broad spectrum of toxicity to microbial pathogens and pest organisms that contribute to basal plant defense to biotic attack. Secretion of glycosyl hydrolases that enzymatically convert saponins into less toxic products was thus far the only mechanism reported to enable fungal pathogens to colonize their saponin-containing host plant(s). We studied the mechanisms that the fungus Botrytis cinerea utilizes to be tolerant to well-characterized, structurally related saponins from tomato and Digitalis purpurea. By gene expression studies, comparative genomics, enzyme assays and testing a large panel of fungal (knockout and complemented) mutants, we unraveled four distinct cellular mechanisms that participate in the mitigation of the toxic activity of these saponins and in virulence on saponin-producing host plants. The enzymatic deglycosylation that we identified is novel and unique to this fungus-saponin combination. The other three tolerance mechanisms operate in the fungal membrane and are mediated by protein families that are widely distributed in the fungal kingdom. We present a spatial and temporal model on how these mechanisms jointly confer tolerance to saponins and discuss the repercussions of these findings for other plant pathogenic fungi, as well as human pathogens.

UNLABELLED: Cryptococcus neoformans causes cryptococcosis, one of the most prevalent fungal diseases, generally characterized by meningitis. There is a limited and not very effective number of drugs available to combat this disease. In this manuscript, we show the host defense peptide mimetic brilacidin (BRI) as a promising antifungal drug against C. neoformans. BRI can affect the organization of the cell membrane, increasing the fungal cell permeability. We also investigated the effects of BRI against the model system Saccharomyces cerevisiae by analyzing libraries of mutants grown in the presence of BRI. In S. cerevisiae, BRI also affects the cell membrane organization, but in addition the cell wall integrity pathway and calcium metabolism. In vivo experiments show BRI significantly reduces C. neoformans survival inside macrophages and partially clears C. neoformans lung infection in an immunocompetent murine model of invasive pulmonary cryptococcosis. We also observed that BRI interacts with caspofungin (CAS) and amphotericin (AmB), potentiating their mechanism of action against C. neoformans. BRI + CAS affects endocytic movement, calcineurin, and mitogen-activated protein kinases. Our results indicate that BRI is a novel antifungal drug against cryptococcosis. IMPORTANCE: Invasive fungal infections have a high mortality rate causing more deaths annually than tuberculosis or malaria. Cryptococcosis, one of the most prevalent fungal diseases, is generally characterized by meningitis and is mainly caused by two closely related species of basidiomycetous yeasts, Cryptococcus neoformans and Cryptococcus gattii. There are few therapeutic options for treating cryptococcosis, and searching for new antifungal agents against this disease is very important. Here, we present brilacidin (BRI) as a potential antifungal agent against C. neoformans. BRI is a small molecule host defense peptide mimetic that has previously exhibited broad-spectrum immunomodulatory/anti-inflammatory activity against bacteria and viruses. BRI alone was shown to inhibit the growth of C. neoformans, acting as a fungicidal drug, but surprisingly also potentiated the activity of caspofungin (CAS) against this species. We investigated the mechanism of action of BRI and BRI + CAS against C. neoformans. We propose BRI as a new antifungal agent against cryptococcosis.

The cell cortex is a dynamic assembly formed by the plasma membrane and underlying cytoskeleton. As the main determinant of cell shape, the cortex ensures its integrity during passive and active deformations by adapting cytoskeleton topologies through yet poorly understood mechanisms. The spectrin meshwork ensures such adaptation in erythrocytes and neurons by adopting different organizations. Erythrocytes rely on triangular-like lattices of spectrin tetramers, whereas in neurons they are organized in parallel, periodic arrays. Since spectrin is ubiquitously expressed, we exploited Expansion Microscopy to discover that, in fibroblasts, distinct meshwork densities co-exist. Through biophysical measurements and computational modeling, we show that the non-polarized spectrin meshwork, with the intervention of actomyosin, can dynamically transition into polarized clusters fenced by actin stress fibers that resemble periodic arrays as found in neurons. Clusters experience lower mechanical stress and turnover, despite displaying an extension close to the tetramer contour length. Our study sheds light on the adaptive properties of spectrin, which participates in the protection of the cell cortex by varying its densities in response to key mechanical features.

Professional phagocytes like neutrophils and macrophages tightly control what they consume, how much they consume, and when they move after cargo uptake. We show that plasma membrane abundance is a key arbiter of these cellular behaviors. Neutrophils and macrophages lacking the G protein subunit Gβ4 exhibited profound plasma membrane expansion, accompanied by marked reduction in plasma membrane tension. These biophysical changes promoted the phagocytosis of bacteria, fungus, apoptotic corpses, and cancer cells. We also found that Gβ4-deficient neutrophils are defective in the normal inhibition of migration following cargo uptake. Sphingolipid synthesis played a central role in these phenotypes by driving plasma membrane accumulation in cells lacking Gβ4. In Gβ4 knockout mice, neutrophils not only exhibited enhanced phagocytosis of inhaled fungal conidia in the lung but also increased trafficking of engulfed pathogens to other organs. Together, these results reveal an unexpected, biophysical control mechanism central to myeloid functional decision-making.

Influenza D virus (IDV) utilizes bovines as a primary reservoir with periodical spillover to other hosts. We have previously demonstrated that IDV binds both 9-O-acetylated N-acetylneuraminic acid (Neu5,9Ac2) and 9-O-acetylated N-glycolylneuraminic acid (Neu5Gc9Ac). Bovines produce both Neu5,9Ac2 and Neu5Gc9Ac, while humans are genetically unable to synthesize Neu5Gc9Ac. 9-O-Acetylation of sialic acids is catalyzed by CASD1 via a covalent acetyl-enzyme intermediate. To characterize the role of Neu5,9Ac2 and Neu5Gc9Ac in IDV infection and determine which form of 9-O-acetylated sialic acids drives IDV entry, we took advantage of a CASD1 knockout (KO) MDCK cell line and carried out feeding experiments using synthetic 9-O-acetyl sialic acids in combination with the single-round and multi-round IDV infection assays. The data from our studies show that (i) CASD1 KO cells are resistant to IDV infection and lack of IDV binding to the cell surface is responsible for the failure of IDV replication; (ii) feeding CASD1 KO cells with Neu5,9Ac2 or Neu5Gc9Ac resulted in a dose-dependent rescue of IDV infectivity; and (iii) diverse IDVs replicated robustly in CASD1 KO cells fed with either Neu5,9Ac2 or Neu5Gc9Ac at a level similar to that in wild-type cells with a functional CASD1. These data demonstrate that IDV can utilize Neu5,9Ac2- or non-human Neu5Gc9Ac-containing glycan receptor for infection. Our findings provide evidence that IDV has acquired the ability to infect and transmit among agricultural animals that are enriched in Neu5Gc9Ac, in addition to posing a zoonotic risk to humans expressing only Neu5,9Ac2.IMPORTANCEInfluenza D virus (IDV) has emerged as a multiple-species-infecting pathogen with bovines as a primary reservoir. Little is known about the functional receptor that drives IDV entry and promotes its cross-species spillover potential among different hosts. Here, we demonstrated that IDV binds exclusively to 9-O-acetylated N-acetylneuraminic acid (Neu5,9Ac2) and non-human 9-O-acetylated N-glycolylneuraminic acid (Neu5Gc9Ac) and utilizes both for entry and infection. This ability in effective engagement of both 9-O-acetylated sialic acids as functional receptors for infection provides an evolutionary advantage to IDV for expanding its host range. This finding also indicates that IDV has the potential to emerge in humans because Neu5,9Ac2 is ubiquitously expressed in human tissues, including lung. Thus, results of our study highlight a need for continued surveillance of IDV in humans, as well as for further investigation of its biology and cross-species transmission mechanism.

The plasma membrane is enriched for receptors and signaling proteins that are accessible from the extracellular space for pharmacological intervention. Here we conducted a series of CRISPR screens using human cell surface proteome and integrin family libraries in multiple cancer models. Our results identified ITGAV (integrin αV) and its heterodimer partner ITGB5 (integrin β5) as the essential integrin α/β pair for cancer cell expansion. High-density CRISPR gene tiling further pinpointed the integral pocket within the β-propeller domain of ITGAV for integrin αVβ5 dimerization. Combined with in silico compound docking, we developed a CRISPR-Tiling-Instructed Computer-Aided (CRISPR-TICA) pipeline for drug discovery and identified Cpd_AV2 as a lead inhibitor targeting the β-propeller central pocket of ITGAV. Cpd_AV2 treatment led to rapid uncoupling of integrin αVβ5 and cellular apoptosis, providing a unique class of therapeutic action that eliminates the integrin signaling via heterodimer dissociation. We also foresee the CRISPR-TICA approach to be an accessible method for future drug discovery studies.

Many G protein-coupled receptors (GPCRs) initiate a second phase of stimulatory heterotrimeric G protein (Gs)-coupled cAMP signaling after endocytosis. The prevailing current view is that the endosomal signal is inherently β-arrestin-dependent because β-arrestin is necessary for receptor internalization and, for some GPCRs, to prolong the endosomal signal. Here we revise this view by showing that the vasoactive intestinal peptide receptor 1 (VIPR1), a secretin-family polypeptide hormone receptor, does not require β-arrestin to internalize or to generate an endosomal signal. β-Arrestin instead resolves the plasma membrane and endosomal signaling phases into sequential cAMP peaks by desensitizing the plasma membrane phase without affecting the endosomal phase. This appears to occur through the formation of functionally distinct VIPR1-β-arrestin complexes at each location that differ in their phosphorylation dependence. We conclude that endosomal GPCR signaling can occur in the absence of β-arrestin and that β-arrestin sculpts the spatiotemporal profile of cellular GPCR-G protein signaling through location-specific remodeling of GPCR-β-arrestin complexes.

Aster proteins mediate the nonvesicular transport of cholesterol from the plasma membrane (PM) to the endoplasmic reticulum (ER). However, the importance of nonvesicular sterol movement for physiology and pathophysiology in various tissues is incompletely understood. Here we show that loss of Aster-B leads to diet-induced obesity in female but not in male mice, and that this sex difference is abolished by ovariectomy. We further demonstrate that Aster-B deficiency impairs nonvesicular cholesterol transport from the PM to the ER in ovaries in vivo, leading to hypogonadism and reduced estradiol synthesis. Female Aster-B-deficient mice exhibit reduced locomotor activity and energy expenditure, consistent with established effects of estrogens on systemic metabolism. Administration of exogenous estradiol ameliorates the diet-induced obesity phenotype of Aster-B-deficient female mice. These findings highlight the key role of Aster-B-dependent nonvesicular cholesterol transport in regulating estradiol production and protecting females from obesity.

Different exogenous electric fields (EF) can guide cell migration, disrupt proliferation, and program cell development. Studies have shown that many of these processes were initiated at the cell membrane, but the mechanism has been unclear, especially for conventionally non-excitable cells. In this study, we focus on the electrostatic aspects of EF coupling with the cell membrane by eliminating Faradaic processes using dielectric-coated microelectrodes. Our data unveil a distinctive biphasic response of the ERK signaling pathway of epithelial cells (MCF10A) to alternate current (AC) EF. The ERK signal exhibits both inhibition and activation phases, with the former triggered by a lower threshold of AC EF, featuring a swifter peaking time and briefer refractory periods than the later-occurring activation phase, induced at a higher threshold. Interestingly, the biphasic ERK responses are sensitive to the waveform and timing of EF stimulation pulses, depicting the characteristics of electrostatic and dissipative interactions. Blocker tests and correlated changes of active Ras on the cell membrane with ERK signals indicated that both EGFR and Ras were involved in the rich ERK dynamics induced by EF. We propose that the frequency-dependent dielectric relaxation process could be an important mechanism to couple EF energy to the cell membrane region and modulate membrane protein-initiated signaling pathways, which can be further explored to precisely control cell behavior and fate with high temporal and spatial resolution.

Androgen binding to the androgen receptor (AR) in the cytoplasm induces the AR to translocate to the nucleus, where it regulates the expression of target genes. Here, we found that androgens rapidly activated a plasma membrane-associated signaling node that enhanced nuclear AR functions. In murine primary osteoblasts, dihydrotestosterone (DHT) binding to a membrane-associated form of AR stimulated plasma membrane-associated protein kinase G type 2 (PKG2), leading to the activation of multiple kinases, including ERK. Phosphorylation of AR at Ser515 by ERK increased the nuclear accumulation and binding of AR to the promoter of Ctnnb1, which encodes the transcription factor β-catenin. In male mouse osteoblasts and human prostate cancer cells, DHT induced the expression of Ctnnb1 and CTNN1B, respectively, as well as β-catenin target genes, stimulating the proliferation, survival, and differentiation of osteoblasts and the proliferation of prostate cancer cells in a PKG2-dependent fashion. Because β-catenin is a master regulator of skeletal homeostasis, these results explain the reported male-specific osteoporotic phenotype of mice lacking PKG2 in osteoblasts and imply that PKG2-dependent AR signaling is essential for maintaining bone mass in vivo. Our results suggest that widely used pharmacological PKG activators, such as sildenafil, could be beneficial for male and estrogen-deficient female patients with osteoporosis but detrimental in patients with prostate cancer.

Invading microbes face a myriad of cidal mechanisms of phagocytes that inflict physical damage to microbial structures. How intracellular bacterial pathogens adapt to these stresses is not fully understood. Here, we report the discovery of a virulence mechanism by which changes to the mechanical stiffness of the mycobacterial cell surface confer refraction to killing during infection. Long-term time-lapse atomic force microscopy was used to reveal a process of "mechanical morphotype switching" in mycobacteria exposed to host intracellular stress. A "soft" mechanical morphotype switch enhances tolerance to intracellular macrophage stress, including cathelicidin. Both pharmacologic treatment, with bedaquiline, and a genetic mutant lacking uvrA modified the basal mechanical state of mycobacteria into a soft mechanical morphotype, enhancing survival in macrophages. Our study proposes microbial cell mechanical adaptation as a critical axis for surviving host-mediated stressors.

The HIV-1 accessory protein Nef hijacks clathrin adaptors to degrade or mislocalize host proteins involved in antiviral defenses. Here, using quantitative live-cell microscopy in genome-edited Jurkat cells, we investigate the impact of Nef on clathrin-mediated endocytosis (CME), a major pathway for membrane protein internalization in mammalian cells. Nef is recruited to CME sites on the plasma membrane, and this recruitment is associated with an increase in the recruitment and lifetime of the CME coat protein AP-2 and the late-arriving CME protein dynamin2. Furthermore, we find that CME sites that recruit Nef are more likely to recruit dynamin2 and transferrin, suggesting that Nef recruitment to CME sites promotes site maturation to ensure high efficiency in host protein downregulation. Implications of these observations for HIV-1 infection are discussed.

Planar cell polarity (PCP) proteins coordinate tissue morphogenesis by governing cell patterning and polarity. Asymmetrically localized on the plasma membrane of cells, transmembrane PCP proteins are trafficked by endocytosis, suggesting they may have intracellular functions that are dependent or independent of their extracellular role, but whether these functions extend to transcriptional control remains unknown. Here, we show the nuclear localization of transmembrane, PCP protein, VANGL2, in the HCC1569 breast cancer cell line, and in undifferentiated, but not differentiated, HC11 cells that serve as a model for mammary lactogenic differentiation. The loss of Vangl2 function results in upregulation of pathways related to STAT5 signaling. We identify DNA binding sites and a nuclear localization signal in VANGL2, and use CUT&RUN to demonstrate recruitment of VANGL2 to specific DNA binding motifs, including one in the Stat5a promoter. Knockdown (KD) of Vangl2 in HC11 cells and primary mammary organoids results in upregulation of Stat5a, Ccnd1 and Csn2, larger acini and organoids, and precocious differentiation; phenotypes are rescued by overexpression of Vangl2, but not Vangl2ΔNLS. Together, these results advance a paradigm whereby PCP proteins coordinate tissue morphogenesis by keeping transcriptional programs governing differentiation in check.

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Drug-carrying nanoparticles can be recognized and captured by macrophages and cleared away by the immune system, resulting in reduced drug efficacy and representing the main drawbacks. Biomimetic nanoparticles, which are coated with cell membranes from natural resources, have been applied to address this problem. This type of nanoparticle maintains some specific biological activities, allowing them to carry drugs reaching designated tissues effectively and have a longer time in circulation. This review article aims to summarize recent progress on biomimetic nanoparticles based on cell membranes. In this paper, we have introduced the classification of biomimetic nanoparticles, their preparation and characterization, and their applications in inflammatory diseases and malignant tumors. We have also analyzed the shortcomings and prospects of this technology, hoping to provide some clues for basic researchers and clinicians engaged in this field. [ABSTRACT FROM AUTHOR]

Surgical resection remains the primary treatment modality for glioblastoma (GBM); however, the infiltrative nature of GBM margins complicates achieving complete tumor removal. Additionally, the blood-brain barrier (BBB) poses a formidable challenge to effective probe delivery, thereby hindering precise imaging-guided surgery. Here, we introduce hybrid cell membrane-coated indocyanine green (ICG) liposomes (HM-Lipo-ICG) as biomimetic near-infrared (NIR) fluorescent probes for targeted BBB penetration and accurate delineation of infiltrative GBM margins. HM-Lipo-ICG encapsulates clinically approved ICG within its core and utilizes a hybrid cell membrane exterior, enabling specific targeting and enhanced BBB permeation. Quantitative assessments demonstrate that HM-Lipo-ICG achieves BBB penetration efficiency 2.8 times higher than conventional ICG liposomes. Mechanistically, CD44 receptor-mediated endocytosis facilitates BBB translocation of HM-Lipo-ICG. Furthermore, HM-Lipo-ICG enables high-contrast NIR imaging, achieving a signal-to-background ratio of 6.5 in GBM regions of an orthotopic glioma mouse model, thereby improving tumor margin detection accuracy fourfold (84.4% vs. 22.7%) compared to conventional ICG liposomes. Application of HM-Lipo-ICG facilitates fluorescence-guided precision surgery, resulting in complete resection of GBM cells. This study underscores the potential of hybrid cell membrane-coated liposomal probes in precisely visualizing and treating infiltrative GBM margins. [ABSTRACT FROM AUTHOR]

The cell (plasma) membrane is enriched with numerous receptors, ligands, enzymes, and phospholipids that play important roles in cell-cell and cell-extracellular matrix interactions governing, for instance, tissue development and repair. We previously showed that plasma membrane nanofragments (PMNFs) act as nucleation sites for bone formation in vivo , and induce in vitro mineralization within 1 day. In this study, we optimized the methods for generating, isolating, and applying PMNFs as a cell-free therapeutic to expedite bone defect repair. The PMNFs were isolated from different mouse cell lines (chondrocytes, osteoblasts, and fibroblasts), pre-conditioned, lyophilized, and subsequently transplanted into 2 mm critical-sized calvarial defects in mice (n = 75). The PMNFs from chondrocytes, following a 3-day pre-incubation, significantly accelerated bone repair within 2 weeks, through a coordinated attraction of macrophages, endothelial cells, and osteoblasts to the healing site. In vitro experiments confirmed that PMNFs enhanced cell adhesion. Comparison of the PMNF efficacy with phosphatidylserine, amorphous calcium phosphate (ACP), and living cells confirmed the unique ability of PMNFs to promote accelerated bone repair. Importantly, PMNFs promoted nearly complete integration of the regenerated bone with native tissue after 6 weeks (% non-integrated bone area = 15.02), contrasting with the partial integration (% non-integrated bone area = 56.10; p < 0.01, Student's test) with transplantation of ACP. Vickers microhardness tests demonstrated that the regenerated bone after 6 weeks (30.10 ± 1.75) exhibited hardness similar to native bone (31.07 ± 2.46). In conclusion, this is the first study to demonstrate that cell membrane can be a promising cell-free material with multifaceted biofunctional properties that promote accelerated bone repair. • For the first time, the cell (plasma) membrane is used as material for bone repair. • The plasma membrane from chondrocytes promoted rapid bone repair within 2 weeks, with complete integration of the newly formed bone with the surrounding native tissue. • The plasma membrane from chondrocytes presented multifaceted effects, strongly promoting the attraction of macrophages, endothelial cells and osteoblasts to the healing site. [Display omitted] [ABSTRACT FROM AUTHOR]

Liquid-liquid phase separation of proteins occurs on both surfaces of cellular membranes during diverse physiological processes. In vitro reconstitution could provide insight into the mechanisms underlying these events. However, most existing reconstitution techniques provide access to only one membrane surface, making it difficult to probe transmembrane phenomena. To study protein phase separation simultaneously on both membrane surfaces, we developed an array of freestanding planar lipid membranes. Interestingly, we observed that liquid-like protein condensates on one side of the membrane colocalized with those on the other side, resulting in transmembrane coupling. Our results, based on lipid probe partitioning and mobility of lipids, suggest that protein condensates locally reorganize membrane lipids, a process which could be explained by multiple effects. These findings suggest a mechanism by which signals originating on one side of a biological membrane, triggered by protein phase separation, can be transferred to the opposite side.

The limitations of traditional drug therapy have driven the creation and development of novel cell membrane-coated nanoparticle (CMNP) platforms. Since the introduction of the CMNP concept and method in 2011, an increasing number of studies focusing on this field have been widely conducted. Despite the growing body of literature, comprehensive bibliometric analysis in this field is still lacking. This study conducted a bibliometric analysis of CMNP-related publications sourced from the Web of Science Core Collection database, covering the period from January 1, 2011, to December 31, 2023. The analysis included co-authorships, co-citations, and co-occurrences of countries, institutions, authors, references, and keywords. Visualized tools such as Citespace, VOSviewer, and R Package Bibliometrix were employed to present the data. A total of 780 studies were included, with China contributing the highest number of publications (75.64%, n = 590). The number of annual publications increased consistently from 2011 to 2023, indicating a growing global interest in the CMNP field. Prof. Liangfang Zhang from the United States is recognized as the founder and leading figure in this area. The top three academic journals in this field, based on publication volume, are ACS Nano (32 publications, IF 2022 = 17.1), ACS Applied Materials Interfaces (32 publications, IF 2022 = 9.5), and Advanced Functional Materials (31 publications, IF 2022 = 19) among 185 scholarly journals. Reference and keyword analysis revealed that erythrocytes and macrophage membranes are significant research hotspots. The primary diseases targeted by CMNP research are cancer and pulmonary inflammation. In addition, CMNPs are frequently studied in conjunction with photothermal and photodynamic therapy. Furthermore, this study also summarized the timelines for various cell membrane coating methods and the three-step preparation process for CMNP. This comprehensive bibliometric analysis provides valuable insights to guide future research in the CMNP field, highlighting the importance of clinical application. Research on cell membrane-coated nanomaterials, particularly those related to cancer and pulmonary inflammation, is expected to remain a focal point. In addition, there is a need for the further development of other potential cell membrane-coated nanomaterials. This bibliometric analysis serves as a resource for researchers to quickly and comprehensively understand the current hotspots and emerging frontiers in this field.

Abstract Drug-carrying nanoparticles can be recognized and captured by macrophages and cleared away by the immune system, resulting in reduced drug efficacy and representing the main drawbacks. Biomimetic nanoparticles, which are coated with cell membranes from natural resources, have been applied to address this problem. This type of nanoparticle maintains some specific biological activities, allowing them to carry drugs reaching designated tissues effectively and have a longer time in circulation. This review article aims to summarize recent progress on biomimetic nanoparticles based on cell membranes. In this paper, we have introduced the classification of biomimetic nanoparticles, their preparation and characterization, and their applications in inflammatory diseases and malignant tumors. We have also analyzed the shortcomings and prospects of this technology, hoping to provide some clues for basic researchers and clinicians engaged in this field.

Abstract Surgical resection remains the primary treatment modality for glioblastoma (GBM); however, the infiltrative nature of GBM margins complicates achieving complete tumor removal. Additionally, the blood-brain barrier (BBB) poses a formidable challenge to effective probe delivery, thereby hindering precise imaging-guided surgery. Here, we introduce hybrid cell membrane-coated indocyanine green (ICG) liposomes (HM-Lipo-ICG) as biomimetic near-infrared (NIR) fluorescent probes for targeted BBB penetration and accurate delineation of infiltrative GBM margins. HM-Lipo-ICG encapsulates clinically approved ICG within its core and utilizes a hybrid cell membrane exterior, enabling specific targeting and enhanced BBB permeation. Quantitative assessments demonstrate that HM-Lipo-ICG achieves BBB penetration efficiency 2.8 times higher than conventional ICG liposomes. Mechanistically, CD44 receptor-mediated endocytosis facilitates BBB translocation of HM-Lipo-ICG. Furthermore, HM-Lipo-ICG enables high-contrast NIR imaging, achieving a signal-to-background ratio of 6.5 in GBM regions of an orthotopic glioma mouse model, thereby improving tumor margin detection accuracy fourfold (84.4% vs. 22.7%) compared to conventional ICG liposomes. Application of HM-Lipo-ICG facilitates fluorescence-guided precision surgery, resulting in complete resection of GBM cells. This study underscores the potential of hybrid cell membrane-coated liposomal probes in precisely visualizing and treating infiltrative GBM margins. Graphical abstract

Human Hsp70-escort protein 1 (hHep1) is a cochaperone that assists in the function and stability of mitochondrial HSPA9. Similar to HSPA9, hHep1 is located outside the mitochondria and can interact with liposomes. In this study, we further investigated the structural and thermodynamic behavior of interactions between hHep1 and negatively charged liposomes, as well as interactions with cellular membranes. Our results showed that hHep1 interacts peripherally with liposomes formed by phosphatidylserine and cardiolipin and remains partially structured, exhibiting similar affinities for both. In addition, after being added to the cell membrane, recombinant hHep1 was incorporated by cells in a dose-dependent manner. Interestingly, the association of HSPA9 with hHep1 improved the incorporation of these proteins into the lipid bilayer. These results demonstrated that hHep1 can interact with lipids also present in the plasma membrane, indicating roles for this cochaperone outside of mitochondria.

Cells execute specific responses to diverse environmental cues by encoding information in distinctly compartmentalized biochemical signalling reactions. Genetically encoded fluorescent biosensors enable the spatial and temporal monitoring of signalling events in live cells. Temporal and spatiotemporal computational models can be used to interpret biosensor experiments in complex biochemical networks and to explore hypotheses that are difficult to test experimentally. In this review, we first provide brief discussions of the experimental toolkit of fluorescent biosensors as well as computational basics with a focus on temporal and spatiotemporal deterministic models. We then describe how we used this combined approach to identify and investigate a protein kinase A (PKA) - cAMP - Ca2+ oscillatory circuit in MIN6 β cells, a mouse pancreatic β cell system. We describe the application of this combined approach to interrogate how this oscillatory circuit is differentially regulated in a nano-compartment formed at the plasma membrane by the scaffolding protein A kinase anchoring protein 79/150. We leveraged both temporal and spatiotemporal deterministic models to identify the key regulators of this oscillatory circuit, which we confirmed with further experiments. The powerful approach of combining live-cell biosensor imaging with quantitative modelling, as discussed here, should find widespread use in the investigation of spatiotemporal regulation of cell signalling.

Candidate cis-regulatory elements (cCREs) in microglia demonstrate the most substantial enrichment for Alzheimer's disease (AD) heritability compared to other brain cell types. However, whether and how these genome-wide association studies (GWAS) variants contribute to AD remain elusive. Here we prioritize 308 previously unreported AD risk variants at 181 cCREs by integrating genetic information with microglia-specific 3D epigenome annotation. We further establish the link between functional variants and target genes by single-cell CRISPRi screening in microglia. In addition, we show that AD variants exhibit allelic imbalance on target gene expression. In particular, rs7922621 is the effective variant in controlling TSPAN14 expression among other nominated variants in the same cCRE and exerts multiple physiological effects including reduced cell surface ADAM10 and altered soluble TREM2 (sTREM2) shedding. Our work represents a systematic approach to prioritize and characterize AD-associated variants and provides a roadmap for advancing genetic association to experimentally validated cell-type-specific phenotypes and mechanisms.

The cell membrane has a fundamental role in the cell life cycle but there's still much to be learned about its heterogeneous structure, regulation, and protein interaction. Additionally, the protein-membrane interaction is often overlooked when studying specific protein dynamics. In this work, we present a new tool for a better understanding of protein dynamics and membrane function using live cells and fast non-invasive techniques without the need for individual particle tracking. To this end, we used the 2D-pair correlation function (2D-pCF) to study protein interactions across cellular membranes. We performed numerical simulations and confocal experiments using a GAP-mEGFP fusion construct known to interact with the plasmatic membrane. Our results demonstrate that based on a quantitative correlation analysis as the 2D pair correlation of the signal intensities, is possible to characterize protein-membrane interactions in live systems and real-time. Combining experimental and numerical results this work presents a new powerful approach to the study of the dynamic protein-membrane interaction.

Protein-membrane interactions (PMIs) are ubiquitous in cellular signaling. Initial steps of signal transduction cascades often rely on transient and dynamic interactions with the inner plasma membrane leaflet to populate and regulate signaling hotspots. Methods to target and modulate these interactions could yield attractive tool compounds and drug candidates. Here, we demonstrate that the conjugation of a medium-chain lipid tail to the covalent K-Ras(G12C) binder MRTX849 at a solvent-exposed site enables such direct modulation of PMIs. The conjugated lipid tail interacts with the tethered membrane and changes the relative membrane orientation and conformation of K-Ras(G12C), as shown by molecular dynamics (MD) simulation-supported NMR studies. In cells, this PMI modulation restricts the lateral mobility of K-Ras(G12C) and disrupts nanoclusters. The described strategy could be broadly applicable to selectively modulate transient PMIs.

Cell membrane-based nanovaccines have demonstrated attractive features due to their inherently multiantigenic nature and ability to be formulated with adjuvants. Here, we report on cellular nanodiscs fabricated from cancer cell membranes and incorporated with a lipid-based adjuvant for antitumor vaccination. The cellular nanodiscs, with their small size and discoidal shape, are readily taken up by antigen-presenting cells and drain efficiently to the lymph nodes. Due to its highly immunostimulatory properties, the nanodisc vaccine effectively stimulates the immune system and promotes tumor-specific immunity. Using a murine colorectal cancer model, strong control of tumor growth is achieved in both prophylactic and therapeutic settings, particularly in combination with checkpoint blockades. Considerable therapeutic efficacy is also observed in treating a weakly immunogenic metastatic melanoma model. This work presents a new paradigm for the design of multiantigenic nanovaccines that can effectively activate antitumor immune responses and may be applicable to a wide range of cancers.

Ligation of retinoic acid receptor alpha (RARα) by RA promotes varied transcriptional programs associated with immune activation and tolerance, but genetic deletion approaches suggest the impact of RARα on TCR signaling. Here, we examined whether RARα would exert roles beyond transcriptional regulation. Specific deletion of the nuclear isoform of RARα revealed an RARα isoform in the cytoplasm of T cells. Extranuclear RARα was rapidly phosphorylated upon TCR stimulation and recruited to the TCR signalosome. RA interfered with extranuclear RARα signaling, causing suboptimal TCR activation while enhancing FOXP3+ regulatory T cell conversion. TCR activation induced the expression of CRABP2, which translocates RA to the nucleus. Deletion of Crabp2 led to increased RA in the cytoplasm and interfered with signalosome-RARα, resulting in impaired anti-pathogen immunity and suppressed autoimmune disease. Our findings underscore the significance of subcellular RA/RARα signaling in T cells and identify extranuclear RARα as a component of the TCR signalosome and a determinant of immune responses.

γδ T cells are potent anticancer effectors with the potential to target tumours broadly, independent of patient-specific neoantigens or human leukocyte antigen background1-5. γδ T cells can sense conserved cell stress signals prevalent in transformed cells2,3, although the mechanisms behind the targeting of stressed target cells remain poorly characterized. Vγ9Vδ2 T cells-the most abundant subset of human γδ T cells4-recognize a protein complex containing butyrophilin 2A1 (BTN2A1) and BTN3A1 (refs. 6-8), a widely expressed cell surface protein that is activated by phosphoantigens abundantly produced by tumour cells. Here we combined genome-wide CRISPR screens in target cancer cells to identify pathways that regulate γδ T cell killing and BTN3A cell surface expression. The screens showed previously unappreciated multilayered regulation of BTN3A abundance on the cell surface and triggering of γδ T cells through transcription, post-translational modifications and membrane trafficking. In addition, diverse genetic perturbations and inhibitors disrupting metabolic pathways in the cancer cells, particularly ATP-producing processes, were found to alter BTN3A levels. This induction of both BTN3A and BTN2A1 during metabolic crises is dependent on AMP-activated protein kinase (AMPK). Finally, small-molecule activation of AMPK in a cell line model and in patient-derived tumour organoids led to increased expression of the BTN2A1-BTN3A complex and increased Vγ9Vδ2 T cell receptor-mediated killing. This AMPK-dependent mechanism of metabolic stress-induced ligand upregulation deepens our understanding of γδ T cell stress surveillance and suggests new avenues available to enhance γδ T cell anticancer activity.

Abstract Glioblastoma (GBM), one of the deadliest brain tumors, accounts for approximately 50% of all primary malignant CNS tumors, therefore novel, highly effective remedies are urgently needed. Boron neutron capture therapy, which has recently repositioned as a promising strategy to treat high-grade gliomas, requires a conspicuous accumulation of boron atoms in the cancer cells. With the aim of selectively deliver sodium borocaptate (BSH, a 12 B atoms-including molecule already employed in the clinics) to GBM cells, we developed novel cell membrane-derived vesicles (CMVs), overcoming the limits of natural extracellular vesicles as drug carriers, while maintaining their inherent homing abilities that make them preferable to fully synthetic nanocarriers. Purified cell membrane fragments, isolated from patient-derived GBM stem-like cell cultures, were used to prepare nanosized CMVs, which retained some membrane proteins specific of the GBM parent cells and were devoid of potentially detrimental genetic material. In vitro tests evidenced the targeting ability of this novel nanosystem and ruled out any cytotoxicity. The CMVs were successfully loaded with BSH, by following two different procedures, i.e. sonication and electroporation, demonstrating their potential applicability in GBM therapy.

Abstract In therapies, curcumin is now commonly formulated in liposomal form, administered through injections or creams. This enhances its concentration at the cellular level compared to its natural form ingestion. Due to its hydrophobic nature, curcumin is situated in the lipid part of the membrane, thereby modifying its properties and influencing processes The aim of the research was to investigate whether the toxicity of specific concentrations of curcumin, assessed through biochemical tests for the SK-N-SH and H-60 cell lines, is related to structural changes in the membranes of these cells, caused by the localization of curcumin in their hydrophobic regions. Biochemical tests were performed using spectrophotometric methods. Langmuir technique were used to evaluate the interaction of the curcumin with the studied lipids. Direct introduction of curcumin into the membranes alters their physicochemical parameters. The extent of these changes depends on the initial properties of the membrane. In the conducted research, it has been demonstrated that curcumin may exhibit toxicity to human cells. The mechanism of this toxicity is related to its localization in cell membranes, leading to their dysfunction. The sensitivity of cells to curcumin presence depends on the saturation level of their membranes; the more rigid the membrane, the lower the concentration of curcumin causes its disruption.

Yao Wang,1,&ast; Mingtang Zeng,1,&ast; Tianfei Fan,1 Ming Jia,2 Ruxi Yin,1 Jia Xue,1 Longjun Xian,3 Ping Fan,1 Mei Zhan1 1Department of Pharmacy, West China Hospital, Sichuan University, Chengdu, People’s Republic of China; 2Nanchong Institute for Food and Drug Control, Nanchong, People’s Republic of China; 3Department of Thoracic Surgery, Institute of Thoracic Oncology, Frontiers Science Center for Disease-Related Molecular Network, West China Hospital, Sichuan University, Chengdu, Sichuan, People’s Republic of China&ast;These authors contributed equally to this workCorrespondence: Ping Fan; Mei Zhan, Department of Pharmacy, West China Hospital, Sichuan University, Chengdu, 610041, People’s Republic of China, Email m13980602288@163.com; mandyzhanmei@163.comAbstract: Metal-organic frameworks (MOFs) are porous materials resulting from the coordination of metal clusters or ions with organic ligands, merging macromolecular and coordination chemistry features. Among these, zeolitic imidazolate framework-8 (ZIF-8) stands out as a widely utilized MOF known for its robust stability in aqueous environments owing to the robust interaction between its constituent zinc ions (Zn2+) and 2-methylimidazole (2-MIM). ZIF-8 readily decomposes under acidic conditions, serving as a promising candidate for pH-responsive drug delivery systems. Moreover, biomimetic materials typically possess good biocompatibility, reducing immune reactions. By mimicking natural structures or surface features within the body, they enhance the targeting of nanoparticles, prolong their circulation time, and increase their bioavailability in vivo. This review explores the latest advancements in biomimetic ZIF-8 nanoparticles for drug delivery, elucidating the primary obstacles and future prospects in utilizing ZIF-8 for drug delivery applications.Keywords: MOF, ZIF-8, cell membrane, biomimetic drug delivery

Blood transfusions save lives and improve health every day. Despite the matching of blood types being stricter than it ever has been, emergency transfusions among incompatible blood types are still inevitable in the clinic when there is a lack of acceptable blood types for recipients. Here to overcome this, a counter measure nanoplatform consisting of a polymeric core coated by a red blood cell (RBC) membrane is developed. With A‐type or B‐type RBC membrane camouflaging, the nanoplatform is capable of specifically capturing anti‐A or anti‐B IgM antibodies within B‐type or A‐type whole blood, thereby decreasing the corresponding IgM antibody levels and then allowing the incompatible blood transfusions. In addition to IgM, the anti‐RBC IgG antibody in a passive immunization murine model can likewise be neutralized by this nanoplatform, leading to prolonged circulation time of incompatible donor RBCs. Noteworthily, nanoplatform made by expired RBCs (>42 days stored hypothermically) and then subjected to lyophilization does not impair their effect on antibody neutralization. Most importantly, antibody‐captured RBC‐NP do not exacerbate the risk of inflammation, complement activation, and coagulopathy in an acute hemorrhagic shock murine model. Overall, this biomimetic nanoplatform can safely neutralize the antibody to enable incompatible blood transfusion. [ABSTRACT FROM AUTHOR]

Ischemic stroke is a serious neurological disease involving multiple complex physiological processes, including vascular obstruction, brain tissue ischemia, impaired energy metabolism, cell death, impaired ion pump function, and inflammatory response. In recent years, there has been significant interest in cell membrane-functionalized biomimetic nanoparticles as a novel therapeutic approach. This review comprehensively explores the mechanisms and importance of using these nanoparticles to treat acute ischemic stroke with a special emphasis on their potential for actively targeting therapies through cell membranes. We provide an overview of the pathophysiology of ischemic stroke and present advances in the study of biomimetic nanoparticles, emphasizing their potential for drug delivery and precision-targeted therapy. This paper focuses on bio-nanoparticles encapsulated in bionic cell membranes to target ischemic stroke treatment. It highlights the mechanism of action and research progress regarding different types of cell membrane-functionalized bi-onic nanoparticles such as erythrocytes, neutrophils, platelets, exosomes, macrophages, and neural stem cells in treating ischemic stroke while emphasizing their potential to improve brain tissue's ischemic state and attenuate neurological damage and dysfunction. Through an in-depth exploration of the potential benefits provided by cell membrane-functionalized biomimetic nanoparticles to improve brain tissue's ischemic state while reducing neurological injury and dysfunction, this study also provides comprehensive research on neural stem cells' potential along with that of cell membrane-functionalized biomimetic nanoparticles to ameliorate neurological injury and dysfunction. However, it is undeniable that there are still some challenges and limitations in terms of biocompatibility, safety, and practical applications for clinical translation. [ABSTRACT FROM AUTHOR]

Protein FadR is known as a fatty acid metabolism global regulator that sustains cell envelope integrity by changing the profile of fatty acid. Here, we present its unique participation in the defense against reactive oxygen species (ROS) in the bacterium. FadR contributes to defending extracellular ROS by maintaining the permeability of the cell membrane. It also facilitates the ROS detoxification process by increasing the expression of ROS neutralizers (KatB, KatG, and AhpCF). FadR also represses the leakage of ROS by alleviating the respiratory action conducted by terminal cytochrome cbb3-type heme-copper oxidases (ccoNOQP). These findings suggest that FadR plays a comprehensive role in modulating the bacterial oxidative stress response, instead of merely strengthening the cellular barrier against the environment. This study sheds light on the complex mechanisms of bacterial ROS defense and offers FadR as a novel target for ROS control research. [ABSTRACT FROM AUTHOR]

In therapies, curcumin is now commonly formulated in liposomal form, administered through injections or creams. This enhances its concentration at the cellular level compared to its natural form ingestion. Due to its hydrophobic nature, curcumin is situated in the lipid part of the membrane, thereby modifying its properties and influencing processes The aim of the research was to investigate whether the toxicity of specific concentrations of curcumin, assessed through biochemical tests for the SK-N-SH and H-60 cell lines, is related to structural changes in the membranes of these cells, caused by the localization of curcumin in their hydrophobic regions. Biochemical tests were performed using spectrophotometric methods. Langmuir technique were used to evaluate the interaction of the curcumin with the studied lipids. Direct introduction of curcumin into the membranes alters their physicochemical parameters. The extent of these changes depends on the initial properties of the membrane. In the conducted research, it has been demonstrated that curcumin may exhibit toxicity to human cells. The mechanism of this toxicity is related to its localization in cell membranes, leading to their dysfunction. The sensitivity of cells to curcumin presence depends on the saturation level of their membranes; the more rigid the membrane, the lower the concentration of curcumin causes its disruption. [ABSTRACT FROM AUTHOR]

Context: Electroporation is a technique that creates electrically generated pores in the cell membrane by modifying transmembrane potential. In this work, the finite element method (FEM) was used to examine the induced transmembrane voltage (ITV) of a spherical-shaped MCF-7 cell, allowing researchers to determine the stationary ITV. A greater ITV than the critical value causes permeabilization of the membrane. Furthermore, the present study shows how a specific surface conductivity can act as a stand-in for the thin layer that constitutes a cell membrane as the barrier between extracellular and intracellular environments. Additionally, the distribution of ITV on the cell membrane and its maximum value were experimentally evaluated for a range of applied electric fields. Consequently, the entire cell surface area was electroporated 66% and 68% for molecular dynamics (MD) simulations and FEM, respectively, when the external electric field of 1500 V/cm was applied to the cell suspension using the previously indicated numerical methods. Furthermore, the lipid bilayers' molecular structure was changed, which led to the development of hydrophilic holes with a radius of 1.33 nm. Applying MD and FEM yielded threshold values for transmembrane voltage of 700 and 739 mV, respectively. Method: Using MD simulations of palmitoyloleoyl-phosphatidylcholine (POPC), pores in cell membranes exposed to external electric fields were numerically investigated. The dependence on the electric field was estimated and developed, and the amount of the electroporated cell surface area matches the applied external electric field. To investigate more, a mathematical model based on an adaptive neuro-fuzzy inference system (ANFIS) is employed to predict the percent cell viability of cancerous cells after applying four pulses during electroporation. For MD simulations, ArgusLab, VMD, and GROMACS software packages were used. Moreover, for FEM analysis, COMSOL software package was used. Also, it is worth mentioning that for mathematical model, MATLAB software is used. [ABSTRACT FROM AUTHOR]

Mechanosensitive mechanisms are often used to sense damage to tissue structure, stimulating matrix synthesis and repair. While this kind of mechanoregulatory process is well recognized in eukaryotic systems, it is not known whether such a process occurs in bacteria. In Vibrio cholerae, antibiotic-induced damage to the load-bearing cell wall promotes increased signaling by the two-component system VxrAB, which stimulates cell wall synthesis. Here we show that changes in mechanical stress within the cell envelope are sufficient to stimulate VxrAB signaling in the absence of antibiotics. We applied mechanical forces to individual bacteria using three distinct loading modalities: extrusion loading within a microfluidic device, direct compression and hydrostatic pressure. In all cases, VxrAB signaling, as indicated by a fluorescent protein reporter, was increased in cells submitted to greater magnitudes of mechanical loading, hence diverse forms of mechanical stimuli activate VxrAB signaling. Reduction in cell envelope stiffness following removal of the endopeptidase ShyA led to large increases in cell envelope deformation and substantially increased VxrAB response, further supporting the responsiveness of VxrAB. Our findings demonstrate a mechanosensitive gene regulatory system in bacteria and suggest that mechanical signals may contribute to the regulation of cell wall homeostasis.

Immune checkpoint immunotherapy (ICI) can re-activate immune reactions against neoantigens, leading to remarkable remission in cancer patients. Nevertheless, only a minority of patients are responsive to ICI, and approaches for prediction of responsiveness are needed to improve the success of cancer treatments. While the tumor mutational burden (TMB) correlates positively with responsiveness and survival of patients undergoing ICI, the influence of the subcellular localizations of the neoantigens remains unclear. Here, we demonstrate in both a mouse melanoma model and human clinical datasets of 1,722 ICI-treated patients that a high proportion of membrane-localized neoantigens, particularly at the plasma membrane, correlate with responsiveness to ICI therapy and improved overall survival across multiple cancer types. We further show that combining membrane localization and TMB analyses can enhance the predictability of cancer patient response to ICI. Our results may have important implications for establishing future clinical guidelines to direct the choice of treatment toward ICI.

Elucidating the structure and interactions of proteins in native environments is a fundamental goal of structural biology. Nuclear magnetic resonance (NMR) spectroscopy is well suited for this task but often suffers from low sensitivity, especially in complex biological settings. Here, we use a sensitivity-enhancement technique called dynamic nuclear polarization (DNP) to overcome this challenge. We apply DNP to capture the membrane interactions of the outer membrane protein Ail, a key component of the host invasion pathway of Yersinia pestis. We show that the DNP-enhanced NMR spectra of Ail in native bacterial cell envelopes are well resolved and enriched in correlations that are invisible in conventional solid-state NMR experiments. Furthermore, we demonstrate the ability of DNP to capture elusive interactions between the protein and the surrounding lipopolysaccharide layer. Our results support a model where the extracellular loop arginine residues remodel the membrane environment, a process that is crucial for host invasion and pathogenesis.

Fungal invasion of the oral epithelium is central to the pathogenesis of oropharyngeal candidiasis (OPC). Candida albicans invades the oral epithelium by receptor-induced endocytosis but this process is incompletely understood. We found that C. albicans infection of oral epithelial cells induces c-Met to form a multi-protein complex with E-cadherin and the epidermal growth factor receptor (EGFR). E-cadherin is necessary for C. albicans to activate both c-Met and EGFR and to induce the endocytosis of C. albicans. Proteomics analysis revealed that c-Met interacts with C. albicans Hyr1, Als3 and Ssa1. Both Hyr1 and Als3 are required for C. albicans to stimulate c-Met and EGFR in oral epithelial cells in vitro and for full virulence during OPC in mice. Treating mice with small molecule inhibitors of c-Met and EGFR ameliorates OPC, demonstrating the potential therapeutic efficacy of blocking these host receptors for C. albicans.

Supported lipid bilayers (SLBs) are commonly used to investigate interactions between cell membranes and their environment. These model platforms can be formed on electrode surfaces and analyzed using electrochemical methods for bioapplications. Carbon nanotube porins (CNTPs) integrated with SLBs have emerged as promising artificial ion channel platforms. In this study, we present the integration and ion transport characterization of CNTPs in in vivo environments. We combine experimental and simulation data obtained from electrochemical analysis to analyze the membrane resistance of the equivalent circuits. Our results show that carrying CNTPs on a gold electrode results in high conductance for monovalent cations (K+ and Na+) and low conductance for divalent cations (Ca2+).

Membrane tension is thought to be a long-range integrator of cell physiology. Membrane tension has been proposed to enable cell polarity during migration through front-back coordination and long-range protrusion competition. These roles necessitate effective tension transmission across the cell. However, conflicting observations have left the field divided as to whether cell membranes support or resist tension propagation. This discrepancy likely originates from the use of exogenous forces that may not accurately mimic endogenous forces. We overcome this complication by leveraging optogenetics to directly control localized actin-based protrusions or actomyosin contractions while simultaneously monitoring the propagation of membrane tension using dual-trap optical tweezers. Surprisingly, actin-driven protrusions and actomyosin contractions both elicit rapid global membrane tension propagation, whereas forces applied to cell membranes alone do not. We present a simple unifying mechanical model in which mechanical forces that engage the actin cortex drive rapid, robust membrane tension propagation through long-range membrane flows.

Through the magic of "active matter"─matter that converts chemical energy into mechanical work to drive emergent properties─biology solves a myriad of seemingly enormous physical challenges. Using active matter surfaces, for example, our lungs clear an astronomically large number of particulate contaminants that accompany each of the 10,000 L of air we respire per day, thus ensuring that the lungs' gas exchange surfaces remain functional. In this Perspective, we describe our efforts to engineer artificial active surfaces that mimic active matter surfaces in biology. Specifically, we seek to assemble the basic active matter components─mechanical motor, driven constituent, and energy source─to design surfaces that support the continuous operation of molecular sensing, recognition, and exchange. The successful realization of this technology would generate multifunctional, "living" surfaces that combine the dynamic programmability of active matter and the molecular specificity of biological surfaces and apply them to applications in biosensors, chemical diagnostics, and other surface transport and catalytic processes. We describe our recent efforts in bio-enabled engineering of living surfaces through the design of molecular probes to understand and integrate native biological membranes into synthetic materials.