Cytoplasmic Material Research Articles

The Immune Escape Strategy of Rabies Virus and Its Pathogenicity Mechanisms.

In contrast to most other rhabdoviruses, which spread by insect vectors, the rabies virus (RABV) is a very unusual member of the Rhabdoviridae family, since it has evolved to be fully adapted to warm-blooded hosts and spread directly between them. There are differences in the immune responses to laboratory-attenuated RABV and wild-type rabies virus infections. Various investigations showed that whilst laboratory-attenuated RABV elicits an innate immune response, wild-type RABV evades detection. Pathogenic RABV infection bypasses immune response by antagonizing interferon induction, which prevents downstream signal activation and impairs antiviral proteins and inflammatory cytokines production that could eliminate the virus. On the contrary, non-pathogenic RABV infection leads to immune activation and suppresses the disease. Apart from that, through recruiting leukocytes into the central nervous system (CNS) and enhancing the blood-brain barrier (BBB) permeability, which are vital factors for viral clearance and protection, cytokines/chemokines released during RABV infection play a critical role in suppressing the disease. Furthermore, early apoptosis of neural cells limit replication and spread of avirulent RABV infection, but street RABV strains infection cause delayed apoptosis that help them spread further to healthy cells and circumvent early immune exposure. Similarly, a cellular regulation mechanism called autophagy eliminates unused or damaged cytoplasmic materials and destroy microbes by delivering them to the lysosomes as part of a nonspecific immune defense mechanism. Infection with laboratory fixed RABV strains lead to complete autophagy and the viruses are eliminated. But incomplete autophagy during pathogenic RABV infection failed to destroy the viruses and might aid the virus in dodging detection by antigen-presenting cells, which could otherwise elicit adaptive immune activation. Pathogenic RABV P and M proteins, as well as high concentration of nitric oxide, which is produced during rabies virus infection, inhibits activities of mitochondrial proteins, which triggers the generation of reactive oxygen species, resulting in oxidative stress, contributing to mitochondrial malfunction and, finally, neuron process degeneration.

Biogenesis of omegasomes and autophagosomes in mammalian autophagy.

Autophagy is a highly conserved catabolic pathway that maintains cellular homeostasis by promoting the degradation of damaged or superfluous cytoplasmic material. A hallmark of autophagy is the generation of membrane cisternae that sequester autophagic cargo. Expansion of these structures allows cargo to be engulfed in a highly selective and exclusive manner. Cytotoxic stress or starvation induces the formation of autophagosomes that sequester bulk cytoplasm instead of selected cargo. This rather nonselective pathway is essential for maintaining vital cellular functions during adverse conditions and is thus a major stress response pathway. Both selective and nonselective autophagy rely on the same molecular machinery. However, due to the different nature of cargo to be sequestered, the involved molecular mechanisms are fundamentally different. Although intense research over the past decades has advanced our understanding of autophagy, fundamental questions remain to be addressed. This review will focus on molecular principles and open questions regarding the formation of omegasomes and phagophores in nonselective mammalian autophagy.

The Antidepressant Drug Amitriptyline Affects Human SH-SY5Y Neuroblastoma Cell Proliferation and Modulates Autophagy.

Amitriptyline is a tricyclic antidepressant commonly used for depressive disorders and is prescribed off-label for several neurological conditions like neuropathic pain, migraines and anxiety. Besides their action on the reuptake of monoaminergic neurotransmitters, tricyclic antidepressants interact with several additional targets that may contribute to either therapeutic or adverse effects. Here, we investigated the effects of amitriptyline on proliferation and autophagy (i.e., an evolutionarily conserved catabolic pathway responsible for the degradation and recycling of cytoplasmic material) in human SH-SY5Y neuroblastoma cell cultures. The dose and time-dependent upregulation of the autophagy marker LC3II and the autophagy receptor p62, with the accumulation of LAMP1 positive compartments, were observed in SH-SY5Y cells exposed to the amitriptyline. These effects were accompanied by reduced cell viability and decreased clonogenic capacity, without a significant induction of apoptosis. Decrease viability and clonogenic activity were still observed in autophagy deficient Atg5-/- MEF and following pre-treatment of SH-SY5Y culture with the autophagy inhibitor chloroquine, suggesting that they were independent from autophagy modulation. Our findings demonstrate that amitriptyline acts on pathways crucial for cell and tissue homeostasis (i.e., autophagy and proliferation) and pose the basis for further studies on the potential therapeutic application of amitriptyline, as well as the consequences of its use for long-term treatments.

The Genetic Diversity of 69 Widely Used Chinese Sorghum Hybrids Released between the 1970s and 2010s

Sorghum has a long history of cultivation in China. In this study, we aimed to clarify the genetic relationships and genetic variation trends in widely used Chinese sorghum hybrids which were released from the 1970s to 2010s and attempted to analyze the changes in sorghum breeding. A total of 257 alleles were detected by 51 polymorphic SSR markers among 69 widely used hybrids; an average of 5.04 alleles were detected by each marker. The average Shannon’s index and polymorphism information content (PIC) of markers were 1.39 and 0.70, respectively. Nei’s genetic diversity index continuously increased in four different breeding development stages (1973–1982, 1983–1992, 1993–2002, and 2003–2014). Genetic diversity gradually increased among the sorghum hybrids. Genetic similarity coefficients in the four breeding development stages first showed an increasing trend, and then a decreasing trend, finally stabilizing with an average value of 0.65. The genetic similarity changes in hybrids in early and late maturing areas were consistent at different breeding development stages. The genetic similarity coefficients in late maturing areas were constantly higher than those in the early maturing areas. This is related to China’s creative utilization of A2 cytoplasmic male sterile materials in the 1990s. A cluster analysis determined that 69 hybrids were divided into two groups, A and B. Group A could be further subdivided into four subgroups. These findings could provide a reference for parental selection and hybrid breeding in sorghum improvement programs.

Dinotoms possess two evolutionary distinct autophagy-related ubiquitin-like conjugation systems

Autophagy is an intracellular degradation mechanism by which cytoplasmic materials are delivered to and degraded in the lysosome-fused autophagosome (autolysosome) and proposed to have been established at an early stage of eukaryotic evolution. Dinoflagellates harboring endosymbiotic diatoms (so-called “dinotoms”), which retain their own nuclei and mitochondria in addition to plastids, have been investigated as an intermediate toward the full integration of a eukaryotic phototroph into the host-controlled organelle (i.e., plastid) through endosymbiosis. Pioneering studies systematically evaluated the degree of host governance on several metabolic pathways in the endosymbiotic diatoms (ESDs). However, little attention has been paid to the impact of the endosymbiotic lifestyle on the autophagy operated in the ESDs. In this study, we searched for ATG3, ATG4, ATG5, ATG7, ATG8, ATG10, and ATG12, which are required for autophagosome formation, in the RNA-seq data from dinotoms Durinskia baltica and Kryptoperidinium foliaceum. We detected two evolutionally distinct sets of the ATG proteins in the dinotom species, one affiliated with the dinoflagellate homologs and the other with the diatom homologs in phylogenetic analyses. The results suggest that the ATG proteins descended from the diatom taken up by the dinoflagellate host persist for autophagosome formation and, most likely, autophagy.

Assessing the Presence of Phosphoinositides on Autophagosomal Membrane in Yeast by Live Cell Imaging.

The formation of autophagosomes mediating the sequestration of cytoplasmic materials is the central step of autophagy. Several phosphoinositides, which are signaling molecules on the membrane, are involved in autophagy. However, it is not always clear whether these phosphoinositides act directly at the site of autophagosome formation, or indirectly via the regulation of other steps or pathways. To address this question, we used a set of phosphoinositide probes to systematically examine their potential presence on autophagosomal membranes in yeast (Saccharomyces cerevisiae). We verified the specificity of these probes using mutant cells deficient in the production of the corresponding phosphoinositides. We then examined starved yeast cells co-expressing a phosphoinositide probe together with an autophagosomal membrane marker, 2Katushka2S-Atg8. Our data revealed that PtdIns(4,5)P2 and PtdIns(3,5)P2 were mainly present on the plasma membrane and vacuolar membrane, respectively. We observed only occasional co-localization between the PtdIns(4)P probe and Atg8, some of which may represent the transient passage of a PtdIns(4)P-containing structure near the autophagosomal membrane. In contrast, substantial colocalization of the PtdIns(3)P probe with Atg8 was observed. Taken together, our data indicate that only PtdIns(3)P is present in a substantial amount on the autophagosomal membrane. For other phosphoinositides involved in autophagy, either their presence on the autophagosomal membrane is very transient, or they act on other cellular membranes to regulate autophagy.

Ultrastructural Characterization of PBMCs and Extracellular Vesicles in Multiple Sclerosis: A Pilot Study.

Growing evidence identifies extracellular vesicles (EVs) as important cell-to-cell signal transducers in autoimmune disorders, including multiple sclerosis (MS). If the etiology of MS still remains unknown, its molecular physiology has been well studied, indicating peripheral blood mononuclear cells (PBMCs) as the main pathologically relevant contributors to the disease and to neuroinflammation. Recently, several studies have suggested the involvement of EVs as key mediators of neuroimmune crosstalk in central nervous system (CNS) autoimmunity. To assess the role of EVs in MS, we applied electron microscopy (EM) techniques and Western blot analysis to study the morphology and content of plasma-derived EVs as well as the ultrastructure of PBMCs, considering four MS patients and four healthy controls. Through its exploratory nature, our study was able to detect significant differences between groups. Pseudopods and large vesicles were more numerous at the plasmalemma interface of cases, as were endoplasmic vesicles, resulting in an activated aspect of the PBMCs. Moreover, PBMCs from MS patients also showed an increased number of multivesicular bodies within the cytoplasm and amorphous material around the vesicles. In addition, we observed a high number of plasma-membrane-covered extensions, with multiple associated large vesicles and numerous autophagosomal vacuoles containing undigested cytoplasmic material. Finally, the study of EV cargo evidenced a number of dysregulated molecules in MS patients, including GANAB, IFI35, Cortactin, Septin 2, Cofilin 1, and ARHGDIA, that serve as inflammatory signals in a context of altered vesicular dynamics. We concluded that EM coupled with Western blot analysis applied to PBMCs and vesiculation can enhance our knowledge in the physiopathology of MS.

The new kids on the block: RNA-binding proteins regulate autophagy in disease.

Mammalian autophagy is a highly regulated and conserved cellular homeostatic process. Its existence allows the degradation of self-components to mediate cell survival in different stress conditions. Autophagy is involved in the regulation of cellular metabolic needs, protecting the cell or tissue from starvation through the degradation and recycling of cytoplasmic materials and organelles to basic molecular building blocks. It also plays a critical role in eliminating damaged or harmful proteins, organelles, and intracellular pathogens. Thus, a deterioration of the process may result in pathological conditions, such as aging-associated disorders and cancer. Understanding the crucial role of autophagy in maintaining the normal physiological function of cells, tissue, or organs has led to copious and expansive research regarding the regulation of this process. So far, most of the research has revolved around transcriptional and post-translational regulation. Here, we discuss the regulation of autophagy-related (ATG) mRNA transcripts by RNA-binding proteins (RBPs). This analysis focuses on how RBPs modulate autophagy in disease. A deeper understanding of the involvement of RBPs in autophagy can facilitate further research and treatment of a variety of human diseases.

VCP/p97 UFMylation stabilizes BECN1 and facilitates the initiation of autophagy

ABSTRACT Macroautophagy/autophagy is essential for the degradation and recycling of cytoplasmic materials. The initiation of this process is determined by phosphatidylinositol-3-kinase (PtdIns3K) complex, which is regulated by factor BECN1 (beclin 1). UFMylation is a novel ubiquitin-like modification that has been demonstrated to modulate several cellular activities. However, the role of UFMylation in regulating autophagy has not been fully elucidated. Here, we found that VCP/p97 is UFMylated on K109 by the E3 UFL1 (UFM1 specific ligase 1) and this modification promotes BECN1 stabilization and assembly of the PtdIns3K complex, suggesting a role for VCP/p97 UFMylation in autophagy initiation. Mechanistically, VCP/p97 UFMylation stabilizes BECN1 through ATXN3 (ataxin 3)-mediated deubiquitination. As a key component of the PtdIns3K complex, stabilized BECN1 facilitates assembly of this complex. Re-expression of VCP/p97, but not the UFMylation-defective mutant, rescued the VCP/p97 depletion-induced increase in MAP1LC3B/LC3B protein expression. We also showed that several pathogenic VCP/p97 mutations identified in a variety of neurological disorders and cancers were associated with reduced UFMylation, thus implicating VCP/p97 UFMylation as a potential therapeutic target for these diseases. Abbreviation: ATG14:autophagy related 14; Baf A1:bafilomycin A1;CMT2Y: Charcot-Marie-Toothdisease, axonal, 2Y; CYB5R3: cytochromeb5 reductase 3; DDRGK1: DDRGK domain containing 1; DMEM:Dulbecco’smodified Eagle’s medium;ER:endoplasmic reticulum; FBS:fetalbovine serum;FTDALS6:frontotemporaldementia and/or amyotrophic lateral sclerosis 6; IBMPFD1:inclusion bodymyopathy with early-onset Paget disease with or withoutfrontotemporal dementia 1; LC-MS/MS:liquid chromatography tandem mass spectrometry; MAP1LC3B/LC3B:microtubule associated protein 1 light chain 3 beta; MS: massspectrometry; NPLOC4: NPL4 homolog, ubiquitin recognition factor;PIK3C3: phosphatidylinositol 3-kinase catalytic subunit type 3;PIK3R4: phosphoinositide-3-kinase regulatory subunit 4; PtdIns3K:phosphatidylinositol 3-kinase; RPL26: ribosomal protein L26; RPN1:ribophorin I; SQSTM1/p62: sequestosome 1; UBA5: ubiquitin likemodifier activating enzyme 5; UFC1: ubiquitin-fold modifierconjugating enzyme 1; UFD1: ubiquitin recognition factor in ERassociated degradation 1; UFL1: UFM1 specific ligase 1; UFM1:ubiquitin fold modifier 1; UFSP2: UFM1 specific peptidase 2; UVRAG:UV radiation resistance associated; VCP/p97: valosin containingprotein; WT: wild-type

Direct interaction between marine cyanobacteria mediated by nanotubes.

Microbial associations and interactions drive and regulate nutrient fluxes in the ocean. However, physical contact between cells of marine cyanobacteria has not been studied thus far. Here, we show a mechanism of direct interaction between the marine cyanobacteria Prochlorococcus and Synechococcus, the intercellular membrane nanotubes. We present evidence of inter- and intra-genus exchange of cytoplasmic material between neighboring and distant cells of cyanobacteria mediated by nanotubes. We visualized and measured these structures in xenic and axenic cultures and in natural samples. We show that nanotubes are produced between living cells, suggesting that this is a relevant system of exchange material in vivo. The discovery of nanotubes acting as exchange bridges in the most abundant photosynthetic organisms in the ocean may have important implications for their interactions with other organisms and their population dynamics.

A review on the nexus of autophagy genes from the perspective of polycystic ovary syndrome.

Polycystic ovary syndrome or PCOS is an endocrine disorder in women of reproductive age. It is a diversified multi factorial disorder and diagnosis is very complicated because of its overlapping symptoms some of which are irregular menstrual cycle, acne in face, excess level of androgen (AE), insulin resistance, obesity, cardiovascular disease, mood disorder and type 2 diabetes (T2DM). PCOS may be caused by hormonal imbalance, genetic and epigenetic vulnerability, hypothalamic and ovarian troubles. PCOS is essentially hyperandrogenimia with oligo-anovulation. This review explains the abnormal regulation of autophagy related genes and proteins in different cells at various stages which leads to the genesis of PCOS. During nutrient starvation cells face stress condition, which it tries to overcome by activating its macroautophagy mechanism and by degrading the cytoplasmic material. This provides energy to the cell facilitating its survival. Downregulation of autophagy related genes in endometria has been observed in PCOS women. PCOS can be managed by maintaining proper lifestyle and medical treatment. Healthy meals and regular exercise can prevent the excessive weight and also reduce the PCOS complications. Medicines such as metformin, clomiphene, and the oral contraceptive pill can also balance the hormonal level. The imbalance in regulation of autophagy genes has been discussed with correlation to PCOS. The different management strategies for PCOS have also been summarized.

Antimicrobial peptides designed by computational analysis of proteomes.

Antimicrobial peptides (AMPs) are promising cationic and amphipathic molecules to fight antibiotic resistance. To search for novel AMPs, we applied a computational strategy to identify peptide sequences within the organisms' proteome, including in-house developed software and artificial intelligence tools. After analyzing 150.450 proteins from eight proteomes of bacteria, plants, a protist, and a nematode, nine peptides were selected and modified to increase their antimicrobial potential. The 18 resulting peptides were validated by bioassays with four pathogenic bacterial species, one yeast species, and two cancer cell-lines. Fourteen of the 18 tested peptides were antimicrobial, with minimum inhibitory concentrations (MICs) values under 10µM against at least three bacterial species; seven were active against Candida albicans with MICs values under 10µM; six had a therapeutic index above 20; two peptides were active against A549 cells, and eight were active against MCF-7 cells under 30µM. This study's most active antimicrobial peptides damage the bacterial cell membrane, including grooves, dents, membrane wrinkling, cell destruction, and leakage of cytoplasmic material. The results confirm that the proposed approach, which uses bioinformatic tools and rational modifications, is highly efficient and allows the discovery, with high accuracy, of potent AMPs encrypted in proteins.

Transplanted human photoreceptors transfer cytoplasmic material but not to the recipient mouse retina

BackgroundThe discovery of material transfer between transplanted and host mouse photoreceptors has expanded the possibilities for utilizing transplanted photoreceptors as potential vehicles for delivering therapeutic cargo. However, previous research has not directly explored the capacity for human photoreceptors to engage in material transfer, as human photoreceptor transplantation has primarily been investigated in rodent models of late-stage retinal disease, which lack host photoreceptors.MethodsIn this study, we transplanted human stem-cell derived photoreceptors purified from human retinal organoids at different ontological ages (weeks 10, 14, or 20) into mouse models with intact photoreceptors and assessed transfer of human proteins and organelles to mouse photoreceptors.ResultsUnexpectedly, regardless of donor age or mouse recipient background, human photoreceptors did not transfer material in the mouse retina, though a rare subset of donor cells (< 5%) integrated into the mouse photoreceptor cell layer. To investigate the possibility that a species barrier impeded transfer, we used a flow cytometric assay to examine material transfer in vitro. Interestingly, dissociated human photoreceptors transferred fluorescent protein with each other in vitro, yet no transfer was detected in co-cultures of human and mouse photoreceptors, suggesting that material transfer is species specific.ConclusionsWhile xenograft models are not a tractable system to study material transfer of human photoreceptors, these findings demonstrate that human retinal organoid-derived photoreceptors are competent donors for material transfer and thus may be useful to treat retinal degenerative disease.

Molecular Mechanism of Autophagosome-Lysosome Fusion in Mammalian Cells.

In eukaryotes, targeting intracellular components for lysosomal degradation by autophagy represents a catabolic process that evolutionarily regulates cellular homeostasis. The successful completion of autophagy initiates the engulfment of cytoplasmic materials within double-membrane autophagosomes and subsequent delivery to autolysosomes for degradation by acidic proteases. The formation of autolysosomes relies on the precise fusion of autophagosomes with lysosomes. In recent decades, numerous studies have provided insights into the molecular regulation of autophagosome-lysosome fusion. In this review, an overview of the molecules that function in the fusion of autophagosomes with lysosomes is provided. Moreover, the molecular mechanism underlying how these functional molecules regulate autophagosome-lysosome fusion is summarized.

Glycogen Granules Are Degraded by Non-Selective Autophagy in Nitrogen-Starved Komagataella phaffii.

Autophagy was initially recognized as a bulk degradation process that randomly sequesters and degrades cytoplasmic material in lysosomes (vacuoles in yeast). In recent years, various types of selective autophagy have been discovered. Glycophagy, the selective autophagy of glycogen granules, is one of them. While autophagy of glycogen is an important contributor to Pompe disease, which is characterized by the lysosomal accumulation of glycogen, its selectivity is still a matter of debate. Here, we developed the Komagataella phaffii yeast as a simple model of glycogen autophagy under nitrogen starvation conditions to address the question of its selectivity. For this, we turned the self-glucosylating initiator of glycogen synthesis, Glg1, which is covalently bound to glycogen, into the Glg1-GFP autophagic reporter. Our results revealed that vacuolar delivery of Glg1-GFP and its processing to free GFP were strictly dependent on autophagic machinery and vacuolar proteolysis. Notably, this process was independent of Atg11, the scaffold protein common for many selective autophagy pathways. Importantly, the non-mutated Glg1-GFP (which synthesizes and marks glycogen) and mutated Glg1Y212F-GFP (which does not synthesize glycogen and is degraded by non-selective autophagy as cytosolic Pgk1-GFP) were equally well delivered to the vacuole and had similar levels of released GFP. Therefore, we concluded that glycogen autophagy is a non-selective process in K. phaffii yeast under nitrogen starvation conditions.

Antibacterial Effect of Eight Essential Oils against Bacteria Implicated in Bovine Mastitis and Characterization of Primary Action Mode of Thymus capitatus Essential Oil.

During the current investigation, eight essential oils (EOs) were tested for their antimicrobial activity against six species, belonging to the genus of staphylococcus, multi-resistant to antibiotics (S. epidermidis, S. cohni, S. wareneri, S. scuiri, S. chromogenes, S. pasteuri), three methicillin-resistant Staphylococcus aureus strains (MRSA) and two strains of Escherichia coli, producing extended-spectrum β-lactamase (ESBL) responsible for bovine mastitis. Our results indicated that the antimicrobial activities of eight EOs varied significantly among the types of EOs and bacterial species. Thymus capitatus and Trachyspermum ammi EOs display important antibacterial activity against all tested strains, with the inhibition zone diameters situated between 20 and 45 mm, while EOs of Artemisia absinthium, Eucalyptus globulus, Eucalyptus camaldulensis, Myrtus communis and Mentha pulegium exerted an intermediate activity. For Cymbopogon citratus, this effect depends on bacteria species. In fact, an important effect was observed against S. warneri, S. epidermidis, S. cohenii, S. pasteuri and MRSA (EC 39+) strains. In addition, the important lytic effect was observed against MRSA strains, showing that Gram-positive bacteria were more sensitive to T. capitatus EO than Gram-negative ones. Concerning the characterization of the mode action of T. capitatus, experiments of kill-time, bacteriolytic, loss of salt tolerance and loss of cytoplasmic material showed that the used EO was able to destroy cell walls and membranes followed by the loss of vital intracellular materials. In addition, it inhibits the normal synthesis of DNA, causing the bacterial death of E. coli and MRSA strains. This study shows the potential of using of EOs, particularly T. capitaus, to inhibit the growth of Gram-positive and Gram-negative bacteria multi-resistant to antibiotics causing bovine mastitis.

Rim aperture of yeast autophagic membranes balances cargo inclusion with vesicle maturation

Autophagy eliminates cytoplasmic material by engulfment in membranous vesicles targeted for lysosome degradation. Nonselective autophagy coordinates sequestration of bulk cargo with the growth of the isolation membrane (IM) in a yet-unknown manner. Here, we show that in the budding yeast Saccharomyces cerevisiae, IMs expand while maintaining a rim sufficiently wide for sequestration of large cargo but tight enough to mature in due time. An obligate complex of Atg24/Snx4 with Atg20 or Snx41 assembles locally at the rim in a spatially extended manner that specifically depends on autophagic PI(3)P. This assembly stabilizes the open rim to promote autophagic sequestration of large cargo in correlation with vesicle expansion. Moreover, constriction of the rim by the PI(3)P-dependent Atg2-Atg18 complex and clearance of PI(3)P by Ymr1 antagonize rim opening to promote autophagic maturation and consumption of small cargo. Tight regulation of membrane rim aperture by PI(3)P thus couples the mechanism and physiology of nonselective autophagy.

Revealing the mechanism of citral induced entry of Vibrio vulnificus into viable but not culturable (VBNC) state based on transcriptomics

Citral has attracted much attention as a safe and effective plant-derived bacteriostatic agent. However, the ability of citral to induce the formation of VBNC state in Vibrio vulnificus has not been evaluated. In the present study, V. vulnificus was shown to be induced to form the VBNC state at 4.5 h and 3 h of citral treatment at 4MIC and 6MIC. Moreover, the citral-induced VBNC state of V. vulnificus maintained some respiratory chain activity and was able to recover well in both APW media, APW media supplemented with 5 % (v/v) Tween 80 and 2 mg/mL sodium pyruvate. Field emission and transmission electron microscopy showed that the external structure of the citral-induced VBNC V. vulnificus cells was shortened to short rods, with folded cell membrane, rough cell surface, and dense cytoplasm and loose nuclear material in the internal cell structure. In addition, the possible molecular mechanisms of citral-induced formation and recovery of V. vulnificus in the VBNC state were explored by transcriptomics. Transcriptome analyses revealed that 1118 genes were significantly altered upon entry into the VBNC state, and 1052 genes were changed after resuscitation. Most of the physiological activities related to energy production were inhibited in the citral-induced VBNC state of V. vulnificus; however, the bacteria retained its pathogenicity. The citral-induced resuscitation of V. vulnificus in the VBNC state selectively restored the activity of some genes related to bacterial growth and reproduction. Meanwhile, the expression levels of other genes may have been influenced by citral-induced resuscitation after the formation of the VBNC state. In conclusion, this study evaluated and analyzed the ability and possible mechanism of citral on the formation of VBNC state and the recovery of VBNC state of V. vulnificus, and made a comprehensive assessment for the safety of citral application in food production.

Meiotic Cytokinesis in Saccharomyces cerevisiae: Spores That Just Need Closure.

In the budding yeast Saccharomyces cerevisiae, sporulation occurs during starvation of a diploid cell and results in the formation of four haploid spores forming within the mother cell ascus. Meiosis divides the genetic material that is encapsulated by the prospore membrane that grows to surround the haploid nuclei; this membrane will eventually become the plasma membrane of the haploid spore. Cellularization of the spores occurs when the prospore membrane closes to capture the haploid nucleus along with some cytoplasmic material from the mother cell, and thus, closure of the prospore membrane is the meiotic cytokinetic event. This cytokinetic event involves the removal of the leading-edge protein complex, a complex of proteins that localizes to the leading edge of the growing prospore membrane. The development and closure of the prospore membrane must be coordinated with other meiotic exit events such as spindle disassembly. Timing of the closure of the prospore membrane depends on the meiotic exit pathway, which utilizes Cdc15, a Hippo-like kinase, and Sps1, an STE20 family GCKIII kinase, acting in parallel to the E3 ligase Ama1-APC/C. This review describes the sporulation process and focuses on the development of the prospore membrane and the regulation of prospore membrane closure.

The mosquito effect: regulatory and effector T cells acquire cytoplasmic material from tumor cells through intercellular transfer.

The phenomenon of intercellular transfer of cellular material, including membranes, cytoplasm, and even organelles, has been observed for decades. The functional impact and molecular mechanisms of such transfer in the immune system remain largely elusive due to the absence of a robust in vivo model. Here, we introduce a new tumor mouse model, where tumor cells express the soluble ultra-bright fluorescent protein ZsGreen, which allows detection and measurement of intercellular transfer of cytoplasm from tumor cells to infiltrating immune cells. We found that in addition to various types of myeloid lineage cells, a large fraction of T regulatory cells and effector CD8 T cells acquire tumor material. Based on the distribution of tumor-derived ZsGreen, the majority of T cells integrate captured cytoplasm into their own, while most myeloid cells store tumor material in granules. Furthermore, scRNA-seq analysis revealed significant alterations in transcriptomes of T cells that acquired tumor cell cytoplasm, suggesting potential impact on T cell function. We identified that the participation of T cells in intercellular transfer requires cell-cell contact and is strictly dependent on the activation status of T lymphocytes. Finally, we propose to name the described phenomenon of intercellular transfer for tumor infiltrating T cells the "mosquito effect".