Pore-forming Toxin Research Articles

Investigating pyroptosis as a mechanism of L. major cell-to-cell spread in the human BLaER1 infection model.

Leishmania is the causative agent of the tropical neglected disease leishmaniasis and infects macrophages as its definitive host cell. In order to sustain and propagate infections, Leishmania parasites have to complete cycles of exit and re-infection. Yet, the mechanism driving the parasite spread to other cells remains unclear. Recent studies reported pro-inflammatory monocytes as replicative niche of Leishmania major and showed prolonged expression of IL-1β at the site of infection, indicating an activation of the NLRP3 inflammasome and pointing toward pyroptosis as a possible mechanism of parasite spread. To address the species-specific inflammasome activation of human cells, we characterized the BLaER1 monocytes as a model for L. major infection. We found that BLaER1 monocytes support infection and activation by Leishmania parasites to the same extent as primary human macrophages. Harnessing the possibilities of this infection model, we first showed that BLaER1 GSDMD-/- cells, which carry a deletion of the pore-forming protein gasdermin D, are more resistant to pyroptotic cell death and, concomitantly, display a strongly delayed release of intracellular parasite. Using that knockout in a co-incubation assay in comparison with wild-type BLaER1 cells, we demonstrate that impairment of the pyroptosis pathway leads to lower rates of parasite spread to new host cells, thus, implicating pyroptotic cell death as a possible exit mechanism of L. major in pro-inflammatory microenvironments.

Remote limb ischemic postconditioning inhibits microglia pyroptosis by modulating HGF after acute ischemia stroke.

The repetitive inflation-deflation of a blood pressure cuff on a limb is known as remote limb ischemic postconditioning (RIPostC). It prevents brain damage induced by acute ischemia stroke (AIS). Pyroptosis, executed by the pore-forming protein gasdermin D (GSDMD), is a type of regulated cell death triggered by proinflammatory signals. It contributes to the pathogenesis of ischemic brain injury. However, the effects of RIPostC on pyroptosis following AIS remain largely unknown. In our study, linear correlation analysis confirmed that serum GSDMD levels in AIS patients upon admission were positively correlated with NIHSS scores. RIPostC treatment significantly reduced GSDMD level compared with patients without RIPostC at 3 days post-treatment. Besides, middle cerebral artery occlusion (MCAO) surgery was performed on C57BL/6 male mice and RIPostC was induced immediately after MCAO. We found that RIPostC suppressed the activation of NLRP3 inflammasome to reduce the maturation of GSDMD, leading to decreased pyroptosis in microglia after AIS. Hepatocyte growth factor (HGF) was identified using the high throughput screening. Importantly, HGF siRNA, exogenous HGF, and ISG15 siRNA were used to reveal that HGF/ISG15 is a possible mechanism of RIPostC regulation in vivo and in vitro.

Pyroptosis and gasdermins-Emerging insights and therapeutic opportunities in metabolic dysfunction-associated steatohepatitis.

In recent years, there has been a rapid expansion in our understanding of regulated cell death, leading to the discovery of novel mechanisms that govern diverse cell death pathways. One recently discovered type of cell death is pyroptosis, initially identified in the 1990s as a caspase-1-dependent lytic cell death. However, further investigations have redefined pyroptosis as a regulated cell death that relies on the activation of pore-forming proteins, particularly the gasdermin family. Among the key regulators of pyroptosis is the inflammasome sensor NOD-like receptor 3 (NLRP3), a critical innate immune sensor responsible for regulating the activation of caspase-1 and gasdermin D. A deeper understanding of pyroptosis and its interplay with other forms of regulated cell death is emerging, shedding light on a complex regulatory network controlling pore-forming proteins and cell fate. Cell death processes play a central role in diseases such as metabolic dysfunction-associated steatotic liver disease, metabolic dysfunction-associated steatohepatitis, autoinflammatory disorders, and cancer. Cell death often acts as a starting point in these diseases, making it an appealing target for drug development. Yet, the complete molecular mechanisms are not fully understood, and new discoveries reveal promising novel avenues for therapeutic interventions. In this review, we summarize recent evidence on pathways and proteins controlling pyroptosis and gasdermins. Furthermore, we will address the role of pyroptosis and the gasdermin family in metabolic dysfunction-associated steatotic liver disease and steatohepatitis. Additionally, we highlight new potential therapeutic targets for treating metabolic dysfunction-associated steatohepatitis and other inflammatory-associated diseases.

Effect of pH on Electrochemical Impedance Response of Tethered Bilayer Lipid Membranes: Implications for Quantitative Biosensing

Tethered bilayer lipid membranes (tBLMs) are increasingly used in biosensor applications where electrochemical impedance spectroscopy (EIS) is the method of choice for amplifying and recording the activity of membrane-damaging agents such as pore-forming toxins or disrupting peptides. While the activity of these biological agents may depend on the pH of the analytes, there is increasing evidence that the sensitivity of tethered bilayer sensors depends on the pH of the solutions. In our study, we addressed the question of what are the fundamental reasons for the variability of the EIS signal of the tBLMs with pH. We designed an experiment to compare the EIS response of tBLMs with natural membrane defects and two different membrane disruptors: vaginolysin and melittin. Our experimental design ensured that the same amount of protein and peptide was present in the tBLMs, while the pH was varied by replacing the buffers with different pH values. Using a recently developed EIS data analysis algorithm from our research group, we were able to demonstrate that, in contrast to previous literature which relates the variability of tBLM, EIS response to the variation in defect density, the main reason for the observed variability in EIS response is the change in the sub-membrane properties of tBLMs with pH. Using surface-enhanced infrared absorption spectroscopy (SEIRAS), we have shown that pH changes from neutral to slightly acidic leads to an expulsion of water, presumably bound to ions, from the sub-membrane reservoir, resulting in a marked decrease in the carrier concentration and specific conductance of the sub-membrane reservoir. Such a decrease is recorded by the EIS as a decrease in the conductance of the tBLM complex and affects the sensitivity of a biosensor. Our data provide important evidence of pH-sensitive effects that should be considered in both the development and operation of biosensors.

MCM8-mediated mitophagy protects vascular health in response to nitric oxide signaling in a mouse model of Kawasaki disease.

Mitophagy is a major quality control pathway that removes unwanted or dysfunctional mitochondria and plays an essential role in vascular health. Here we show that MCM8 expression is significantly decreased in children with Kawasaki disease (KD) who developed coronary artery aneurysms. Mechanistically, we discovered that nitric oxide signaling promotes TRIM21-mediated MCM8 ubiquitination, which disrupts its interaction with MCM9 and promotes its cytosolic export. In the cytosol, MCM8 relocates to the mitochondria pore-forming proteins and promotes their ubiquitination by TRIM21. In addition, MCM8 directly recruits LC3 via its LC3-interacting region (LIR) motif and initiates mitophagy. This suppresses mitochondrial DNA-mediated activation of type I interferon via cGAS and STING. Mice that are deficient in Mcm8, Trim21 and Nos2 or reconstituted with the East-Asian-specific MCM8-P276 variant develop more severe coronary artery vasculopathy in the Lactobacillus casei extract-induced KD model. Collectively, the data suggest that MCM8 protects vascular health in the KD setting.

Role of the Alteration in Calcium Homeostasis in Cell Death Induced by Clostridioides difficile Toxin A and Toxin B.

Clostridioides difficile (C. difficile), responsible for 15-25% of gastrointestinal infections, causes health problems mainly due to the toxic activity of toxins A and B (Tcds). These are responsible for its clinical manifestations, including diarrhea, pseudomembranous colitis, toxic megacolon and death, with a mortality of 5-30% in primary infection, that increase following relapses. Studies on Tcd-induced cell death have highlighted a key role of caspases, calpains, and cathepsins, with involvement of mitochondria and reactive oxygen species (ROS) in a complex signaling pathway network. The complex response in the execution of various types of cell death (apoptosis, necrosis, pyroptosis and pyknosis) depends on the amount of Tcd, cell types, and Tcd receptors involved, and could have as initial/precocious event the alterations in calcium homeostasis. The entities, peculiarities and cell types involved in these alterations will decide the signaling pathways activated and cell death type. Calcium homeostasis alterations can be caused by calcium influx through calcium channel activation, transient intracellular calcium oscillations, and leakage of calcium from intracellular stores. These increases in cytoplasmic calcium have important effects on all calcium-regulated molecules, which may play a direct role in several cell death types and/or activate other cell death effectors, such as caspases, calpains, ROS and proapoptotic Bcl-2 family members. Furthermore, some support for the possible role of the calcium homeostasis alteration in Tcd-induced cell death originates from the similarity with cytotoxic effects that cause pore-forming toxins, based mainly on calcium influx through plasma membrane pores.

Albumin pre-opsonized membrane-active iPep nanomedicine potentiates chemo to immunotherapy of cancer

Chemotherapy-conjugated immunotherapy in clinical oncology conceptually resembles the combined effects of cytoreduction and immunostimulation in membrane targeted cell killings mediated by pore-forming proteins or host defense peptides. Of the similar concept, targeting cancer cell membrane using membrane active peptides is a hopeful therapeutic modality but had long been hindered from in vivo application. Here we report an enabling strategy of pre-opsonizing a membrane penetrating Ir-complexed octa-arginine peptide (iPep) with serum albumin via intrinsic amphipathicity-driven bimodal interactions into nanoparticles (NP). We found that NP triggered stress-mediated 4T1 cell oncosis which induced potent immunological activation, surpassing several well-known immunogenic medicines. Vested with albumin-enhanced in vivo tumor targeting specificity and pharmacokinetic properties, NP showed combined chemo to immunotherapies of s. c. tumors in mice, with decreased percentages of MDSC, Treg, M2-like macrophage and improved infiltration of CTLs in tumor site, caused complete regression of 4T1 and CT26 tumors, outperforming clinical medicines. In a challenging orthotopic breast cancer model, boost i. v. injections of NP acted as in situ tumor vaccine that drastically enhanced 4T1-specific cellular and humoral immunities to reverse disease progression. Thus, with combined effects of direct cytoreduction, immune activation and tumor vaccine, iPep-NP presents the promise and potential of a new modality of cancer medicine.

Transcriptomic and proteomic analyses reveal the first occurrence of diverse toxin groups in Millepora alcicornis

Millepora alcicornis is a reef-forming cnidarian widely distributed in the Mexican Caribbean. Millepora species or “fire corals" inflict a painful stinging reaction in humans when touched. Even though hundreds of organic and polypeptide toxins have been characterized from sea anemones and jellyfish, there are few reports regarding the diversity of toxins synthesized by fire corals. Here, based on transcriptomic analysis of M. alcicornis, several predicted proteins that show amino acid sequence similarity to toxins were identified, including neurotoxins, metalloproteases, hemostasis-impairing toxins, serin proteases, cysteine-rich venom proteins, phospholipases, complement system-impairing toxins, phosphodiesterases, pore-forming toxins, and L-aminoacid oxidases. The soluble nematocyst proteome of this organism was shown to induce hemolytic, proteolytic, and phospholipase A2 effects by gel zymography. Protein bands or spots on 1D- and 2D-PAGE gels corresponding to zones of hemolytic and enzymatic activities were excised, subjected to in-gel digestion with trypsin, and analyzed by mass spectrometry. These proteins exhibited sequence homology to PLA2s, metalloproteinases, pore-forming toxins, and neurotoxins, such as actitoxins and CrTX-A. The complex array of venom-related transcripts that were identified in M. alcicornis, some of which are first reported in “fire corals”, provide novel insight into the structural richness of Cnidarian toxins and their distribution among species. SignificanceMarine organisms are a promising source of bioactive compounds with valuable contributions in diverse fields such as human health, pharmaceuticals, and industrial application. Currently, not much attention has been paid to the study of fire corals, which possess a variety of molecules that exhibit diverse toxic effects and therefore have great pharmaceutical and biotechnological potential. The isolation and identification of novel marine-derived toxins by classical approaches are time-consuming and have low yields. Thus, next-generation strategies, like base-’omics technologies, are essential for the high-throughput characterization of venom compounds such as those synthesized by fire corals.This study moves the field forward because it provides new insights regarding the first occurrence of diverse toxin groups in Millepora alcicornis. The findings presented here will contribute to the current understanding of the mechanisms of action of Millepora toxins. This research also reveals important information related to the potential role of toxins in the defense and capture of prey mechanisms and for designing appropriate treatments for fire coral envenomation. Moreover, due to the lack of information on the taxonomic identification of Millepora, the insights presented here can advise the taxonomic classification of the species of this genus.

Protein engineering of pores for separation, sensing, and sequencing.

Proteins are critical to cellular function and survival. They are complex molecules with precise structuresand chemistries, which allow them to serve diverse functions for maintaining overall cell homeostasis. Since the discovery of the first enzyme in 1833, a gamut of advanced experimental and computational tools has been developed and deployed for understanding protein structure and function. Recent studies have demonstrated the ability to redesign/alter natural proteins for applications in industrial processes of interest and to make customized, novel synthetic proteins in the laboratory through protein engineering. We comprehensively review the successes in engineering pore-forming proteins and correlate the amino acid-level biochemistry of different pore modification strategies to the intended applications limited to nucleotide/peptide sequencing, single-molecule sensing, and precise molecular separations.

Effects of sodium metabisulfite on pyroptosis, mitophagy and degranulation in mast cells.

Sodium metabisulphite (SMB) is the most used foods and drugs antioxidant among sulfites. So far, there were few studies about its harm, especially in mast cells. Our study was to investigate the effects of SMB on mitophagy and pyroptosis in mast cells. The results revealed that SMB dose-dependently promoted the expressions of NLRP3, GSDMD-N and other marker proteins of pyroptosis. Knockdown of GSDMD, NLRP3 inhibitor, mitophagy activator and mtROS inhibitor all reversed the changes in pyroptosis indicators caused by SMB. Considering the degranulation characteristics of mast cells and the sensitization of sulfite, we examined the effects of the above inhibitors on the degranulation of mast cells caused by SMB. The results showed that SMB-mediated mast cell degranulation was significantly inhibited by the above inhibitors. Meanwhile, we used immunofluorescence co-localization experiments and found that GSDMD pore-forming protein and histamine co-localized near the cell membrane. Overall, evidence suggested that SMB caused pyroptosis by inhibiting mitophagy, leading to mast cell degranulation. These findings are of great significance to the sensitization mechanism of SMB and provide a new insight into SMB toxicology and mast cell degranulation.

Gating of β-Barrel Protein Pores, Porins, and Channels: An Old Problem with New Facets.

β barrels are ubiquitous proteins in the outer membranes of mitochondria, chloroplasts, and Gram-negative bacteria. These transmembrane proteins (TMPs) execute a wide variety of tasks. For example, they can serve as transporters, receptors, membrane-bound enzymes, as well as adhesion, structural, and signaling elements. In addition, multimeric β barrels are common structural scaffolds among many pore-forming toxins. Significant progress has been made in understanding the functional, structural, biochemical, and biophysical features of these robust and versatile proteins. One frequently encountered fundamental trait of all β barrels is their voltage-dependent gating. This process consists of reversible or permanent conformational transitions between a large-conductance, highly permeable open state and a low-conductance, solute-restrictive closed state. Several intrinsic molecular mechanisms and environmental factors modulate this universal property of β barrels. This review article outlines the typical signatures of voltage-dependent gating. Moreover, we discuss recent developments leading to a better qualitative understanding of the closure dynamics of these TMPs.

Fast drosophila enterocyte regrowth after infection involves a reverse metabolic flux driven by an amino acid transporter

Upon exposure to a bacterial pore-forming toxin, enterocytes rapidly purge their apical cytoplasm into the gut lumen, resulting in a thin intestinal epithelium. The enterocytes regain their original shape and thickness within 16h after the ingestion of the bacteria. Here, we show that the regrowth of Drosophila enterocytes entails an inversion of metabolic fluxes from the organism back toward the intestine. We identify a proton-assisted transporter, Arcus, that is required for the reverse absorption of amino acids and the timely recovery of the intestinal epithelium. Arcus is required for a peak of amino acids appearing in the hemolymph shortly after infection. The regrowth of enterocytes involves the insulin signaling pathway and Myc. The purge decreases Myc mRNA levels, which subsequently remain at low levels in the arcus mutant. Interestingly, the action of arcus and Myc in the intestinal epithelium is not cell-autonomous, suggesting amino acid fluxes within the intestinal epithelium.

Surface plasmon resonance and microscale thermophoresis approaches for determining the affinity of perforin for calcium ions.

Perforin is a pore-forming protein that plays a crucial role in the immune system by clearing virus-infected or tumor cells. It is released from cytotoxic granules of immune cells and forms pores in targeted lipid membranes to deliver apoptosis-inducing granzymes. It is a very cytotoxic protein and is therefore adapted not to act in producing cells. Its activity is regulated by the requirement for calcium ions for optimal activity. However, the exact affinity of perforin for calcium ions has not yet been determined. We conducted a molecular dynamics simulation in the absence or presence of calcium ions that showed that binding of at least three calcium ions is required for stable perforin binding to the lipid membrane. Biophysical studies using surface plasmon resonance and microscale thermophoresis were then performed to estimate the binding affinities of native human and recombinant mouse perforin for calcium ions. Both approaches showed that mouse perforin has a several fold higher affinity for calcium ions than that of human perforin. This was attributed to a particular residue, tryptophan at position 488 in mouse perforin, which is replaced by arginine in human perforin. This represents an additional mechanism to control the activity of human perforin.

Activation of inflammasomes and mechanisms for intracellular recognition of Listeria monocytogenes.

The high mortality rate associated with Listeria monocytogenes can be attributed to its ability to invade the body systemically and to activate inflammasomes. Both of these processes are facilitated by expressing a major virulence factor known as listeriolysin O, a 56 kDa pore-forming protein encoded by the hly gene. Listeriolysin O plays a crucial role in the pathogenesis of the bacterium by facilitating the escape of the pathogen from the phagosome into the cytosol. This process is essential for the successful establishment of infection. In addition, listeriolysin O is known as an immunomodulator that activates host signal transduction. In addition to listeriolysin O, Listeria expresses a variety of bacterial ligands, such as lipoteichoic acid, nucleotide, and flagellin, that are recognized by host intracellular pattern-recognition receptors including Nod-like receptors, AIM2-like receptors, and RIG-I-like receptors. This review introduces intracellular recognition of Listeria monocytogenes since recent studies have revealed that the activation of inflammasome exacerbates Gram-positive bacteria infection.

Bacterial Pesticidal Protein Mpp51Aa1 Delivered via Transgenic Citrus Severely Impacts the Fecundity of Asian Citrus Psyllid, Diaphorina citri.

The Asian citrus psyllid (ACP) Diaphorina citri vectors the causative agent of citrus greening disease that has the capacity to decimate citrus production. As an alternative and more sustainable approach to manage D. citri than repeated application of chemical insecticides, we investigated the potential use of the bacteria-derived pesticidal protein, Mpp51Aa1, when delivered by transgenic Citrus sinensis cv. Valencia sweet orange or Citrus paradisi cv. Duncan grapefruit. Following confirmation of transcription and translation of mpp51aa1 by transgenic plants, no impact of Mpp51Aa1 expression was seen on D. citri host plant choice between transgenic and control Duncan grapefruit plants. A slight but significant drop in survival of adult psyllids fed on these transgenic plants was noted relative to those fed on control plants. In line with this result, damage to the gut epithelium consistent with that caused by pore-forming proteins was only observed in a minority of adult D. citri fed on the transgenic Duncan grapefruit. However, greater impacts were observed on nymphs than on adults, with a 40% drop in the survival of nymphs fed on transgenic Duncan grapefruit relative to those fed on control plants. For Valencia sweet orange, a 70% decrease in the number of eggs laid by adult D. citri on transgenic plants was noted relative to those on control plants, with a 90% drop in emergence of progeny. These impacts that contrast with those associated with other bacterial pesticidal proteins and the potential for use of Mpp51Aa1-expressing transgenic plants for suppression of D. citri populations are discussed. IMPORTANCE Pesticidal proteins derived from bacteria such as Bacillus thuringiensis are valuable tools for management of agricultural insect pests and provide a sustainable alternative to the application of chemical insecticides. However, relatively few bacterial pesticidal proteins have been used for suppression of hemipteran or sap-sucking insects such as the Asian citrus psyllid, Diaphorina citri. This insect is particularly important as the vector of the causative agent of citrus greening, or huanglongbing disease, which severely impacts global citrus production. In this study, we investigated the potential of transgenic citrus plants that produce the pesticidal protein Mpp51Aa1. While adult psyllid mortality on transgenic plants was modest, the reduced number of eggs laid by exposed adults and the decreased survival of progeny was such that psyllid populations dropped by more than 90%. These results provide valuable insight for potential deployment of Mpp51Aa1 in combination with other control agents for the management of D. citri.

Aminopeptidase MNP-1 triggers intestine protease production by activating daf-16 nuclear location to degrade pore-forming toxins in Caenorhabditis elegans.

Pore-forming toxins (PFTs) are effective tools for pathogens infection. By disrupting epithelial barriers and killing immune cells, PFTs promotes the colonization and reproduction of pathogenic microorganisms in their host. In turn, the host triggers defense responses, such as endocytosis, exocytosis, or autophagy. Bacillus thuringiensis (Bt) bacteria produce PFT, known as crystal proteins (Cry) which damage the intestinal cells of insects or nematodes, eventually killing them. In insects, aminopeptidase N (APN) has been shown to act as an important receptor for Cry toxins. Here, using the nematode Caenorhabditis elegans as model, an extensive screening of APN gene family was performed to analyze the potential role of these proteins in the mode of action of Cry5Ba against the nematode. We found that one APN, MNP-1, participate in the toxin defense response, since the mnp-1(ok2434) mutant showed a Cry5Ba hypersensitive phenotype. Gene expression analysis in mnp-1(ok2434) mutant revealed the involvement of two protease genes, F19C6.4 and R03G8.6, that participate in Cry5Ba degradation. Finally, analysis of the transduction pathway involved in F19C6.4 and R03G8.6 expression revealed that upon Cry5Ba exposure, the worms up regulated both protease genes through the activation of the FOXO transcription factor DAF-16, which was translocated into the nucleus. The nuclear location of DAF-16 was found to be dependent on mnp-1 under Cry5Ba treatment. Our work provides evidence of new host responses against PFTs produced by an enteric pathogenic bacterium, resulting in activation of host intestinal proteases that degrade the PFT in the intestine.

Characterization of TelE, a T7SS LXG Effector Exhibiting a Conserved C-Terminal Glycine Zipper Motif Required for Toxicity.

Streptococcus gallolyticus subsp. gallolyticus (SGG) is an opportunistic bacterial pathogen strongly associated with colorectal cancer. Here, through comparative genomics analysis, we demonstrated that the genetic locus encoding the type VIIb secretion system (T7SSb) machinery is uniquely present in SGG in two different arrangements. SGG UCN34 carrying the most prevalent T7SSb genetic arrangement was chosen as the reference strain. To identify the effectors secreted by this secretion system, we inactivated the essC gene encoding the motor of this machinery. A comparison of the proteins secreted by UCN34 wild type and its isogenic ΔessC mutant revealed six T7SSb effector proteins, including the expected WXG effector EsxA and three LXG-containing proteins. In this work, we characterized an LXG-family toxin named herein TelE promoting the loss of membrane integrity. Seven homologs of TelE harboring a conserved glycine zipper motif at the C terminus were identified in different SGG isolates. Scanning mutagenesis of this motif showed that the glycine residue at position 470 was crucial for TelE membrane destabilization activity. TelE activity was antagonized by a small protein TipE belonging to the DUF5085 family. Overall, we report herein a unique SGG T7SSb effector exhibiting a toxic activity against nonimmune bacteria. IMPORTANCE In this study, 38 clinical isolates of Streptococcus gallolyticus subsp. gallolyticus (SGG) were sequenced and a genetic locus encoding the type VIIb secretion system (T7SSb) was found conserved and absent from 16 genomes of the closely related S. gallolyticus subsp. pasteurianus (SGP). The T7SSb is a bona fide pathogenicity island. Here, we report that the model organism SGG strain UCN34 secretes six T7SSb effectors. One of the six effectors named TelE displayed a strong toxicity when overexpressed in Escherichia coli. Our results indicate that TelE is probably a pore-forming toxin whose activity can be antagonized by a specific immunity protein named TipE. Overall, we report a unique toxin-immunity protein pair and our data expand the range of effectors secreted through T7SSb.

Horizontal gene transfer underlies the painful stings of asp caterpillars (Lepidoptera: Megalopygidae)

Larvae of the genus Megalopyge (Lepidoptera: Zygaenoidea: Megalopygidae), known as asp or puss caterpillars, produce defensive venoms that cause severe pain. Here, we present the anatomy, chemistry, and mode of action of the venom systems of caterpillars of two megalopygid species, the Southern flannel moth Megalopyge opercularis and the black-waved flannel moth Megalopyge crispata. We show that megalopygid venom is produced in secretory cells that lie beneath the cuticle and are connected to the venom spines by canals. Megalopygid venoms consist of large aerolysin-like pore-forming toxins, which we have named megalysins, and a small number of peptides. The venom system differs markedly from those of previously studied venomous zygaenoids of the family Limacodidae, suggestive of an independent origin. Megalopygid venom potently activates mammalian sensory neurons via membrane permeabilization and induces sustained spontaneous pain behavior and paw swelling in mice. These bioactivities are ablated by treatment with heat, organic solvents, or proteases, indicating that they are mediated by larger proteins such as the megalysins. We show that the megalysins were recruited as venom toxins in the Megalopygidae following horizontal transfer of genes from bacteria to the ancestors of ditrysian Lepidoptera. Megalopygids have recruited aerolysin-like proteins as venom toxins convergently with centipedes, cnidarians, and fish. This study highlights the role of horizontal gene transfer in venom evolution.

Functional Diversity of Novel Lectins with Unique Structural Features in Marine Animals.

Due to their remarkable structural diversity, glycans play important roles as recognition molecules on cell surfaces of living organisms. Carbohydrates exist in numerous isomeric forms and can adopt diverse structures through various branching patterns. Despite their relatively small molecular weights, they exhibit extensive structural diversity. On the other hand, lectins, also known as carbohydrate-binding proteins, not only recognize and bind to the diverse structures of glycans but also induce various biological reactions based on structural differences. Initially discovered as hemagglutinins in plant seeds, lectins have been found to play significant roles in cell recognition processes in higher vertebrates. However, our understanding of lectins in marine animals, particularly marine invertebrates, remains limited. Recent studies have revealed that marine animals possess novel lectins with unique structures and glycan recognition mechanisms not observed in known lectins. Of particular interest is their role as pattern recognition molecules in the innate immune system, where they recognize the glycan structures of pathogens. Furthermore, lectins serve as toxins for self-defense against foreign enemies. Recent discoveries have identified various pore-forming proteins containing lectin domains in fish venoms and skins. These proteins utilize lectin domains to bind target cells, triggering oligomerization and pore formation in the cell membrane. These findings have spurred research into the new functions of lectins and lectin domains. In this review, we present recent findings on the diverse structures and functions of lectins in marine animals.

The Global Burden of Community-Acquired Pneumonia in Adults, Encompassing Invasive Pneumococcal Disease and the Prevalence of Its Associated Cardiovascular Events, with a Focus on Pneumolysin and Macrolide Antibiotics in Pathogenesis and Therapy

Despite innovative advances in anti-infective therapies and vaccine development technologies, community-acquired pneumonia (CAP) remains the most persistent cause of infection-related mortality globally. Confronting the ongoing threat posed by Streptococcus pneumoniae (the pneumococcus), the most common bacterial cause of CAP, particularly to the non-immune elderly, remains challenging due to the propensity of the elderly to develop invasive pneumococcal disease (IPD), together with the predilection of the pathogen for the heart. The resultant development of often fatal cardiovascular events (CVEs), particularly during the first seven days of acute infection, is now recognized as a relatively common complication of IPD. The current review represents an update on the prevalence and types of CVEs associated with acute bacterial CAP, particularly IPD. In addition, it is focused on recent insights into the involvement of the pneumococcal pore-forming toxin, pneumolysin (Ply), in subverting host immune defenses, particularly the protective functions of the alveolar macrophage during early-stage disease. This, in turn, enables extra-pulmonary dissemination of the pathogen, leading to cardiac invasion, cardiotoxicity and myocardial dysfunction. The review concludes with an overview of the current status of macrolide antibiotics in the treatment of bacterial CAP in general, as well as severe pneumococcal CAP, including a consideration of the mechanisms by which these agents inhibit the production of Ply by macrolide-resistant strains of the pathogen.