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Journal of Virology Sep 2023Nascent nucleocapsids of herpesviruses acquire a primary envelope during their nuclear export by budding through the inner nuclear membrane into the perinuclear space...
Nascent nucleocapsids of herpesviruses acquire a primary envelope during their nuclear export by budding through the inner nuclear membrane into the perinuclear space between the inner and outer nuclear membranes. This process is mediated by a conserved viral heterodimeric complex designated the nuclear egress complex, which consists of the nuclear matrix protein and the nuclear membrane protein. In addition to its essential roles during nuclear egress, the nuclear matrix protein has been shown to interact with intracellular signaling pathway molecules including NF-κB and IFN-β to affect viral or cellular gene expression. The human herpesvirus 6A (HHV-6A) U37 gene encodes a nuclear matrix protein, the role of which has not been analyzed. Here, we show that HHV-6A U37 activates the heat shock element promoter and induces the accumulation of the molecular chaperone Hsp90. Mechanistically, HHV-6A U37 interacts with heat shock transcription factor 1 (HSF1) and induces its phosphorylation at Ser-326. We report that pharmacological inhibition of HSF1, Hsp70, or Hsp90 decreases viral protein accumulation and viral replication. Taken together, our results lead us to propose a model in which HHV-6A U37 activates the heat shock response to support viral gene expression and replication. IMPORTANCE Human herpesvirus 6A (HHV-6A) is a dsDNA virus belonging to the genus within the subfamily. It is frequently found in patients with neuroinflammatory disease, although its pathogenetic role, if any, awaits elucidation. The heat shock response is important for cell survival under stressful conditions that disrupt homeostasis. Our results indicate that HHV-6A U37 activates the heat shock element promoter and leads to the accumulation of heat shock proteins. Next, we show that the heat shock response is important for viral replication. Overall, our findings provide new insights into the function of HHV-6A U37 in host cell signaling and identify potential cellular targets involved in HHV-6A pathogenesis and replication.
Topics: Humans; Heat Shock Transcription Factors; Heat-Shock Response; Herpesvirus 6, Human; Viral Matrix Proteins; HSP90 Heat-Shock Proteins; HSP70 Heat-Shock Proteins; Promoter Regions, Genetic; Virus Replication; Phosphorylation; Gene Expression Regulation, Viral; Signal Transduction
PubMed: 37671864
DOI: 10.1128/jvi.00718-23 -
Frontiers in Cellular and Infection... 2023Ring finger protein 213 (RNF213) is a large E3 ubiquitin ligase with a molecular weight of 591 kDa that is associated with moyamoya disease, a rare cerebrovascular... (Review)
Review
Ring finger protein 213 (RNF213) is a large E3 ubiquitin ligase with a molecular weight of 591 kDa that is associated with moyamoya disease, a rare cerebrovascular disease. It is located in the cytosol and perinuclear space. Missense mutations in this gene have been found to be more prevalent in patients with moyamoya disease compared with that in healthy individuals. Understanding the molecular function of RNF213 could provide insights into moyamoya disease. RNF213 contains a C3HC4-type RING finger domain with an E3 ubiquitin ligase domain and six AAA+ adenosine triphosphatase (ATPase) domains. It is the only known protein with both AAA+ ATPase and ubiquitin ligase activities. Recent studies have highlighted the role of RNF213 in fighting against microbial infections, including viruses, parasites, bacteria, and chlamydiae. This review aims to summarize the recent research progress on the mechanisms of RNF213 in pathogenic infections, which will aid researchers in understanding the antimicrobial role of RNF213.
Topics: Humans; Moyamoya Disease; Ubiquitin-Protein Ligases; Genes, Regulator; Transcription Factors; Anti-Infective Agents; Adenosine Triphosphatases
PubMed: 37655297
DOI: 10.3389/fcimb.2023.1205355 -
BioRxiv : the Preprint Server For... Jul 2023TorsinA is an atypical ATPase that lacks intrinsic activity unless it is bound to its activators lamina-associated polypeptide 1 (LAP1) in the perinuclear space or...
TorsinA is an atypical ATPase that lacks intrinsic activity unless it is bound to its activators lamina-associated polypeptide 1 (LAP1) in the perinuclear space or luminal domain-like LAP1 (LULL1) throughout the endoplasmic reticulum. However, the interaction of torsinA with LAP1 and LULL1 has not yet been shown to modulate a defined physiological process in mammals . We previously demonstrated that depletion of torsinA from mouse hepatocytes leads to reduced liver triglyceride secretion and marked steatosis, whereas depletion of LAP1 had more modest similar effects. We now show that depletion of LULL1 alone does not significantly decrease liver triglyceride secretion or cause steatosis. However, simultaneous depletion of both LAP1 and LULL1 from hepatocytes leads to defective triglyceride secretion and marked steatosis similar to that observed with depletion of torsinA. Our results demonstrate that torsinA and its activators dynamically regulate a physiological process in mammals .
PubMed: 37547008
DOI: 10.1101/2023.06.21.545957 -
Nature Plants Jul 2023The nuclear lamina is a complex network of nuclear lamins and lamin-associated nuclear membrane proteins, which scaffold the nucleus to maintain structural integrity. In...
The nuclear lamina is a complex network of nuclear lamins and lamin-associated nuclear membrane proteins, which scaffold the nucleus to maintain structural integrity. In Arabidopsis thaliana, nuclear matrix constituent proteins (NMCPs) are essential components of the nuclear lamina and are required to maintain the structural integrity of the nucleus and specific perinuclear chromatin anchoring. At the nuclear periphery, suppressed chromatin overlapping with repetitive sequences and inactive protein-coding genes are enriched. At a chromosomal level, plant chromatin organization in interphase nuclei is flexible and responds to various developmental cues and environmental stimuli. On the basis of these observations in Arabidopsis, and given the role of NMCP genes (CRWN1 and CRWN4) in organizing chromatin positioning at the nuclear periphery, one can expect considerable changes in chromatin-nuclear lamina interactions when the global chromatin organization patterns are being altered in plants. Here we report the highly flexible nature of the plant nuclear lamina, which disassembles substantially under various stress conditions. Focusing on heat stress, we reveal that chromatin domains, initially tethered to the nuclear envelope, remain largely associated with CRWN1 and become scattered in the inner nuclear space. By investigating the three-dimensional chromatin contact network, we further reveal that CRWN1 proteins play a structural role in shaping the changes in genome folding under heat stress. Also, CRWN1 acts as a negative transcriptional coregulator to modulate the shift of the plant transcriptome profile in response to heat stress.
Topics: Nuclear Lamina; Cell Nucleus; Chromatin; Nuclear Envelope; Lamins; Arabidopsis
PubMed: 37400513
DOI: 10.1038/s41477-023-01457-2 -
Pharmaceutics Jun 2023Fluorescent micellar carriers with controlled release of a novel anticancer drug were developed to enable intracellular imaging and cancer treatment simultaneously. The...
Novel Fluorescent Benzimidazole-Hydrazone-Loaded Micellar Carriers for Controlled Release: Impact on Cell Toxicity, Nuclear and Microtubule Alterations in Breast Cancer Cells.
Fluorescent micellar carriers with controlled release of a novel anticancer drug were developed to enable intracellular imaging and cancer treatment simultaneously. The nanosized fluorescent micellar systems were embedded with a novel anticancer drug via the self-assembling behavior of well-defined block copolymers based on amphiphilic poly(acrylic acid)-block-poly(n-butyl acrylate) (PAA-b-PnBA) copolymer obtained by Atom Transfer Radical Polymerization (ATRP) and hydrophobic anticancer benzimidazole-hydrazone drug (BzH). Through this method, well-defined nanosized fluorescent micelles were obtained consisting of a hydrophilic PAA shell and a hydrophobic PnBA core embedded with the BzH drug due to the hydrophobic interactions, thus reaching very high encapsulation efficiency. The size, morphology, and fluorescent properties of blank and drug-loaded micelles were investigated using dynamic light scattering (DLS), transmission electron microscopy (TEM), and fluorescent spectroscopy, respectively. Additionally, after 72 h of incubation, drug-loaded micelles released 3.25 μM of BzH, which was spectrophotometrically determined. The BzH drug-loaded micelles were found to exhibit enhanced antiproliferative and cytotoxic effects on MDA-MB-231 cells, with long-lasting effects on microtubule organization, with apoptotic alterations and preferential localization in the perinuclear space of cancer cells. In contrast, the antitumor effect of BzH alone or incorporated in micelles on non-cancerous cells MCF-10A was relatively weak.
PubMed: 37376201
DOI: 10.3390/pharmaceutics15061753 -
Diseases (Basel, Switzerland) Jun 2023Oxymetholone is one of the anabolic steroids that has widely been used among teenagers and athletes to increase their muscle bulk. It has undesirable effects on male...
Oxymetholone is one of the anabolic steroids that has widely been used among teenagers and athletes to increase their muscle bulk. It has undesirable effects on male health and fertility. In this study, the therapeutic effects of platelet-rich plasma (PRP) on oxymetholone-induced testicular toxicity were investigated in adult albino rats. During the experiments, 49 adult male albino rats were divided into 4 main groups: Group 0 (donor group) included 10 rats for the donation of PRP, Group I (control group) included 15 rats, Group II included 8 rats that received 10 mg/kg of oxymetholone orally, once daily, for 30 days, and Group III included 16 rats and was subdivided into 2 subgroups (IIIa and IIIb) that received oxymetholone the same as group II and then received PRP once and twice, respectively. Testicular tissues of all examined rats were obtained for processing and histological examination and sperm smears were stained and examined for sperm morphology. Oxymetholone-treated rats revealed wide spaces in between the tubules, vacuolated cytoplasm, and dark pyknotic nuclei of most cells, as well as deposition of homogenous acidophilic material between the tubules. Electron microscopic examination showed vacuolated cytoplasm of most cells, swollen mitochondria, and perinuclear dilatation. Concerning subgroup IIIa (PRP once), there was a partial improvement in the form of decreased vacuolations and regeneration of spermatogenic cells, as well as a reasonable improvement in sperm morphology. Regarding subgroup IIIb (PRP twice), histological sections revealed restoration of the normal testicular structure to a great extent, regeneration of the spermatogenic cells, and most sperms had normal morphology. Thus, it is recommended to use PRP to minimize structural changes in the testis of adult albino rats caused by oxymetholone.
PubMed: 37366872
DOI: 10.3390/diseases11020084 -
Nature Communications Jun 2023The endoplasmic reticulum (ER) is an organelle of nucleated cells that produces proteins, lipids and oligosaccharides. ER volume and activity are increased upon...
The endoplasmic reticulum (ER) is an organelle of nucleated cells that produces proteins, lipids and oligosaccharides. ER volume and activity are increased upon induction of unfolded protein responses (UPR) and are reduced upon activation of ER-phagy programs. A specialized domain of the ER, the nuclear envelope (NE), protects the cell genome with two juxtaposed lipid bilayers, the inner and outer nuclear membranes (INM and ONM) separated by the perinuclear space (PNS). Here we report that expansion of the mammalian ER upon homeostatic perturbations results in TMX4 reductase-driven disassembly of the LINC complexes connecting INM and ONM and in ONM swelling. The physiologic distance between ONM and INM is restored, upon resolution of the ER stress, by asymmetric autophagy of the NE, which involves the LC3 lipidation machinery, the autophagy receptor SEC62 and the direct capture of ONM-derived vesicles by degradative LAMP1/RAB7-positive endolysosomes in a catabolic pathway mechanistically defined as micro-ONM-phagy.
Topics: Animals; Nuclear Envelope; Endoplasmic Reticulum Stress; Autophagy; Unfolded Protein Response; Endoplasmic Reticulum; Mammals
PubMed: 37311770
DOI: 10.1038/s41467-023-39172-3 -
Mass Spectrometry (Tokyo, Japan) 2023Aberrant glycosylation of membrane proteins is a hallmark of cancer and a useful molecular marker for the diagnosis of breast cancer (BC). However, the molecular...
Aberrant glycosylation of membrane proteins is a hallmark of cancer and a useful molecular marker for the diagnosis of breast cancer (BC). However, the molecular mechanisms by which altered glycosylation affects the malignant transformations associated with BC are poorly understood. Accordingly, we performed comparative membrane -glycoproteomics using the human BC cell line pair, Hs578T, and its syngeneic normal cell line, Hs578Bst. A total of 359 -glycoforms derived from 113 proteins were identified in both cell lines, of which 27 were found only in Hs578T cells. Significant changes in -glycosylation were found in the lysosome-associated membrane protein 1 (LAMP1), the integrin family, and laminin. Confocal immunofluorescence microscopy images revealed the accumulation of lysosomes in the perinuclear space in cancer cells, which could be associated with marked changes in LAMP1 glycosylation, such as a decreased level of polylactosamine chains. Overall, the alterations in glycosylation may be involved in changes in the adhesion and degradation of BC cells.
PubMed: 37250596
DOI: 10.5702/massspectrometry.A0117 -
Journal of Virology May 2023Adeno-associated viruses (AAVs) are small, helper-dependent, single-stranded DNA viruses that exploit a broad spectrum of host factors for cell entry. During the course...
Adeno-associated viruses (AAVs) are small, helper-dependent, single-stranded DNA viruses that exploit a broad spectrum of host factors for cell entry. During the course of infection, several AAV serotypes have been shown to transit through the -Golgi network within the host cell. In the current study, we investigated whether the Golgi-localized, calcium-dependent protease furin influences AAV transduction. While CRISPR/Cas9-mediated knockout (KO) of the gene minimally affected the transduction efficiency of most recombinant AAV serotypes tested, we observed a striking increase in transgene expression (~2 log orders) for the African green monkey isolate AAV4. Interrogation of different steps in the infectious pathway revealed that AAV4 binding, uptake, and transcript levels are increased in furin KO cells, but postentry steps such as uncoating or nuclear entry remain unaffected. Recombinant furin does not cleave AAV4 capsid proteins nor alter cellular expression levels of essential factors such as AAVR or GPR108. Interestingly, fluorescent lectin screening revealed a marked increase in 2,3--linked sialoglycan staining on the surface and perinuclear space of furin KO cells. The essential nature of increased sialoglycan expression in furin KO cells in enhancing AAV4 transduction was further corroborated by (i) increased transduction by the closely related isolates AAVrh.32.33 and sea lion AAV and (ii) selective blockade or removal of cellular 2,3--linked sialoglycans by specific lectins or neuraminidase, respectively. Based on the overall findings, we postulate that furin likely plays a key role in regulating expression of cellular sialoglycans, which in turn can influence permissivity to AAVs and possibly other viruses. Adeno-associated viruses (AAVs) are a proven recombinant vector platform for gene therapy and have demonstrated success in the clinic. Continuing to improve our knowledge of AAV-host cell interactions is critical for improving the safety and efficacy. The current study dissects the interplay between furin, a common intracellular protease, and certain cell surface sialoglycans that serve as viral attachment factors for cell entry. Based on the findings, we postulate that differential expression of furin in host cells and tissues is likely to influence gene expression by certain recombinant AAV serotypes.
Topics: Animals; Chlorocebus aethiops; Dependovirus; Virus Internalization; Furin; Genetic Vectors; Capsid Proteins; Transduction, Genetic
PubMed: 37097176
DOI: 10.1128/jvi.00093-23 -
Annual Review of Virology Sep 2023Nuclear egress of herpesvirus capsids across the intact nuclear envelope is an exceptional vesicle-mediated nucleocytoplasmic translocation resulting in the delivery of... (Review)
Review
Nuclear egress of herpesvirus capsids across the intact nuclear envelope is an exceptional vesicle-mediated nucleocytoplasmic translocation resulting in the delivery of herpesvirus capsids into the cytosol. Budding of the (nucleo)capsid at and scission from the inner nuclear membrane (INM) is mediated by the viral nuclear egress complex (NEC) resulting in a transiently enveloped virus particle in the perinuclear space followed by fusion of the primary envelope with the outer nuclear membrane (ONM). The dimeric NEC oligomerizes into a honeycomb-shaped coat underlining the INM to induce membrane curvature and scission. Mutational analyses complemented structural data defining functionally important regions. Questions remain, including where and when the NEC is formed and how membrane curvature is mediated, vesicle formation is regulated, and directionality is secured. The composition of the primary enveloped virion and the machinery mediating fusion of the primary envelope with the ONM is still debated. While NEC-mediated budding apparently follows a highly conserved mechanism, species and/or cell type-specific differences complicate understanding of later steps.
Topics: Viral Proteins; Herpesviridae; Nuclear Envelope; Capsid Proteins; Capsid; Cell Nucleus; Virus Release
PubMed: 37040797
DOI: 10.1146/annurev-virology-111821-105518