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Current Issues in Molecular Biology 2021During viral replication, herpesviruses utilize a unique strategy, termed nuclear egress, to translocate capsids from the nucleus into the cytoplasm. This initial... (Review)
Review
During viral replication, herpesviruses utilize a unique strategy, termed nuclear egress, to translocate capsids from the nucleus into the cytoplasm. This initial budding step transfers a newly formed capsid from within the nucleus, too large to fit through nuclear pores, through the inner nuclear membrane to the perinuclear space. The perinuclear enveloped virion must then fuse with the outer nuclear membrane to be released into the cytoplasm for further maturation, undergoing budding once again at the trans-Golgi network or early endosomes, and ultimately exit the cell non-lytically to spread infection. This first budding process is mediated by two conserved viral proteins, UL31 and UL34, that form a heterodimer called the nuclear egress complex (NEC). This review focuses on what we know about how the NEC mediates capsid transport to the perinuclear space, including steps prior to and after this budding event. Additionally, we discuss the involvement of other viral proteins in this process and how NEC-mediated budding may be regulated during infection.
Topics: Capsid; Cell Nucleus; Cytoplasm; Herpesviridae; Herpesviridae Infections; Humans; Nuclear Envelope; Viral Proteins; Virion
PubMed: 32764158
DOI: 10.21775/cimb.041.125 -
Anatomical Record (Hoboken, N.J. : 2007) Apr 2021Sofosbuvir is a promising antiviral drug against chronic hepatitis C virus. Although it is characterized by its high efficacy, its adverse effects on nervous tissue are...
Sofosbuvir is a promising antiviral drug against chronic hepatitis C virus. Although it is characterized by its high efficacy, its adverse effects on nervous tissue are still unclear. Saffron is known for its neuroprotective property. This is a biochemical, histological and immunohistochemical study of the effect of sofosbuvir on the cerebellar cortex of rat and the possible ameliorating role of saffron's aqueous extract. Twenty-four adult male Wistar albino rats were equally divided into four groups; control, saffron extract-treated, sofosbuvir-treated (41.1 mg/kg/day for 6 weeks) and group concomitantly treated with saffron extract and sofosbuvir. Sofosbuvir-treated group recorded a significant increase in cerebellar malondialdehyde level coupling with a significant decrease in tissue glutathione and superoxide dismutase. Light microscopy revealed reduced number of Purkinje cells. The granular layer depicted many granular cells and Bergmann astrocytes with nuclear and cytoplasmic alterations. Electron microscopy revealed disorganized molecular layer with disarranged myelinated axons and disrupted mitochondria. Few shrunken Purkinje cells showed electron-dense cytoplasm and rarefied nuclei, indistinct nuclear envelope and dilated perinuclear space, areas of vacuolated cytoplasm, fragmented rough endoplasmic reticulum and few dark mitochondria. Some axons with tiny mitochondria were detected. A significant upregulation in immunohistochemical expression of GFAP-positive astrocytes was recorded. Concomitant administration of saffron extract significantly improved all studied parameters. Saffron extract is beneficial in ameliorating sofosbuvir-induced cerebellar morphological changes mainly through its antioxidant and neuroprotective properties.
Topics: Animals; Antiviral Agents; Astrocytes; Cerebellar Cortex; Crocus; Glutathione; Male; Malondialdehyde; Plant Extracts; Purkinje Cells; Rats; Rats, Wistar; Sofosbuvir; Superoxide Dismutase
PubMed: 32721089
DOI: 10.1002/ar.24501 -
Virus Research Oct 2020During herpesvirus replication, newly synthesized nucleocapsids exit the nucleus by a vesicle-mediated transport, which requires the nuclear egress complex (NEC),...
During herpesvirus replication, newly synthesized nucleocapsids exit the nucleus by a vesicle-mediated transport, which requires the nuclear egress complex (NEC), composed of the conserved viral proteins designated as pUL31 and pUL34 in the alphaherpesviruses pseudorabies virus (PrV) and herpes simplex viruses. Oligomerization of the heterodimeric NEC at the inner nuclear membrane (INM) results in membrane bending and budding of virus particles into the perinuclear space. The INM-derived primary envelope then fuses with the outer nuclear membrane to release nucleocapsids into the cytoplasm. The two NEC components are necessary and sufficient for induction of vesicle budding and scission as shown after co-expression in eukaryotic cells or in synthetic membranes. However, where and when the NEC is formed, how membrane curvature is mediated and how it is regulated, remains unclear. While monospecific antisera raised against the different components of the PrV NEC aided in the characterization and intracellular localization of the individual proteins, no NEC specific tools have been described yet for any herpesvirus. To gain more insight into vesicle budding and scission, we aimed at generating NEC specific monoclonal antibodies (mAbs). To this end, mice were immunized with bacterially expressed soluble PrV NEC, which was previously used for structure determination. Besides pUL31- and pUL34-specific mAbs, we also identified mAbs, which reacted only in the presence of both proteins indicating specificity for the complex. Confocal microscopy with those NEC-specific mAbs revealed small puncta (approx. 0.064 μm) along the nuclear rim in PrV wild type infected cells. In contrast, ca. 5-fold larger speckles (approx. 0.35 μm) were detectable in cells infected with a PrV mutant lacking the viral protein kinase pUS3, which is known to accumulate primary enveloped virions in the PNS within large invaginations of the INM, or in cells co-expressing pUL31 and pUL34. Kinetic experiments showed that while the individual proteins were detectable already between 2-4 hours after infection, the NEC-specific mAbs produced significant staining only after 4-6 hours in accordance with timing of nuclear egress. Taken together, the data indicate that these mAbs specifically label the PrV NEC.
Topics: Animals; Antibodies, Monoclonal; Cell Line; Female; Herpesvirus 1, Suid; Mice; Mice, Inbred BALB C; Microscopy, Immunoelectron; Nuclear Envelope; Nucleocapsid; Rabbits; Viral Proteins; Virus Release
PubMed: 32682818
DOI: 10.1016/j.virusres.2020.198096 -
The Plant Journal : For Cell and... Oct 2020Soybean cyst nematode (SCN; Heterodera glycines) is the largest pathogenic cause of soybean yield loss. The Rhg1 locus is the most used and best characterized SCN...
Soybean cyst nematode (SCN; Heterodera glycines) is the largest pathogenic cause of soybean yield loss. The Rhg1 locus is the most used and best characterized SCN resistance locus, and contains three genes including one encoding an α-SNAP protein. Although the Rhg1 α-SNAP is known to play an important role in vesicle trafficking and SCN resistance, the protein's binding partners and the molecular mechanisms underpinning SCN resistance remain unclear. In this report, we show that the Rhg1 α-SNAP strongly interacts with two syntaxins of the t-SNARE family (Glyma.12G194800 and Glyma.16G154200) in yeast and plants; importantly, the genes encoding these syntaxins co-localize with SCN resistance quantitative trait loci. Fluorescent visualization revealed that the α-SNAP and the two interacting syntaxins localize to the plasma membrane and perinuclear space in both tobacco epidermal and soybean root cells. The two syntaxins and their two homeologs were mutated, individually and in combination, using the CRISPR-Cas9 system in the SCN-resistant Peking and SCN-susceptible Essex soybean lines. Peking roots with deletions introduced into syntaxin genes exhibited significantly reduced resistance to SCN, confirming that t-SNAREs are critical to resisting SCN infection. The results presented here uncover a key step in the molecular mechanism of SCN resistance, and will be invaluable to soybean breeders aiming to develop highly SCN-resistant soybean varieties.
Topics: Animals; Clustered Regularly Interspaced Short Palindromic Repeats; Disease Resistance; Host-Parasite Interactions; Plant Diseases; Plant Proteins; Plant Roots; Plants, Genetically Modified; Qa-SNARE Proteins; Quantitative Trait Loci; SNARE Proteins; Glycine max; Two-Hybrid System Techniques; Tylenchoidea
PubMed: 32645235
DOI: 10.1111/tpj.14923 -
Journal of Experimental Botany Aug 2020The nuclear envelope delineates the eukaryotic cell nucleus. The membrane system of the nuclear envelope consists of an outer nuclear membrane and an inner nuclear... (Review)
Review
The nuclear envelope delineates the eukaryotic cell nucleus. The membrane system of the nuclear envelope consists of an outer nuclear membrane and an inner nuclear membrane separated by a perinuclear space. It serves as more than just a static barrier, since it regulates the communication between the nucleoplasm and the cytoplasm and provides the anchoring points where chromatin is attached. Fewer nuclear envelope proteins have been identified in plants in comparison with animals and yeasts. Here, we review the current state of knowledge of the nuclear envelope in plants, focusing on its role as a chromatin organizer and regulator of gene expression, as well as on the modifications that it undergoes to be efficiently disassembled and reassembled with each cell division. Advances in knowledge concerning the mitotic role of some nuclear envelope constituents are also presented. In addition, we summarize recent progress on the contribution of the nuclear envelope elements to telomere tethering and chromosome dynamics during the meiotic division in different plant species.
Topics: Animals; Cell Division; Chromatin; Cytoplasm; Nuclear Envelope; Telomere
PubMed: 32589712
DOI: 10.1093/jxb/eraa299 -
Biochemical and Biophysical Research... Jun 2020Apoptosis is the prototype for a regulated form of cell death, but recent studies have revealed other types of regulated forms of cell death, including necroptosis and...
Apoptosis is the prototype for a regulated form of cell death, but recent studies have revealed other types of regulated forms of cell death, including necroptosis and ferroptosis. The molecular mechanisms underlying the execution of these processes have been intensively investigated, yet the hallmarks of their morphology are not fully understood. Here, we report that electron lucent cytoplasm was a common feature of both necroptosis and ferroptosis, which was consistent with cytoplasmic vacuolization due to a defect in the cytoplasmic membrane integrity. Notably, the perinuclear space was dilated in necroptosis, but such dilation did not occur in ferroptosis. Cells undergoing ferroptosis, but not necroptosis, exhibited an electron lucent nucleus. We previously reported that one of the nuclear danger-associated molecular patterns (DAMPs), high mobility group box (HMGB)1, is rapidly released from the nucleus to the extracellular spaces of cells undergoing necroptosis through the ruptured nuclear and cytoplasmic membrane. Via time-lapse imaging of cells stably expressing HMGB1 fused to a fluorescence protein, we found that HMGB1 was also released from the nucleus to the cytosol, and then eventually released into the extracellular spaces in cells undergoing ferroptosis. Thus, nuclear membrane damage was induced prior to cytoplasmic membrane rupture in ferroptosis. Thus, dilation of the perinuclear space and an electron lucent nucleus may be the hallmarks of necroptosis and ferroptosis, respectively.
Topics: Cell Line, Tumor; Cell Membrane; Cell Nucleus; Cytoplasm; Ferroptosis; HMGB1 Protein; Humans; Microscopy, Electron, Transmission; Necroptosis
PubMed: 32430176
DOI: 10.1016/j.bbrc.2020.04.127 -
The Journal of Clinical Investigation Jun 2020Although autophagy is generally protective, uncontrolled or excessive activation of autophagy can be detrimental. However, it is often difficult to distinguish death by...
Although autophagy is generally protective, uncontrolled or excessive activation of autophagy can be detrimental. However, it is often difficult to distinguish death by autophagy from death with autophagy, and whether autophagy contributes to death in cardiomyocytes (CMs) is still controversial. Excessive activation of autophagy induces a morphologically and biochemically defined form of cell death termed autosis. Whether autosis is involved in tissue injury induced under pathologically relevant conditions is poorly understood. In the present study, myocardial ischemia/reperfusion (I/R) induced autosis in CMs, as evidenced by cell death with numerous vacuoles and perinuclear spaces, and depleted intracellular membranes. Autosis was observed frequently after 6 hours of reperfusion, accompanied by upregulation of Rubicon, attenuation of autophagic flux, and marked accumulation of autophagosomes. Genetic downregulation of Rubicon inhibited autosis and reduced I/R injury, whereas stimulation of autosis during the late phase of I/R with Tat-Beclin 1 exacerbated injury. Suppression of autosis by ouabain, a cardiac glycoside, in humanized Na+,K+-ATPase-knockin mice reduced I/R injury. Taken together, these results demonstrate that autosis is significantly involved in I/R injury in the heart and triggered by dysregulated accumulation of autophagosomes due to upregulation of Rubicon.
Topics: Animals; Autophagosomes; Autophagy; Intracellular Signaling Peptides and Proteins; Mice; Mice, Transgenic; Myocardial Reperfusion Injury; Myocardium; Up-Regulation
PubMed: 32364533
DOI: 10.1172/JCI132366 -
Scientific Reports Apr 2020The duck plague virus (DPV) US3 protein, a homolog of the herpes simplex virus-1 (HSV-1) US3 protein that is reported to be critical for viral replication, has been...
The duck plague virus (DPV) US3 protein, a homolog of the herpes simplex virus-1 (HSV-1) US3 protein that is reported to be critical for viral replication, has been minimally studied. Therefore, to investigate the function of the DPV US3 protein, we used scarless Red recombination technology based on an infectious bacterial artificial chromosome (BAC) containing the DPV Chinese virulent strain (CHv) genome and successfully constructed and rescued a US3-deleted mutant and the corresponding revertant virus (BAC-CHv-ΔUS3 and BAC-CHv-ΔUS3R, respectively). For viral growth characteristics, compared to the parental and revertant viruses, the US3-deleted mutant showed an approximately 100-fold reduction in viral titers but no significant reduction in genome copies, indicating that the US3-deleted mutant exhibited decreased viral replication but not decreased viral DNA generation. In addition, the US3-deleted mutant formed viral plaques that were 33% smaller on average than those formed by the parental and revertant viruses, demonstrating that US3 protein affected the viral cell-to-cell spread of DPV. Finally, the results of electron microscopy showed that the deletion of US3 resulted in a large number of virions accumulating in the nucleus and perinuclear space, thus blocking virion nuclear egress. In this study, we found that the DPV US3 protein played pivotal roles in viral replication by promoting viral cell-to-cell spread and virion nuclear egress, which may provide some references for research on the function of the DPV US3 protein.
Topics: Animals; Cells, Cultured; Ducks; Herpesviridae Infections; Mardivirus; Poultry Diseases; Viral Proteins; Virion; Virus Release
PubMed: 32346128
DOI: 10.1038/s41598-020-64190-2 -
Zhen Ci Yan Jiu = Acupuncture Research Apr 2020To investigate the effect of acupuncture stimulation of head acupoints "Jin San Zhen" (JIN's Three Acupuncture Needles Therapy) on behavior reactions, hippocampal...
OBJECTIVE
To investigate the effect of acupuncture stimulation of head acupoints "Jin San Zhen" (JIN's Three Acupuncture Needles Therapy) on behavior reactions, hippocampal neuronal autophagy and expression of autophagy associated proteins (Beclin-1 and light chain 3 Ⅱ/Ⅰ [LC 3 Ⅱ/Ⅰ]) in rats with hypoxic-ischemic brain damage (HIBD) due to fetal intrauterine distress, so as to reveal its underlying mechanisms in improving neonatal HIBD.
METHODS
Pregnant SD rats were used in the present study. The HIBD model was established by delayed caesarean delivery and bilateral uterine arteries clipping for 10 minutes. The HIBD rats were randomly divided into model group and acupuncture groups (=9 rats in each group). The other 9 rats delivered naturally were used as the normal control group. On day 14 after delivery, the neonatal rats in the acupuncture group received acupuncture stimulation of head acupoints ("Nao San Zhen""Nie San Zhen" and "Zhi San Zhen") by twirling each needle leftward and rightward for 10 times, once a day for 14 d. The open field test and Morris water maze test were used to determine the locomotive activity and spatial learning-memory ability, respectively. The ultrastructure and autophagosomes in the hippocampal neurons were observed by transmission electron microscope. The contents and expression levels of Beclin-1 and LC3 Ⅱ/Ⅰ in the hippocampus tissues were detected by flow cytometry and Western blot, separately.
RESULTS
Compared with the normal control group, the time to go out of the central region of open field test, and the escape latency and duration of first platform-quadrant-crossing of spatial exploration of Morris water maze tests were significantly increased (<0.01,<0.05,<0.001), and the total distance and number of activities in the central region, and the target quadrant resistance time and number of platform-cros-sing remarkably decreased in the model group (<0.01, <0.05), suggesting a decline of both locomotor activity and learning-memory ability after modeling. The expression level (%) of Beclin-1 protein and ratio of LC3 Ⅱ/Ⅰ proteins were considerably increased in the model group (<0.01). Following acupuncture interventions, the locomotor activity and spatial learning-memory ability were obviously increased (<0.05,<0.01,<0.001), and the expression of Beclin-1 protein and ratio of LC3 Ⅱ/Ⅰ were further up-regulated relevant to the model group (<0.001). Moreover, ultrastructural observation showed serrated change of nuclear membrane and widened perinuclear space, vacuolization in the mitochondria, dilation of endoplasmic reticulum and increase of autophagosomes in the hippocampal neurons in the model group. These situations were relatively milder in the acupuncture group.
CONCLUSION
Acupuncture of head acupoints of "JIN San Zhen" may increase locomotor activity and learning-memory abi-lity in rats with HIBD due to fetal intrauterine anoxia, which is closely with its effect in promoting hippocampal neuronal autophagy via up-regulating the expression of Beclin-1 and LC3 Ⅱ/Ⅰ.
Topics: Acupuncture Therapy; Animals; Autophagy; Female; Hippocampus; Locomotion; Pregnancy; Rats; Rats, Sprague-Dawley
PubMed: 32333531
DOI: 10.13702/j.1000-0607.190704 -
Toxicology and Industrial Health Feb 2020In this study, the effects of a potent antioxidant, selenium, on apoptosis induced by acrolein, a cytotoxic and genotoxic environmental pollutant, were investigated by... (Comparative Study)
Comparative Study
In this study, the effects of a potent antioxidant, selenium, on apoptosis induced by acrolein, a cytotoxic and genotoxic environmental pollutant, were investigated by immunohistochemical and electron microscopic methods. One hundred adult male Wistar albino rats were used in the study. The rats were divided into four main groups: control, acrolein, selenium, and acrolein + selenium. The animals in the experimental groups were given 1 mg/kg/day selenium and 4 mg/kg/day acrolein daily for 7 days by gavage. After drug administration, each group was divided into subgroups according to the time they were to be euthanized: 12th hour, 1st, 2nd, 3rd, and 5th day. The rats in each group at the determined time were euthanized and their livers were removed. Routine histological procedures were performed for light and electron microscopy examinations. After applying the Terminal Deoxynucleotidyl Transferase dUTP nick end labeling assay on the liver sections, apoptotic index values were calculated. Comparing the liver sections of the rats in the acrolein group and the control group, acrolein was found to cause a significant increase in the apoptotic index. The apoptotic index values of the acrolein + selenium group decreased compared to the acrolein group. In the electron microscopic examinations, apoptotic findings were observed in the liver tissues of the rats given acrolein, such as chromatin condensation in the nucleus of hepatocytes, dilatations in the perinuclear space, and cytoplasmic vacuolization. These apoptotic findings were not observed in the acrolein + selenium group after the 12th hour. These findings show that selenium may potentially be useful as a protective agent for people exposed to acrolein.
Topics: Acrolein; Animals; Antioxidants; Apoptosis; Euthanasia, Animal; Liver; Male; Rats; Rats, Wistar; Selenium
PubMed: 32279646
DOI: 10.1177/0748233720909043