Large Organelles Research Articles

Dynamic protein assembly and architecture of the large solitary membraneless organelle during germline development in the wasp Nasonia vitripennis.

Throughout metazoa, germ cells assemble RNA-protein organelles (germ granules). In Drosophila ovaries, perinuclear nuage forms in the nurse cells, while compositionally similar polar granules form in the oocyte. A similar system appears to exist in the distantly related (∼350 million years) wasp Nasonia, with some surprising divergences. Nuage is similarly formed in Nasonia, except that anterior nurse cells accumulate significantly more nuage, in association with high levels of DNA double-strand breaks, suggesting that increased transposon activity in anterior is silenced by high nuage levels. In the oocyte, the germ plasm forms a single granule that is 40 times larger than a homologous Drosophila polar granule. While conserved germ granule proteins are recruited to the oosome, they show unusual localization: Tudor protein forms a shell encapsulating the embryonic oosome, while small Oskar/Vasa/Aubergine granules coalesce interiorly. Wasp Vasa itself is unusual since it has an alternative splice form that includes a novel nucleoporin-like phenylalanine-glycine repeat domain. Our work is consistent with the high degree of evolutionary plasticity of membraneless organelles and describes new experimental model and resources to study biomolecular condensates.

Live-cell imaging under centrifugation characterized the cellular force for nuclear centration in the Caenorhabditis elegans embryo

Organelles in cells are appropriately positioned, despite crowding in the cytoplasm. However, our understanding of the force required to move large organelles, such as the nucleus, inside the cytoplasm is limited, in part owing to a lack of accurate methods for measurement. We devised a method to apply forces to the nucleus of living Caenorhabditis elegans embryos to measure the force generated inside the cell. We used a centrifuge polarizing microscope to apply centrifugal force and orientation-independent differential interference contrast microscopy to characterize the mass density of the nucleus and cytoplasm. The cellular forces moving the nucleus toward the cell center increased linearly at ~12 pN/μm depending on the distance from the center. The frictional coefficient was ~980 pN s/μm. The measured values were smaller than the previously reported estimates for sea urchin embryos. The forces were consistent with the centrosome-organelle mutual pulling model for nuclear centration. The frictional coefficient was reduced when microtubules were shorter or detached from nuclei in mutant embryos, demonstrating the contribution of astral microtubules. Finally, the frictional coefficient was higher than a theoretical estimate, indicating the contribution of uncharacterized properties of the cytoplasm.

The mutual influence of microtubules and the cortical ER on their coordinated organisation.

The endoplasmic reticulum (ER) is the largest organelle in terms of membrane content, occupying the entire cytoplasmic volume. It is tethered to the cell cortex through ER-plasma membrane contact sites (EPCS). Previous studies have shown that EPCSs labelled by VAP27 align with cortical microtubules, and that ER tubules elongate along microtubules. Here, we addressed the question whether this relationship is bidirectional, with EPCSs influencing microtubule organisation. Using TIRF microscopy to track EPCSs and microtubule dynamics simultaneously, we demonstrate that while EPCSs remain stable, microtubules are highly dynamic and can adjust their positioning based on nearby EPCS in Arabidopsis cotyledon epidermis. In lobes of epidermal cells enclosed by two indentations, where microtubules bundle together, EPCSs flank the bundles and exhibit a distinctive arrangement, forming symmetric arcs in relation to the lobe axis. In guard cells, transversely oriented ER tubules co-align with microtubules. Disrupting microtubules with the drug oryzalin leads to transient guard cells-ER remodelling, followed by its reorganisation into transverse tubules before microtubule recovery. Taken together our observations suggest, that the positioning of EPCSs and cortical microtubules, can affect each other and the organisation of cortical ER.

Large attachment organelle mediates interaction between Nanobdellota archaeon YN1 and its host.

DPANN archaea are an enigmatic superphylum that are difficult to isolate and culture in the laboratory due to their specific culture conditions and apparent ectosymbiotic lifestyle. Here, we successfully isolated and cultivated a coculture system of a novel Nanobdellota archaeon YN1 and its host Sulfurisphaera ohwakuensis YN1HA. We characterized the coculture system by complementary methods, including metagenomics and metabolic pathway analysis, fluorescence microscopy, and high-resolution electron cryo-tomography (cryoET). We show that YN1 is deficient in essential metabolic processes and requires host resources to proliferate. CryoET imaging revealed an enormous attachment organelle present in the YN1 envelope that forms a direct interaction with the host cytoplasm, bridging the two cells. Together, our results unravel the molecular and structural basis of ectosymbiotic relationship between YN1 and YN1HA. This research broadens our understanding of DPANN biology and the versatile nature oftheir ectosymbiotic relationships.

Gigantic animal cells suggest organellar scaling mechanisms across a 50-fold range in cell volume.

The largest cells are orders of magnitude bigger than the smallest cells. Organelle content scales to maintain cell function, with different organelles increasing in volume, length, or number as cells increase in size. Scaling may also reflect functional demands placed on organelles by increased cell size. Amphibians exhibit exceptional diversity in cell size. Using transmission electron microscopy, we analyzed 3 species whose enterocyte cell volumes range from 228 to 10,593 μm3. We show that nuclear volume increases by an increase in radius while mitochondrial volume increases by an increase in total network length; the endoplasmic reticulum and Golgi apparatus, with their complex shapes, are intermediate. Notably, all 4 organelle types increase in total volume proportional to cell volume, despite variation in functional (i.e., metabolic, transport) demands. This pattern suggests that organellar building blocks are incorporated into more or larger organelles following the same rules across species that vary ~50-fold in cell sizes, consistent with a "limited precursor" model for organellar scaling that, in turn, assumes equivalent cytoplasmic concentrations of organellar building block proteins. Taken together, our results lead us to hypothesize that salamanders have evolved increased biosynthetic capacity to maintain functional protein concentrations despite huge cell volumes.

Vacuolar H+-ATPase and BZR1 form a feedback loop to regulate the homeostasis of BR signaling in Arabidopsis

Brassinosteroid (BR) is a vital plant hormone that regulates plant growth and development. BRASSINAZOLE RESISTANT 1 (BZR1) is a key transcription factor in BR signaling, and its nucleocytoplasmic localization is crucial for BR signaling. However, the mechanisms that regulate BZR1 nucleocytoplasmic distribution and thus the homeostasis of BR signaling remain largely unclear. The vacuole is the largest organelle in mature plant cells and plays a key role in maintenance of cellular pH, storage of intracellular substances, and transport of ions. In this study, we uncovered a novel mechanism of BR signaling homeostasis regulated by the vacuolar H+-ATPase (V-ATPase) and BZR1 feedback loop. Our results revealed that the vha-a2 vha-a3 mutant (vha2, lacking V-ATPase activity) exhibits enhanced BR signaling with increased total amount of BZR1, nuclear-localized BZR1, and the ratio of BZR1/phosphorylated BZR1 in the nucleus. Further biochemical assays revealed that VHA-a2 and VHA-a3 of V-ATPase interact with the BZR1 protein through a domain that is conserved across multiple species. VHA-a2 and VHA-a3 negatively regulate BR signaling by interacting with BZR1 and promoting its retention in the tonoplast. Interestingly, a series of molecular analyses demonstrated that nuclear-localized BZR1 could bind directly to specific motifs in the promoters of VHA-a2 and VHA-a3 to promote their expression. Taken together, these results suggest that V-ATPase and BZR1 may form a feedback regulatory loop to maintain the homeostasis of BR signaling in Arabidopsis, providing new insights into vacuole-mediated regulation of hormone signaling.

Machine perfusion preservation with hemoglobin based oxygen vesicles alleviate ultrastructural damages in porcine liver donated after cardiac death.

Midthermic machine perfusion (MMP) of post-circulatory arrest donor liver grafts has the advantage of preserving the functional ultrastructure of hepatocytes in donor grafts. It was reported that oxygenation during MMP reduces portal venous resistance and increases bile production. The MMP with hemoglobin-based oxygen vesicles (HbV) keeps the lower aspartate aminotransferase level (an indicator of liver injury) and maintains the functional ultrastructure of mitochondria in the hepatocytes. To evaluated differences of ultrastructural damages in donor livers between the MMP with and without HbV, porcine liver grafts after 60 min of warm ischemia were perfused at 22°C for 4 h with or without HbV, and a part of liver grafts were analyzed by transmission electron microscopy (TEM) and osmium-maceration scanning electron microscopy (OM-SEM). The remaining grafts were perfused with autologous blood at 38°C for 2 h in an isolated liver reperfusion model (IRM) that mimics the inside of the body after transplantation, and then analyzed by TEM and OM-SEM. Hepatocytes after MMP had small round mitochondria with rod-shaped cristae and reticulovesicular rough endoplasmic reticulum (rER) in both HbV(+) and HbV(-) livers. After IRM of HbV(+) livers, the well-developed lamellar rER was often found in hepatocytes. Liver sinusoidal endothelial cells (LSECs) after MMP contained some large vacuolar structures containing amorphous garbage in the cytoplasm, and their size along with appearance frequency were smaller and lower, respectively, in HbV(+) livers than HbV(-). Oxygenation during the MMP by using HbV suppressed the ultrastructural damages in donor livers, in particular for the LSECs. RESEARCH HIGHLIGHTS: Liver sinusoidal endothelial cells after midthermic machine perfusion had large vacuolar organelles with amorphous garbage. Oxygenation during the perfusion made them less and smaller, ultrastructurally supporting its utility.

SARS-CoV-2 infection and viral fusogens cause neuronal and glial fusion that compromises neuronal activity.

Numerous viruses use specialized surface molecules called fusogens to enter host cells. Many of these viruses, including the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), can infect the brain and are associated with severe neurological symptoms through poorly understood mechanisms. We show that SARS-CoV-2 infection induces fusion between neurons and between neurons and glia in mouse and human brain organoids. We reveal that this is caused by the viral fusogen, as it is fully mimicked by the expression of the SARS-CoV-2 spike (S) protein or the unrelated fusogen p15 from the baboon orthoreovirus. We demonstrate that neuronal fusion is a progressive event, leads to the formation of multicellular syncytia, and causes the spread of large molecules and organelles. Last, using Ca2+ imaging, we show that fusion severely compromises neuronal activity. These results provide mechanistic insights into how SARS-CoV-2 and other viruses affect the nervous system, alter its function, and cause neuropathology.

Pharmacoinformatic Investigation of Silymarin as a Potential Inhibitor against Nemopilema nomurai Jellyfish Metalloproteinase Toxin-like Protein.

Jellyfish stings pose a major threat to swimmers and fishermen worldwide. These creatures have explosive cells containing one large secretory organelle called a nematocyst in their tentacles, which contains venom used to immobilize prey. Nemopilema nomurai, a venomous jellyfish belonging to the phylum Cnidaria, produces venom (NnV) comprising various toxins known for their lethal effects on many organisms. Of these toxins, metalloproteinases (which belong to the toxic protease family) play a significant role in local symptoms such as dermatitis and anaphylaxis, as well as systemic reactions such as blood coagulation, disseminated intravascular coagulation, tissue injury, and hemorrhage. Hence, a potential metalloproteinase inhibitor (MPI) could be a promising candidate for reducing the effects of venom toxicity. For this study, we retrieved the Nemopilema nomurai venom metalloproteinase sequence (NnV-MPs) from transcriptome data and modeled its three-dimensional structure using AlphaFold2 in a Google Colab notebook. We employed a pharmacoinformatics approach to screen 39 flavonoids and identify the most potent inhibitor against NnV-MP. Previous studies have demonstrated the efficacy of flavonoids against other animal venoms. Based on our analysis, Silymarin emerged as the top inhibitor through ADMET, docking, and molecular dynamics analyses. In silico simulations provide detailed information on the toxin and ligand binding affinity. Our results demonstrate that Silymarin's strong inhibitory effect on NnV-MP is driven by hydrophobic affinity and optimal hydrogen bonding. These findings suggest that Silymarin could serve as an effective inhibitor of NnV-MP, potentially reducing the toxicity associated with jellyfish envenomation.

Wuzang Wenyang Huayu Tang promoting learning-memory ability in vascular dementia rats via brain-gut-microbiome axis

IntroductionLots of research has revealed that vascular dementia (VD), which is closely related to the metabolic disorders of intestinal microbiota, can be treated through traditional Chinese drug by regulating the brain-gut-microbiome axis. However, the specific interaction between VD and intestinal microflora, as well as the mechanism of Wuzang Wenyang Huayu Tang (WWHT) in treating VD by regulating the brain-gut-microbiome axi remains unknown on the whole and needs to be further studied. ObjectivesTo explore the effect of WWHT on the learning and memory ability of rats with VD based on brain-gut-microbiome axis. MethodsIn this study, taking the specific pathogen-free (SPF) SD rats as the subjects, a vascular dementia rat model was created by using the two-vessel occlusion model (2-VO). The subjects were divided into six groups, namely control group, sham group, VD model group, the piracetam group, the WWHT group, and the WWHT combined with piracetam (WCP) group. After 22 days of the gavage administration method, the Morris water maze experiment was adopted to determine the learning and memory abilities of each group through the water maze, observe the effects of hematoxylin and eosin (HE) staining and transmission electron microscopy in the hippocampal area of each group, and analyze the relevant proteins in the whole brain by Western Blot (WB). In addition, the feces of each group were collected and analyzed by fecal metabolomics combined with 16S rRNA gene sequencing to explore the interaction between VD and intestinal microbiota. ResultsThrough Morris water maze experiment, it was found that the learning and memory ability of VD rats in WWHT, piracetam and WCP groups was improved. The morphology of hippocampal neuron cells were improved in the WWHT group and the number of hippocampal neuron cells were increased in the VD rats by HE. By observing the ultrastructure of hippocampal cells, it can be found that the VD model group showed cell membrane edema and large organelle lysis. The ultrastructure of the hippocampus of all the administered groups was improved. In terms of the effect on the expression levels of related proteins in the whole brain of rats, compared with the VD model group, the expression levels of Caspase-3 protein decreased in all groups after drug administration. The protein expression levels of APG5L/ATG5, Beclin-1 and LC3A/B were upregulated, indicating that the drug administration group could regulate cellular autophagy in the whole brain to further protect brain tissue. Fecal metabolite profiles revealed 27 different metabolites related to VD. These metabolites are mainly involved in related metabolic pathways, such as vitamin B metabolism, purine metabolism, pyrimidine metabolism, and histidine metabolism. Further analysis of the gut microbiota by 16S rRNA gene sequencing data showed that WWHT-treated VD rats were different in gut microbiota composition. DiscussionThrough the effect of brain-gut-microbiome axis, WWHT regulates the metabolism of vitamin B6 and tryptophan in VD rats, which can improve the intestinal microbiota of VD rats and regulate autophagy in the brain to improve cell morphology in the hippocampus, thus increasing the number of hippocampal cells to protect hippocampal neuron cells2510100, and finally improving the learning and memory ability of VD rats.

Refractile bodies of Eimeria tenella are proteinaceous membrane-less organelles that undergo dynamic changes during infection.

Refractile bodies (RB) are large membrane-less organelles (MLO) of unknown function found as a prominent mismatched pair within the sporozoite stages of all species of Eimeria, parasitic coccidian protozoa. High resolution imaging methods including time-lapse live confocal microscopy and serial block face-scanning electron microscopy (SBF-SEM) were used to investigate the morphology of RB and other intracellular organelles before and after sporozoite invasion of host cells. Live cell imaging of MDBK cells infected with E. tenella sporozoites confirmed previous reports that RB reduce from two to one post-infection and showed that reduction in RB number occurs via merger of the anterior RB with the posterior RB, a process that lasts 20-40 seconds and takes place between 2- and 5-hours post-infection. Ultrastructural studies using SBF-SEM on whole individual sporozoites, both pre- and post-host cell invasion, confirmed the live cell imaging observations and showed also that changes to the overall sporozoite cell shape accompanied RB merger. Furthermore, the single RB post-merger was found to be larger in volume than the two RB pre-merger. Actin inhibitors were used to investigate a potential role for actin in RB merger, Cytochalasin D significantly inhibited both RB merger and the accompanying changes in sporozoite cell shape. MLOs in eukaryotic organisms are characterised by their lack of a membrane and ability to undergo liquid-liquid phase separation (LLPS) and fusion, usually in an actin-mediated fashion. Based on the changes in sporozoite cell shape observed at the time of RB merger together with a potential role for actin in this process, we propose that RB are classed as an MLO and recognised as one of the largest MLOs so far characterised.

Size- and position-dependent cytoplasm viscoelasticity through hydrodynamic interactions with the cell surface

Many studies of cytoplasm rheology have focused on small components in the submicrometer scale. However, the cytoplasm also baths large organelles like nuclei, microtubule asters, or spindles that often take significant portions of cells and move across the cytoplasm to regulate cell division or polarization. Here, we translated passive components of sizes ranging from few up to ~50 percents of the cell diameter, through the vast cytoplasm of live sea urchin eggs, with calibrated magnetic forces. Creep and relaxation responses indicate that for objects larger than the micron size, the cytoplasm behaves as a Jeffreys material, viscoelastic at short timescales, and fluidizing at longer times. However, as component size approached that of cells, cytoplasm viscoelastic resistance increased in a nonmonotonic manner. Flow analysis and simulations suggest that this size-dependent viscoelasticity emerges from hydrodynamic interactions between the moving object and the static cell surface. This effect also yields to position-dependent viscoelasticity with objects initially closer to the cell surface being harder to displace. These findings suggest that the cytoplasm hydrodynamically couples large organelles to the cell surface to restrain their motion, with important implications for cell shape sensing and cellular organization.

Wandering the intracellular maze.

The endoplasmic reticulum (ER) is the largest organelle in eukaryotic cells and consists of a dynamic network of stacked sheets and lipid membrane tubules stretching from the nucleus to the cell periphery. The ER serves an essential role in the distribution of lipids, ions and proteins throughout the cell, but the impact of network morphology on transport remains poorly understood. To that end, analytic mean first passage times for particles diffusing on extracted ER network structures are used as a measure of accessibility for different regions of the ER, which exhibit a wide heterogeneity due to varying tubule density and connectivity patterns. Moving beyond average search times, we combine agent-based simulations of particles diffusing on tubular networks with in vivo data on the spreading of photoactivated membrane proteins in COS-7 cells, in order to quantify how local ER structure determines protein spreading. We demonstrate that static local connectivity serves to explain some but not all of the observed behavior of proteins in the ER. Network dynamics, such as tubule growth and rearrangement are also implicated. In order to quantify these effects, we develop a dynamic model of the peripheral ER that aims to capture its unique, constantly evolving structure. This minimal network model reproduces key features that are observed experimentally in the ER of COS-7 cells such as length and area distributions. Intriguingly, the effective node diffusivity predicted from the model is similar to the observed diffusivity of ER exit site structures. Pairing analytic calculations and theoretical modeling with live-cell imaging, we highlight the role the heterogeneous ER network morphology plays in the non-uniform transport of proteins and gain fresh insights into the dynamic nature of the network itself.

Unbiased Quantification of Golgi Scattering and Golgi-Centrosome Association.

The vertebrate Golgi complex is a large dynamic organelle which undergoes morphological changes and fragmentation both as a part of normal physiological dynamics and under disease conditions. The Golgi is known to have a functionally important relationship with the centrosome. The extent of the spatial association between these two organelles varies in a dynamic and regulated manner. It is essential to have a reliable unbiased approach to evaluate Golgi volume, Golgi extension/scattering in the 3D cell space, and spatial association of the Golgi with the centrosome. It is also important that each of these features is evaluated by a simple metric, one measurement per cell, so that the variability and deviations in the cell population can be easily assessed. Here, we present an approach to analyze confocal microscopy image stacks to easily measure Golgi volume, scattering, and association with the centrosome. The approach is based on a custom MATLAB script, provided here as a supplement, and also uses widely available software (ImageJ and/or Imaris). The output of the script is a table with the following parameters: Golgi volume in voxels, Golgi volume in μm3, "Golgi-Golgi" distance (averaged distance between all Golgi voxels), Golgi-centrosome distance (averaged distance between each Golgi voxel and the nearest mother centriole), and centrosome-centrosome distance (for cells with duplicated centrosome, the distance between the mother centrioles). The approach can also be applied to analyze distribution of any fluorescently- labeled structure within a cell and its association with the centrosome or any single point within the cell volume.

Studies of functional properties of espin 1: Its interaction to actin filaments.

Actin is a multifunctional biomolecule that forms not only basic structural bodies such as filopodia and lamellipodia, but also large microvilli-like organelles like stereocilia. Actin consists of four sub-domains (S1, S2, S3, and S4), and the "target-binding groove" formed between S1 and S3 is the major binding site for various actin binding proteins. Actin filament dynamics are regulated by numerous actin binding proteins with different mechanisms of actin binding, assembly, and disassembly such as actin severing, branching, and bundling. Ectoplasmic specialization protein 1 (espin 1) is an actin binding and bundling protein that is specifically implicated in the elongation and stabilization of stereocilia as a binding partner with myosin III. However, little is known about the molecular structure, actin bundling, and stabilizing mechanism of espin 1; hence, we investigated the interaction between actin and espin 1 through structural data. In this study, we first purified human espin 1 in an E. coli system following a new detergent-free approach and then demonstrated the 2D structure of full-length espin 1 using transmission electron microscopy along with Nickel nitrilotriacetic acid nanogold labeling and 2D averaging using SPIDER. Furthermore, we also determined the espin 1 binding domain of actin using a co-sedimentation assay along with gelsolin and myosin S1. These findings are not only beneficial for understanding the actin binding and bundling mechanism of espin 1, but also shed light on its elongation, stabilization, and tip-localization mechanisms with myosin III. This study thus provides a basis for understanding the molecular structure of espin 1 and can contribute to various hearing-related diseases, such as hearing loss and vestibular dysfunction.

ER-Phagy: Quality and Quantity Control of the Endoplasmic Reticulum by Autophagy.

The endoplasmic reticulum (ER) is the largest organelle and has multiple roles in various cellular processes such as protein secretion, lipid synthesis, calcium storage, and organelle biogenesis. The quantity and quality of this organelle are controlled by the ubiquitin-proteasome system and autophagy (termed "ER-phagy"). ER-phagy is defined as the degradation of part of the ER by the vacuole or lysosomes, and there are at least two types of ER-phagy: macro-ER-phagy and micro-ER-phagy. In macro-ER-phagy, ER fragments are enclosed by autophagosomes, which is mediated by ER-phagy receptors. In micro-ER-phagy, a portion of the ER is engulfed directly by the vacuole or lysosomes. In these two pathways, some proteins in the ER lumen can be recognized selectively and subjected to ER-phagy. This review summarizes our current knowledge of ER-phagy, focusing on its membrane dynamics, molecular mechanisms, substrate specificity, and physiological significance.

Endoplasmic reticulum-targeted polymer dots encapsulated with ultrasonic synthesized near-infrared carbon nanodots and their application for in vivo monitoring of Cu2+

Endoplasmic reticulum (ER) is the largest organelle in eukaryotic cells and plays a variety of functions in living cells include protein folding, calcium homeostasis, and lipid biosynthesis. Normal function of ER is crucial for cell survival, while disequilibrium of ER can cause misfolding of proteins and ER stress, leading to many serious diseases. It has been documented that ER stress is closely related to the metabolism of Cu2+, as ER is the main intracellular accumulation space of Cu2+ and toxic reactive oxygen species can be generated by Cu2+ via Fenton and Haber-Weiss reactions. In this context, developing a powerful tool capable of selective and sensitive monitoring of Cu2+ in ER and investigating its role in physiological and pathological processes is of great importance. Herein, we report the first ER targeted near infrared (NIR) nanosensor, polymer dots encapsulated with NIR hydrophobic carbon nanodots, for detecting Cu2+ in biosystems. This nanosensor with stable fluorescence showed a fast response toward Cu2+ (120 s) and can be used for the quantification of Cu2+ in a linear range covering from 0.25 to 9.0 μM with a detection limit of 13 nM. In addition, the fluorescence variations of the nanosensor are remarkably specific to Cu2+ in comparison with the other metal ions and amino acids. Moreover, the developed nanosensor exhibited low cytotoxicity, good biocompatibility, and ER targeting ability. Because of these excellent spectroscopic features, the nanosensor was successfully utilized for visualizing Cu2+ fluctuations at the living cell, zebrafish and mouse levels, which further proved its potential application in biological systems.

Inside Front Cover: Continuous organelle separation in an insulator‐based dielectrophoretic device

DOI: 10.1002/elps.202100326 The (inside front) cover art shows a scanning electron image of the device used for the iDEP-based continuous separation of organelles. The inset demonstrates the working mechanism of the devices showing a mixture of analytes injected in the center of a post-array and being separated as they are steered through the posts. Small and large organelles are collected at the opposing outlets of the post-array.

The ER-Mitochondria Interface as a Dynamic Hub for T Cell Efficacy in Solid Tumors.

The endoplasmic reticulum (ER) is a large continuous membranous organelle that plays a central role as the hub of protein and lipid synthesis while the mitochondria is the principal location for energy production. T cells are an immune subset exhibiting robust dependence on ER and mitochondrial function based on the need for protein synthesis and secretion and metabolic dexterity associated with foreign antigen recognition and cytotoxic effector response. Intimate connections exist at mitochondrial-ER contact sites (MERCs) that serve as the structural and biochemical platforms for cellular metabolic homeostasis through regulation of fission and fusion as well as glucose, Ca2+, and lipid exchange. Work in the tumor immunotherapy field indicates that the complex interplay of nutrient deprivation and tumor antigen stimulation in the tumor microenvironment places stress on the ER and mitochondria, causing dysfunction in organellar structure and loss of metabolic homeostasis. Here, we assess prior literature that establishes how the structural interface of these two organelles is impacted by the stress of solid tumors along with recent advances in the manipulation of organelle homeostasis at MERCs in T cells. These findings provide strong evidence for increased tumor immunity using unique therapeutic avenues that recharge cellular metabolic homeostasis in T cells.

A ratiometric fluorescence probe for imaging endoplasmic reticulum (ER) hypochlorous acid in living cells undergoing excited state intramolecular proton transfer

Hypochlorous acid (HOCl), one of the most important ROS in living organisms, appears to serve an important role in the immune system in vivo. Endoplasmic reticulum (ER), the largest organelle in cells, manages many biological processes connected to vital activities. To better obtain insight into the relationship of ER stress and HOCl level, a ratiometric fluorescent probe RHE, based on rhodamine combined with HBT and ER-targeting group, was designed and synthesized for HOCl detection in the ER. Probe RHE shows a large stokes shift about 155 nm, which is derived to ESIPT principle. In addition, probe RHE exhibited excellent properties such as fast response (<80 s), high sensitivity with a low detection limit (40 nM), high selectivity and anti-interference. Moreover, probe RHE displayed an excellent ER-targeting ability and had been successfully applied for detection of exogenous and endogenous HOCl in HepG2 cells.