Mucopolysaccharidosis-Plus Syndrome: Is This a Type of Mucopolysaccharidosis or a Separate Kind of Metabolic Disease?

Several years ago, dozens of cases were described in patients with symptoms very similar to mucopolysaccharidosis (MPS). This new disease entity was described as mucopolysaccharidosis-plus syndrome (MPSPS). The name of the disease indicates that in addition to the typical symptoms of conventional MPS, patients develop other features such as congenital heart defects and kidney and hematopoietic system disorders. The symptoms are highly advanced, and patients usually do not survive past the second year of life. MPSPS is inherited in an autosomal recessive manner and is caused by a homozygous-specific mutation in the gene encoding the VPS33A protein. To date, it has been described in 41 patients. Patients with MPSPS exhibited excessive excretion of glycosaminoglycans (GAGs) in the urine and exceptionally high levels of heparan sulfate in the plasma, but the accumulation of substrates is not caused by a decrease in the activity of any lysosomal enzymes. Here, we discuss the pathomechanisms and symptoms of MPSPS, comparing them to those of MPS. Moreover, we asked the question whether MPSPS should be classified as a type of MPS or a separate disease, as contrary to 'classical' MPS types, despite GAG accumulation, no defects in lysosomal enzymes responsible for degradation of these compounds could be detected in MPSPS. The molecular mechanism of the appearance of GAG accumulation in MPSPS is suggested on the basis of results available in the literature.

Characterization and functional analysis of Toxoplasma Golgi-associated proteins identified by proximity labeling.

Toxoplasma gondii possesses a highly polarized secretory pathway that contains both broadly conserved eukaryotic organelles and unique apicomplexan organelles, which play essential roles in the parasite's lytic cycle. As in other eukaryotes, the T. gondii Golgi apparatus sorts and modifies proteins prior to their distribution to downstream organelles. Many of the typical trafficking factors found involved in these processes are missing from apicomplexan genomes, suggesting that these parasites have evolved unique proteins to fill these roles. Here, we identify a Golgi-localizing protein (ULP1), which is structurally similar to the eukaryotic trafficking factor p115/Uso1. We demonstrate that depletion of ULP1 leads to a dramatic reduction in parasite fitness that is the result of defects in microneme secretion, invasion, replication, and egress. Using ULP1 as bait for TurboID proximity labeling and immunoprecipitation, we identify 11 more Golgi-associated proteins and demonstrate that ULP1 interacts with the T. gondii-conserved oligomeric Golgi (COG) complex. These proteins include both conserved trafficking factors and parasite-specific proteins. Using a conditional knockdown approach, we assess the effect of each of these 11 proteins on parasite fitness. Together, this work reveals a diverse set of T. gondii Golgi-associated proteins that play distinct roles in the secretory pathway. As several of these proteins are absent outside of the Apicomplexa, they represent potential targets for the development of novel therapeutics against these parasites. IMPORTANCE: Apicomplexan parasites such as Toxoplasma gondii infect a large percentage of the world's population and cause substantial human disease. These widespread pathogens use specialized secretory organelles to infect their host cells, modulate host cell functions, and cause disease. While the functions of the secretory organelles are now better understood, the Golgi apparatus of the parasite remains largely unexplored, particularly regarding parasite-specific innovations that may help direct traffic intracellularly. In this work, we characterize ULP1, a protein that is unique to parasites but shares structural similarity to the eukaryotic trafficking factor p115/Uso1. We show that ULP1 plays an important role in parasite fitness and demonstrate that it interacts with the conserved oligomeric Golgi (COG) complex. We then use ULP1 proximity labeling to identify 11 additional Golgi-associated proteins, which we functionally analyze via conditional knockdown. This work expands our knowledge of the Toxoplasma Golgi apparatus and identifies potential targets for therapeutic intervention.

The Chlamydia effector IncE employs two short linear motifs to reprogram host vesicle trafficking.

Chlamydia trachomatis, a leading cause of bacterial sexually transmitted infections, creates a specialized intracellular replicative niche by translocation and insertion of a diverse array of effectors (Incs [inclusion membrane proteins]) into the inclusion membrane. Here, we characterize IncE, a multifunctional Inc that encodes two non-overlapping short linear motifs (SLiMs) within its short cytosolic C terminus. The proximal SLiM, by mimicking just a small portion of an R-N-ethylmaleimide-sensitive factor adaptor protein receptor (SNARE) motif, binds and recruits syntaxin (STX)7- and STX12-containing vesicles to the inclusion. The distal SLiM mimics the sorting nexin (SNX)5 and SNX6 cargo binding site to recruit SNX6-containing vesicles to the inclusion. By simultaneously binding two distinct vesicle classes, IncE brings these vesicles in close apposition with each other at the inclusion to facilitate C. trachomatis intracellular development. Our work suggests that Incs may have evolved SLiMs to enable rapid evolution in a limited protein space to disrupt host cell processes.

Invasion of the stigma by oomycete pathogenic hyphae or pollen tubes : striking similarities and differences.

Both filamentous pathogens' hyphae and pollen tube penetrate the host's outer layer and involve growth within the host tissues. Early epidermal responses are decisive for the outcome of these two-cell interaction processes. We identified a single cell type, the papilla of Arabidospis thaliana's stigma, as a tool to conduct a comprehensive comparative analysis on how an epidermal cell responds to the invasion of an unwanted pathogen or a welcomed pollen tube. We showed that Phytophtora parasitica, a root oomycete, effectively breaches the stigmatic cell wall and develops as a biotroph within the papilla cytoplasm. These invasive features resemble the behaviour exhibited by the pathogen within its natural host cells, but diverge from the manner in which the pollen tube progresses, being engulfed within the papilla cell wall. Quantitative analysis revealed that both invaders trigger reorganisation of the stigmatic endomembrane system and the actin cytoskeleton. While some remodelling processes are shared between the two interactions, others appear more specific towards the respective invader. These findings underscore the remarkable ability of an epidermal cell to differentiate between two types of invaders, thereby enabling it to trigger the most suitable response during the onset of invasion.

Tight Junction Component Occludin Binds to FIP5 to Regulate Endosome Trafficking and Mitotic Spindle Function.

The genetic basis of vertebrate emergence during metazoan evolution has remained largely unknown. Understanding vertebrate-specific genes, such as the tight junction protein Occludin (Ocln), may help answer this question. Here, it is shown that mammary glands lacking Ocln exhibit retarded epithelial branching, owing to reduced cell proliferation and surface expansion. Interestingly, Ocln regulates mitotic spindle orientation and function, and its loss leads to a range of defects, including prolonged prophase and failed nuclear and/or cytoplasmic division. Mechanistically, Ocln binds to the RabGTPase-11 adaptor FIP5 and recruits recycling endosomes to the centrosome to participate in spindle assembly and function. FIP5 loss recapitulates Ocln null, leading to prolonged prophase, reduced cell proliferation, and retarded epithelial branching. These results identify a novel role in OCLN-mediated endosomal trafficking and potentially highlight its involvement in mediating membranous vesicle trafficking and function, which is evolutionarily conserved and essential.

Genome-Wide Identification of SNARE Family Genes and Functional Characterization of an R-SNARE Gene BbSEC22 in a Fungal Insect Pathogen Beauveria bassiana.

Soluble N-ethylmaleimide-sensitive factor attachment protein receptors (SNAREs) are central components of the machinery mediating cell membrane fusion and intracellular vesicular trafficking in eukaryotic cells, and have been well-documented to play critical roles in growth, development, and pathogenesis in the filamentous fungal plant pathogens. However, little is known about the contributions of SNAREs to the physiology and biocontrol potential in entomopathogenic filamentous fungi. Here, a genome-wide analysis of SNARE genes was performed taking advantage of the available whole genome sequence of Beauveria bassiana, a classical entomopathogenic fungus. Based on the compared genomic method, 22 genes encoding putative SNAREs were identified from the whole genome of B. bassiana, and were classified into four groups (7 Qa-, 4 Qb-, 6 Qc-, and 5 R-SNAREs) according to the conserved structural features of their encoding proteins. An R-SNARE encoding gene BbSEC22 was further functionally characterized by gene disruption and complementation. The BbSEC22 null mutant showed a fluffy appearance in mycelial growth and an obvious lag in conidial germination. The null mutant also exhibited significantly increased sensitivity to oxidative stress and cell wall perturbing agents and reduced the yield of conidia production by 43.1% compared with the wild-type strain. Moreover, disruption of BbSEC22 caused a significant decrease in conidial virulence to Spodoptera litura larvae. Overall, our results provide an overview of vesicle trafficking in B. bassiana and revealed that BbSec22 was a multifunctional protein associated with mycelial growth, sporulation, conidial germination, stress tolerance, and insecticidal virulence.

Exploring the Implications of Golgi Apparatus Dysfunction in Bone Diseases.

The Golgi apparatus is an organelle responsible for protein processing, sorting, and transport in cells. Recent research has shed light on its possible role in the pathogenesis of various bone diseases. This review seeks to explore its significance in osteoporosis, osteogenesis imperfecta, and other bone conditions such as dysplasias. Numerous lines of evidence demonstrate that perturbations to Golgi apparatus function can disrupt post-translational protein modification, folding and trafficking functions crucial for bone formation, mineralization, and remodeling. Abnormalities related to glycosylation, protein sorting, or vesicular transport in Golgi have been associated with altered osteoblast and osteoclast function, compromised extracellular matrix composition, as well as disrupted signaling pathways involved with homeostasis of bones. Mutations or dysregulation of Golgi-associated proteins, including golgins and coat protein complex I and coat protein complex II coat components, have also been implicated in bone diseases. Such genetic alterations may disrupt Golgi structure, membrane dynamics, and protein transport, leading to bone phenotype abnormalities. Understanding the links between Golgi apparatus dysfunction and bone diseases could provide novel insights into disease pathogenesis and potential therapeutic targets. Future research should focus on unraveling specific molecular mechanisms underlying Golgi dysfunction associated with bone diseases to develop targeted interventions for restoring normal bone homeostasis while decreasing clinical manifestations associated with these issues.

Identification, Classification, and Transcriptional Analysis of Rab GTPase Genes from Tomato (Solanum lycopersicum) Reveals Salt Stress Response Genes.

Salinity in plants generates an osmotic and ionic imbalance inside cells that compromises the viability of the plant. Rab GTPases, the largest family within the small GTPase superfamily, play pivotal roles as regulators of vesicular trafficking in plants, including the economically important and globally cultivated tomato (Solanum lycopersicum). Despite their significance, the specific involvement of these small GTPases in tomato vesicular trafficking and their role under saline stress remains poorly understood. In this work, we identified and classified 54 genes encoding Rab GTPases in cultivated tomato, elucidating their genomic distribution and structural characteristics. We conducted an analysis of duplication events within the S. lycopersicum genome, as well as an examination of gene structure and conserved motifs. In addition, we investigated the transcriptional profiles for these Rab GTPases in various tissues of cultivated and wild tomato species using microarray-based analysis. The results showed predominantly low expression in most of the genes in both leaves and vegetative meristem, contrasting with notably high expression levels observed in seedling roots. Also, a greater increase in gene expression in shoots from salt-tolerant wild tomato species was observed under normal conditions when comparing Solanum habrochaites, Solanum pennellii, and Solanum pimpinellifolium with S. lycopersicum. Furthermore, an expression analysis of Rab GTPases from Solanum chilense in leaves and roots under salt stress treatment were also carried out for their characterization. These findings revealed that specific Rab GTPases from the endocytic pathway and the trans-Golgi network (TGN) showed higher induction in plants exposed to saline stress conditions. Likewise, disparities in gene expression were observed both among members of the same Rab GTPase subfamily and between different subfamilies. Overall, this work emphasizes the high degree of conservation of Rab GTPases, their high functional diversification in higher plants, and the essential role in mediating salt stress tolerance and suggests their potential for further exploration of vesicular trafficking mechanisms in response to abiotic stress conditions.

Toxoplasma and Plasmodium associate with host Arfs during infection.

The apicomplexans Toxoplasma gondii and Plasmodium are intracellular parasites that reside within a host-derived compartment termed the parasitophorous vacuole (PV). During infection, the parasites must acquire critical host resources and transport them across their PV for development. However, the mechanism by which host resources are trafficked to and across the PV remains uncertain. Here, we investigated host ADP ribosylation factors (Arfs), a class of proteins involved in vesicular trafficking that may be exploited by T. gondii and Plasmodium berghei for nutrient acquisition. Using overexpressed Arf proteins coupled with immunofluorescence microscopy, we found that all Arfs were internalized into the T. gondii PV, with most vacuoles containing at least one punctum of Arf protein by the end of the lytic cycle. We further characterized Arf1, the most abundant Arf inside the T. gondii PV, and observed that active recycling between its GDP/GTP-bound state influenced Arf1 internalization independent of host guanine nucleotide exchange factors (GEFs). In addition, Arf1 colocalized with vesicle coat complexes and exogenous sphingolipids, suggesting a role in nutrient acquisition. While Arf1 and Arf4 were not observed inside the PV during P. berghei infection, our gene depletion studies showed that liver stage development and survival depended on the expression of Arf4 and the host GEF, GBF1. Collectively, these observations indicate that apicomplexans use distinct mechanisms to subvert the host vesicular trafficking network and efficiently replicate. The findings also pave the way for future studies to identify parasite proteins critical to host vesicle recruitment and the components of vesicle cargo. IMPORTANCE: The parasites Toxoplasma gondii and Plasmodium live complex intracellular lifestyles where they must acquire essential host nutrients while avoiding recognition. Although previous work has sought to identify the specific nutrients scavenged by apicomplexans, the mechanisms by which host materials are transported to and across the parasite vacuole membrane are largely unknown. Here, we examined members of the host vesicular trafficking network to identify specific pathways subverted by T. gondii and Plasmodium berghei. Our results indicate that T. gondii selectively internalizes host Arfs, a class of proteins involved in intracellular trafficking. For P. berghei, host Arfs were restricted by the parasite's vacuole membrane, but proteins involved in vesicular trafficking were identified as essential for liver stage development. A greater exploration into how and why apicomplexans subvert host vesicular trafficking could help identify targets for host-directed therapeutics.

Disorders of vesicular trafficking presenting with recurrent acute liver failure: NBAS, RINT1, and SCYL1 deficiency.

Among genetic disorders of vesicular trafficking, there are three causing recurrent acute liver failure (RALF): NBAS, RINT1, and SCYL1-associated disease. These three disorders are characterized by liver crises triggered by febrile infections and account for a relevant proportion of RALF causes. While the frequency and severity of liver crises in NBAS and RINT1-associated disease decrease with age, patients with SCYL1 variants present with a progressive, cholestatic course. In all three diseases, there is a multisystemic, partially overlapping phenotype with variable expression, including liver, skeletal, and nervous systems, all organ systems with high secretory activity. There are no specific biomarkers for these diseases, and whole exome sequencing should be performed in patients with RALF of unknown etiology. NBAS, SCYL1, and RINT1 are involved in antegrade and retrograde vesicular trafficking. Pathomechanisms remain unclarified, but there is evidence of a decrease in concentration and stability of the protein primarily affected by the respective gene defect and its interaction partners, potentially causing impairment of vesicular transport. The impairment of protein secretion by compromised antegrade transport provides a possible explanation for different organ manifestations such as bone alteration due to lack of collagens or diabetes mellitus when insulin secretion is affected. Dysfunction of retrograde transport impairs membrane recycling and autophagy. The impairment of vesicular trafficking results in increased endoplasmic reticulum stress, which, in hepatocytes, can progress to hepatocytolysis. While there is no curative therapy, an early and consequent implementation of an emergency protocol seems crucial for optimal therapeutic management.

Phosphorylation-state dependent intraneuronal sorting of Aß differentially impairs autophagy and the endo-lysosomal system.

ABBREVIATIONS: AD: Alzheimer disease; APP: amyloid beta precursor protein; ATG: autophagy related; Aß: amyloid-ß; CTSD: cathepsin D; DAPI: 4',6-diamidino-2-phenylindole; EEA1: early endosome antigen 1; FA: formic acid; GFP: green fluorescent protein; LAMP2: lysosomal-associated membrane protein 2; MAP1LC3/LC3: microtubule-associated protein 1 light chain 3; MAP2: microtubule-associated protein 2; nmAß: non-modified amyloid-ß; npAß: non-phosphorylated amyloid-ß; pAß: phosphorylated amyloid-ß; p-Ser26Aß: amyloid-ß phosphorylated at serine residue 26; p-Ser8Aß: amyloid-ß phosphorylated at serine residue 8; RAB: RAB, member RAS oncogene family; RFP: red fluorescent protein; SQSTM1/p62: sequestome 1; YFP: yellow fluorescent protein.

Spontaneous transfer of small peripheral peptides between supported lipid bilayer and giant unilamellar vesicles.

Vesicular trafficking facilitates material transport between membrane-bound organelles. Membrane protein cargos are trafficked for relocation, recycling, and degradation during various physiological processes. In vitro fusion studies utilized synthetic lipid membranes to study the molecular mechanisms of vesicular trafficking and to develop synthetic materials mimicking the biological membrane trafficking. Various fusogenic conditions which can induce vesicular fusion have been used to establish synthetic systems that can mimic biological systems. Despite these efforts, the mechanisms underlying vesicular trafficking of membrane proteins remain limited and robust in vitro methods that can construct synthetic trafficking systems for membrane proteins between large membranes (>1 µm2) are unavailable. Here, we provide data to show the spontaneous transfer of small membrane-bound peptides (∼4 kD) between a supported lipid bilayer (SLB) and giant unilamellar vesicles (GUVs). We found that the contact between the SLB and GUVs led to the occasional but notable transfer of membrane-bound peptides in a physiological saline buffer condition (pH 7.4, 150 mM NaCl). Quantitative and dynamic time-lapse analyses suggested that the observed exchange occurred through the formation of hemi-fusion stalks between the SLB and GUVs. Larger protein cargos with a size of ∼77 kD could not be transferred between the SLB and GUVs, suggesting that the larger-sized cargos limited diffusion across the hemi-fusion stalk, which was predicted to have a highly curved structure. Compositional study showed Ni-chelated lipid head group was the essential component catalyzing the process. Our system serves as an example synthetic platform that enables the investigation of small-peptide trafficking between synthetic membranes and reveals hemi-fused lipid bridge formation as a mechanism of peptide transfer.

Mechanisms of aquaporin-4 vesicular trafficking in mammalian cells.

The aquaporin-4 (AQP4) water channel is abundantly expressed in the glial cells of the central nervous system and facilitates brain swelling following diverse insults, such as traumatic injury or stroke. Lack of specific and therapeutic AQP4 inhibitors highlights the need to explore alternative routes to control the water permeability of glial cell membranes. The cell surface abundance of AQP4 in mammalian cells fluctuates rapidly in response to changes in oxygen levels and tonicity, suggesting a role for vesicular trafficking in its translocation to and from the cell surface. However, the molecular mechanisms of AQP4 trafficking are not fully elucidated. In this work, early and recycling endosomes were investigated as likely candidates of rapid AQP4 translocation together with changes in cytoskeletal dynamics. In transiently transfected HEK293 cells a significant amount of AQP-eGFP colocalised with mCherry-Rab5-positive early endosomes and mCherry-Rab11-positive recycling endosomes. When exposed to hypotonic conditions, AQP4-eGFP rapidly translocated from intracellular vesicles to the cell surface. Co-expression of dominant negative forms of the mCherry-Rab5 and -Rab11 with AQP4-eGFP prevented hypotonicity-induced AQP4-eGFP trafficking and led to concentration at the cell surface or intracellular vesicles respectively. Use of endocytosis inhibiting drugs indicated that AQP4 internalisation was dynamin-dependent. Cytoskeleton dynamics-modifying drugs also affected AQP4 translocation to and from the cell surface. AQP4 trafficking mechanisms were validated in primary human astrocytes, which express high levels of endogenous AQP4. The results highlight the role of early and recycling endosomes and cytoskeletal dynamics in AQP4 translocation in response to hypotonic and hypoxic stress and suggest continuous cycling of AQP4 between intracellular vesicles and the cell surface under physiological conditions.

Vesicular Trafficking, a Mechanism Controlled by Cascade Activation of Rab Proteins: Focus on Rab27.

Vesicular trafficking is essential for the cell to internalize useful proteins and soluble substances, for cell signaling or for the degradation of pathogenic elements such as bacteria or viruses. This vesicular trafficking also enables the cell to engage in secretory processes for the elimination of waste products or for the emission of intercellular communication vectors such as cytokines, chemokines and extracellular vesicles. Ras-related proteins (Rab) and their effector(s) are of crucial importance in all of these processes, and mutations/alterations to them have serious pathophysiological consequences. This review presents a non-exhaustive overview of the role of the major Rab involved in vesicular trafficking, with particular emphasis on their involvement in the biogenesis and secretion of extracellular vesicles, and on the role of Rab27 in various pathophysiological processes. Therefore, Rab and their effector(s) are central therapeutic targets, given their involvement in vesicular trafficking and their importance for cell physiology.

Analysis of CDPK1 targets identifies a trafficking adaptor complex that regulates microneme exocytosis in Toxoplasma.

Apicomplexan parasites use Ca2+-regulated exocytosis to secrete essential virulence factors from specialized organelles called micronemes. Ca2+-dependent protein kinases (CDPKs) are required for microneme exocytosis; however, the molecular events that regulate trafficking and fusion of micronemes with the plasma membrane remain unresolved. Here, we combine sub-minute resolution phosphoproteomics and bio-orthogonal labeling of kinase substrates in Toxoplasma gondii to identify 163 proteins phosphorylated in a CDPK1-dependent manner. In addition to known regulators of secretion, we identify uncharacterized targets with predicted functions across signaling, gene expression, trafficking, metabolism, and ion homeostasis. One of the CDPK1 targets is a putative HOOK activating adaptor. In other eukaryotes, HOOK homologs form the FHF complex with FTS and FHIP to activate dynein-mediated trafficking of endosomes along microtubules. We show the FHF complex is partially conserved in T. gondii, consisting of HOOK, an FTS homolog, and two parasite-specific proteins (TGGT1_306920 and TGGT1_316650). CDPK1 kinase activity and HOOK are required for the rapid apical trafficking of micronemes as parasites initiate motility. Moreover, parasites lacking HOOK or FTS display impaired microneme protein secretion, leading to a block in the invasion of host cells. Taken together, our work provides a comprehensive catalog of CDPK1 targets and reveals how vesicular trafficking has been tuned to support a parasitic lifestyle.

Synergism of vesicle trafficking and cytoskeleton during regulation of plant growth and development: A mechanistic outlook.

The cytoskeleton is a fundamental component found in all eukaryotic organisms, serving as a critical factor in various essential cyto-biological mechanisms, particularly in the locomotion and morphological transformations of plant cells. The cytoskeleton is comprised of three main components: microtubules (MT), microfilaments (MF), and intermediate filaments (IF). The cytoskeleton plays a crucial role in the process of cell wall formation and remodeling throughout the growth and development of cells. It is a highly organized and regulated network composed of filamentous components. In the basic processes of intracellular transport, such as mitosis, cytokinesis, and cell polarity, the plant cytoskeleton plays a crucial role according to recent studies. The major flaws in the organization of the cytoskeletal framework are at the root of the aberrant organogenesis currently observed in plant mutants. The regulation of protein compartmentalization and abundance within cells is predominantly governed by the process of vesicle/membrane transport, which plays a crucial role in several signaling cascades.The regulation of membrane transport in eukaryotic cells is governed by a diverse array of proteins. Recent developments in genomics have provided new tools to study the evolutionary relationships between membrane proteins in different plant species. It is known that members of the GTPases, COP, SNAREs, Rabs, tethering factors, and PIN families play essential roles in vesicle transport between plant, animal, and microbial species. This Review presents the latest research on the plant cytoskeleton, focusing on recent developments related to the cytoskeleton and summarizing the role of various proteins in vesicle transport. In addition, the report predicts future research direction of plant cytoskeleton and vesicle trafficking, potential research priorities, and provides researchers with specific pointers to further investigate the significant link between cytoskeleton and vesicle trafficking.

Five Inhibitory Receptors Display Distinct Vesicular Distributions in Murine T Cells.

T cells can express multiple inhibitory receptors. Upon induction of T cell exhaustion in response to a persistent antigen, prominently in the anti-tumor immune response, many are expressed simultaneously. Key inhibitory receptors are CTLA-4, PD-1, LAG3, TIM3, and TIGIT, as investigated here. These receptors are important as central therapeutic targets in cancer immunotherapy. Inhibitory receptors are not constitutively expressed on the cell surface, but substantial fractions reside in intracellular vesicular structures. It remains unresolved to which extent the subcellular localization of different inhibitory receptors is distinct. Using quantitative imaging of subcellular distributions and plasma membrane insertion as complemented by proximity proteomics and biochemical analysis of the association of the inhibitory receptors with trafficking adaptors, the subcellular distributions of the five inhibitory receptors were discrete. The distribution of CTLA-4 was most distinct, with preferential association with lysosomal-derived vesicles and the sorting nexin 1/2/5/6 transport machinery. With a lack of evidence for the existence of specific vesicle subtypes to explain divergent inhibitory receptor distributions, we suggest that such distributions are driven by divergent trafficking through an overlapping joint set of vesicular structures. This extensive characterization of the subcellular localization of five inhibitory receptors in relation to each other lays the foundation for the molecular investigation of their trafficking and its therapeutic exploitation.

Bacterial outer membrane vesicles provide an alternative pathway for trafficking of Escherichia coli O157 type III secreted effectors to epithelial cells.

IMPORTANCE: Bacteria can package protein cargo into nanosized membrane blebs that are shed from the bacterial membrane and released into the environment. Here, we report that a type of pathogenic bacteria called enterohemorrhagic Escherichia coli O157 (EHEC) uses their membrane blebs (outer membrane vesicles) to package components of their type 3 secretion system and send them into host cells, where they can manipulate host signaling pathways including those involved in infection response, such as immunity. Usually, EHEC use a needle-like apparatus to inject these components into host cells, but packaging them into membrane blebs that get taken up by host cells is another way of delivery that can bypass the need for a functioning injection system.

Organelles are miscommunicating: Membrane contact sites getting hijacked by pathogens.

Membrane Contact Sites (MCS) are areas of close apposition of organelles that serve as hotspots for crosstalk and direct transport of lipids, proteins and metabolites. Contact sites play an important role in Ca2+ signalling, phospholipid synthesis, and micro autophagy. Initially, altered regulation of vesicular trafficking was regarded as the key mechanism for intracellular pathogen survival. However, emerging studies indicate that pathogens hijack MCS elements - a novel strategy for survival and replication in an intracellular environment. Several pathogens exploit MCS to establish direct contact between organelles and replication inclusion bodies, which are essential for their survival within the cell. By establishing this direct control, pathogens gain access to cytosolic compounds necessary for replication, maintenance, escaping endocytic maturation and circumventing lysosome fusion. MCS components such as VAP A/B, OSBP, and STIM1 are targeted by pathogens through their effectors and secretion systems. In this review, we delve into the mechanisms which operate in the evasion of the host immune system when intracellular pathogens hostage MCS. We explore targeting MCS components as a novel therapeutic approach, modifying molecular pathways and signalling to address the disease's mechanisms and offer more effective, tailored treatments for affected individuals.