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The Journal of Neuroscience : the... Apr 2020During differentiation, oligodendrocyte precursor cells (OPCs) extend a network of processes that make contact with axons and initiate myelination. Recent studies...
During differentiation, oligodendrocyte precursor cells (OPCs) extend a network of processes that make contact with axons and initiate myelination. Recent studies revealed that actin polymerization is required for initiation of myelination whereas actin depolymerization promotes myelin wrapping. Here, we used primary OPCs in culture isolated from neonatal rat cortices of both sexes and young male and female mice with oligodendrocyte-specific deletion of mechanistic target of rapamycin (mTOR) to demonstrate that mTOR regulates expression of specific cytoskeletal targets and actin reorganization in oligodendrocytes during developmental myelination. Loss or inhibition of mTOR reduced expression of profilin2 and ARPC3, actin polymerizing factors, and elevated levels of active cofilin, which mediates actin depolymerization. The deficits in actin polymerization were revealed in reduced phalloidin and deficits in oligodendrocyte cellular branching complexity at the peak of morphologic differentiation and a delay in initiation of myelination. We further show a critical role for mTOR in expression and localization of myelin basic protein () mRNA and MBP protein to the cellular processes where it is necessary at the myelin membrane for axon wrapping. mRNA transport deficits were confirmed by single molecule RNA FISH. Moreover, expression of the kinesin family member 1B, an mRNA transport protein, was reduced in CC1+ cells in the and in mTOR inhibited oligodendrocytes undergoing differentiation These data support the conclusion that mTOR regulates both initiation of myelination and axon wrapping by targeting cytoskeletal reorganization and MBP localization to oligodendrocyte processes. Myelination is essential for normal CNS development and adult axon preservation and function. The mechanistic target of rapamycin (mTOR) signaling pathway has been implicated in promoting CNS myelination; however, there is a gap in our understanding of the mechanisms by which mTOR promotes developmental myelination through regulating specific downstream targets. Here, we present evidence that mTOR promotes the initiation of myelination through regulating specific cytoskeletal targets and cellular process expansion by oligodendrocyte precursor cells as well as expression and cellular localization of myelin basic protein.
Topics: Actin-Related Protein 2-3 Complex; Actins; Animals; Axons; Cell Differentiation; Cytoskeleton; Kinesins; Mice; Mice, Knockout; Myelin Basic Protein; Myelin Proteolipid Protein; Myelin Sheath; Oligodendroglia; Rats; Rats, Sprague-Dawley; Stem Cells; TOR Serine-Threonine Kinases; Zebrafish
PubMed: 32139584
DOI: 10.1523/JNEUROSCI.1434-18.2020 -
Cellular and Molecular Life Sciences :... Apr 2022Proteolipids are proteins with unusual lipid-like properties. It has long been established that PLP and plasmolipin, which are two unrelated membrane-tetra-spanning...
Proteolipids are proteins with unusual lipid-like properties. It has long been established that PLP and plasmolipin, which are two unrelated membrane-tetra-spanning myelin proteolipids, can be converted in vitro into a water-soluble form with a distinct conformation, raising the question of whether these, or other similar proteolipids, can adopt two different conformations in the cell to adapt their structure to distinct environments. Here, we show that MALL, another proteolipid with a membrane-tetra-spanning structure, distributes in membranes outside the nucleus and, within the nucleus, in membrane-less, liquid-like PML body biomolecular condensates. Detection of MALL in one or other environment was strictly dependent on the method of cell fixation used, suggesting that MALL adopts different conformations depending on its physical environment -lipidic or aqueous- in the cell. The acquisition of the condensate-compatible conformation requires PML expression. Excess MALL perturbed the distribution of the inner nuclear membrane proteins emerin and LAP2β, and that of the DNA-binding protein BAF, leading to the formation of aberrant nuclei. This effect, which is consistent with studies identifying overexpressed MALL as an unfavorable prognostic factor in cancer, could contribute to cell malignancy. Our study establishes a link between proteolipids, membranes and biomolecular condensates, with potential biomedical implications.
Topics: Biomolecular Condensates; Cell Nucleus; Humans; Molecular Conformation; Neoplasms; Proteolipids
PubMed: 35399121
DOI: 10.1007/s00018-022-04270-w -
Reproductive Biology Mar 2022Circular RNAs (circRNAs) have been identified as critical regulators in human cancers, including cervical cancer (CC). However, the precise action of circ_0084904 in...
Circular RNAs (circRNAs) have been identified as critical regulators in human cancers, including cervical cancer (CC). However, the precise action of circ_0084904 in cervical carcinogenesis remains to be elucidated. The levels of circ_0084904, microRNA (miR)-802, and Mal, T cell differentiation protein 2 (MAL2) were checked by quantitative real-time PCR (qRT-PCR) or western blot. Ribonuclease R (RNase R) and subcellular localization assays were used to detect the stability and localization of circ_0084904, respectively. Cell colony formation ability was assessed by colony formation assay. Cell cycle and apoptosis were detected by flow cytometry. Cell migration and invasion abilities were gauged by transwell assay. Dual-luciferase reporter and RNA immunoprecipitation (RIP) assays were applied to determine the direct relationship between miR-802 and circ_0084904 or MAL2. The xenograft experiments were performed to evaluate the role of circ_0084904 in tumor growth in vivo. Circ_0084904 was markedly up-regulated in CC tissues and cell lines. Silencing endogenous circ_0084904 impeded cell colony formation, cell cycle progression, migration, invasion, epithelial-mesenchymal transition (EMT), and promoted apoptosis in vitro, as well as diminished tumor growth in vivo. Mechanistically, circ_0084904 targeted miR-802, and the effects of circ_0084904 silencing were mediated by miR-802. MAL2 was directly targeted and inhibited by miR-802, and MAL2 was a functional target of miR-802. Moreover, circ_0084904 modulated MAL2 expression via miR-802. Our study identified circ_0084904 as a novel oncogenic driver in CC depending on the modulation of the miR-802/MAL2 axis, establishing the notion that silencing of circ_0084904 might represent a promising targeted therapy for CC.
Topics: Animals; Cell Line, Tumor; Cell Proliferation; Female; Humans; MicroRNAs; Myelin and Lymphocyte-Associated Proteolipid Proteins; RNA, Circular; Uterine Cervical Neoplasms
PubMed: 35033901
DOI: 10.1016/j.repbio.2021.100600 -
Cellular and Molecular Biology... Oct 2023Soluble epoxide hydrolase (sEH) inhibition has currently emerged as a therapeutic target in the treatment of various neuroinflammatory neurodegenerative diseases,...
Pro-inflammatory GPR75 and anti-apoptotic phospholipase signaling pathways contribute to the ameliorating effect of soluble epoxide hydrolase inhibition on chronic experimental autoimmune encephalomyelitis in mice.
Soluble epoxide hydrolase (sEH) inhibition has currently emerged as a therapeutic target in the treatment of various neuroinflammatory neurodegenerative diseases, including multiple sclerosis. Previously, we reported that treatment of mice with a sEH-selective inhibitor, 1-(1-propanoylpiperidin-4-yl)-3-[4-(trifluoromethoxy)phenyl]urea; TPPU), ameliorated chronic experimental autoimmune encephalomyelitis (EAE) induced by myelin oligodendrocyte glycoprotein 35-55 peptide immunization followed by injection of pertussis toxin to mice via regulating pro-inflammatory and anti-inflammatory pathways in the central nervous system. This study tested the hypothesis that the pro-inflammatory G protein-coupled receptor (GPR) 75 and anti-apoptotic phospholipase C (PLC) signaling pathways also contribute to the ameliorating effect of TPPU on chronic EAE. Brains and spinal cords of phosphate-buffered saline-, dimethyl sulfoxide-, or TPPU (3 mg/kg)-treated mice were used for the measurement of sEH, GPR75, Gaq/11, activator protein (AP)-1, PLC β4, phosphoinositide 3-kinase (PI3K) p85a, Akt1, mitogen-activated protein kinase kinase (MEK) 1/2, extracellular signal-regulated kinase (ERK) 1/2, cyclic adenosine monophosphate-response element-binding protein (CREB) 1, B-cell lymphoma (Bcl)-2, semaphorin (SEMA) 3A, and myelin proteolipid protein (PLP) expression and/or activity by using the immunoblotting method. Expression of sEH, GPR75, Gaq/11, c-jun, phosphorylated c-Jun, and SEMA3A was lower, while PLCβ4, phosphorylated PI3K p85a, phosphorylated Akt1, phosphorylated MEK1/2, phosphorylated ERK1/2, phosphorylated CREB1, Bcl-2, and myelin PLP expression was higher in the tissues of TPPU (3 mg/kg)-treated mice as compared with the EAE and vehicle control groups. Inhibition of sEH by TPPU ameliorates chronic EAE through suppressing pro-inflammatory GPR75/Gaq/11/AP-1 pathway and reducing expression of the remyelination inhibitor, SEMA3A, as well as increasing anti-apoptotic PLC/PI3K/Akt1/MEK1/2/ERK1/2/CREB1/Bcl-2 pathway activity and myelin PLP expression.
Topics: Animals; Mice; Encephalomyelitis, Autoimmune, Experimental; Epoxide Hydrolases; Mice, Inbred C57BL; Myelin Proteolipid Protein; Phosphatidylinositol 3-Kinases; Phospholipases; Proto-Oncogene Proteins c-bcl-2; Semaphorin-3A; Signal Transduction; Receptors, G-Protein-Coupled
PubMed: 37953590
DOI: 10.14715/cmb/2023.69.10.2 -
Food Chemistry Feb 2024Naringenin (NG) belongs to the class of flavanones having impressive pharmacological properties. Unfortunately, the in vivo bioavailability of NG is very low due to its...
Naringenin (NG) belongs to the class of flavanones having impressive pharmacological properties. Unfortunately, the in vivo bioavailability of NG is very low due to its higher hydrophobicity, which limits its practical use. Thus, in this study, we tried to develop NG-loaded macrophage membrane-coated liposome-based biomimetic nanoparticles with distinct physicochemical compositions and biological attributes for improving their bioavailability at the target site. The developed biomimetic nanoparticle (BNP) has shown good biocompatibility, stability, satisfactory particle size, pH-responsive drug (NG) release kinetics, and higher cellular uptake in vitro. The anti-metastatic efficacy of NGBNP has confirmed in syngeneic athymic BALB/c nude experimental models. By western blot analysis, semi-quantitative PCR, real-time PCR, and IHC, we conclude that NGBNP gets localized on the metastatic niche via its surface receptor α4, β1 integrin, and VCAM1 of metastatic cells and reduces the number of metastatic colonies in the lungs via regulating the apoptotic signaling axis.
PubMed: 37741236
DOI: 10.1016/j.foodchem.2023.137445 -
Oxidative Medicine and Cellular... 2020Numerous evidences suggest that plant polyphenols may have therapeutic benefits in regulating oxidative stress and providing neuroprotection in many neurodegenerative...
Olive Leaf Polyphenols Attenuate the Clinical Course of Experimental Autoimmune Encephalomyelitis and Provide Neuroprotection by Reducing Oxidative Stress, Regulating Microglia and SIRT1, and Preserving Myelin Integrity.
Numerous evidences suggest that plant polyphenols may have therapeutic benefits in regulating oxidative stress and providing neuroprotection in many neurodegenerative diseases, including multiple sclerosis (MS). However, these mechanisms are not yet completely understood. In this study, we investigated the effect of olive leaf polyphenols on oxidative stress through oxidation marker level and activity (TBARS, SOD, and GPX) and their protein expression (SOD1, SOD2, and GPX1), as well as the protein expression of Sirtuin 1 (SIRT1) and microglia markers (Iba-1, CD206, and iNOS) and myelin integrity (proteolipid protein expression) in the brain of rats with induced experimental autoimmune encephalomyelitis (EAE) and subjected to olive leaf therapy. Experiments were performed in male EAE DA rats, which were randomly divided into 2 main groups: EAE groups treated with the therapy of olive leaf (EAE+TOL) and untreated EAE control groups. The EAE treated groups consumed olive leaf tea instead of drinking water () from the beginning to the end of the experiment. In addition, olive leaf extract was injected intraperitoneally (.) for the 10 continuous days and started on the 8 day after EAE induction. The clinical course was monitored in both groups until the 30 day after EAE induction. Our results demonstrated that TOL attenuated the clinical course of EAE; reduced the oxidative stress (by decreasing the concentration of MDA); upregulated antioxidant enzymes (SOD1, SOD2, and GPX1), SIRT1 (overall and microglial), and anti-inflammatory M2 microglia; downregulated proinflammatory M1 type; and preserved myelin integrity. These data support the idea that TOL may be an effective therapeutic approach for treating MS and other neurodegenerative diseases.
Topics: Animals; Disease Models, Animal; Encephalomyelitis, Autoimmune, Experimental; Male; Microglia; Myelin Sheath; Olea; Oxidative Stress; Plant Leaves; Polyphenols; Rats; Sirtuin 1
PubMed: 32802267
DOI: 10.1155/2020/6125638 -
Science Advances Mar 2023The cytoskeletal proteins tubulin and actin are the obligate substrates of TCP-1 ring complex/Chaperonin containing TCP-1 (TRiC/CCT), and their folding involves...
The cytoskeletal proteins tubulin and actin are the obligate substrates of TCP-1 ring complex/Chaperonin containing TCP-1 (TRiC/CCT), and their folding involves co-chaperone. Through cryo-electron microscopy analysis, we present a more complete picture of TRiC-assisted tubulin/actin folding along TRiC adenosine triphosphatase cycle, under the coordination of co-chaperone plp2. In the open S1/S2 states, plp2 and tubulin/actin engaged within opposite TRiC chambers. Notably, we captured an unprecedented TRiC-plp2-tubulin complex in the closed S3 state, engaged with a folded full-length -tubulin and loaded with a guanosine triphosphate, and a plp2 occupying opposite rings. Another closed S4 state revealed an actin in the intermediate folding state and a plp2. Accompanying TRiC ring closure, plp2 translocation could coordinate substrate translocation on the CCT6 hemisphere, facilitating substrate stabilization and folding. Our findings reveal the folding mechanism of the major cytoskeletal proteins tubulin/actin under the coordination of the biogenesis machinery TRiC and plp2 and extend our understanding of the links between cytoskeletal proteostasis and related human diseases.
Topics: Humans; Actins; Cryoelectron Microscopy; MARVEL Domain-Containing Proteins; Molecular Chaperones; Protein Folding; Proteolipids; Tubulin; Cytoskeletal Proteins
PubMed: 36921056
DOI: 10.1126/sciadv.ade1207 -
Frontiers in Genetics 2022We aim to identify the crucial genes or potential biomarkers associated with uterine fibroids (UFs), which may provide clinicians with evidence about the diagnostic...
We aim to identify the crucial genes or potential biomarkers associated with uterine fibroids (UFs), which may provide clinicians with evidence about the diagnostic biomarker of UFs and reveal the mechanism of its progression. The gene expression and genome-wide DNA methylation profiles were obtained from Gene Expression Omnibus database (GEO). GSE45189, GSE31699, and GSE593 datasets were included. GEO2R and Venn diagrams were used to analyze the differentially expressed genes (DEGs) and extract the hub genes. Gene Ontology (GO) analysis was performed by the online tool Database for Annotation, Visualization, and Integrated Discovery (DAVID). The mRNA and protein expression of hub genes were validated by RT-qPCR, western blot, and immunohistochemistry. The receiver operating characteristic (ROC) curve was used to evaluate the diagnostic value. We detected 22 DEGs between UFs and normal myometrium, which were enriched in cell maturation, apoptotic process, hypoxia, protein binding, and cytoplasm for cell composition. By finding the intersection of the data between differentially expressed mRNA and DNA methylation profiles, 3 hub genes were identified, including transmembrane 4 L six family member 1 (TM4SF1), TNF superfamily member 10 (TNFSF10), and proteolipid protein 1 (PLP1). PLP1 was validated to be up-regulated significantly in UFs both at mRNA and protein levels. The area under the ROC curve (AUC) of PLP1 was 0.956, with a sensitivity of 79.2% and a specificity of 100%. Conclusion: Overall, our results indicate that PLP1 may be a potential diagnostic biomarker for uterine fibroids.
PubMed: 36386836
DOI: 10.3389/fgene.2022.1045395 -
Aging Sep 2021Ovarian carcinoma (OC) is the deadliest gynecologic malignancy in females worldwide. Circular RNA Foxo3 (Foxo3) plays essential roles in various cancers. However, the...
BACKGROUND
Ovarian carcinoma (OC) is the deadliest gynecologic malignancy in females worldwide. Circular RNA Foxo3 (Foxo3) plays essential roles in various cancers. However, the detailed function of Foxo3 in OC remains unclear. This study aimed to investigate the role of Foxo3 in OC and the underlying molecular mechanism.
METHODS
The abundance of Foxo3 was detected in OC cell lines by qPCR. Lentivirus transduction, CCK-8, wound healing assays, transwell migration and invasion assays, luciferase reporter assay, western blotting, fluorescence hybridization (FISH), transmission electron microscopy, nanoparticle tracking analysis, and bioinformatics analysis were performed to investigate the underlying mechanism.
RESULTS
The results demonstrated that Foxo3 was significantly upregulated in OC cell lines. Overexpression and knockdown of Foxo3 promoted and inhibited the proliferation, migration, and invasion of OC cells, respectively. Foxo3 could bind to miR-422a to negatively regulate miR-422a expression. Also, proteolipid protein 2 (PLP2) was a targeting gene of miR-422a. Additionally, Foxo3 was highly expressed in exosomes derived from OC cells. Furthermore, Foxo3 could be shuttled to OC cells by exosomes and promoted OC progression.
CONCLUSIONS
Foxo3 promoted OC progression through exosome-mediated intercellular interaction to target miR-422a/PLP2 axis. Foxo3 may serve as a potential biomarker for OC.
Topics: Carcinoma, Ovarian Epithelial; Computational Biology; Exosomes; Female; Forkhead Box Protein O3; Humans; In Situ Hybridization, Fluorescence; MARVEL Domain-Containing Proteins; MicroRNAs; Ovarian Neoplasms; Proteolipids; RNA, Circular; Up-Regulation
PubMed: 34555810
DOI: 10.18632/aging.203550 -
Biophysical Journal Jan 2020Membrane proteins are embedded in a complex lipid environment that influences their structure and function. One key feature of nearly all biological membranes is a...
Membrane proteins are embedded in a complex lipid environment that influences their structure and function. One key feature of nearly all biological membranes is a distinct lipid asymmetry. However, the influence of membrane asymmetry on proteins is poorly understood, and novel asymmetric proteoliposome systems are beneficial. To our knowledge, we present the first study on a multispanning protein incorporated in large unilamellar liposomes showing a stable lipid asymmetry. These asymmetric proteoliposomes contain the Na/H antiporter NhaA from Salmonella Typhimurium. Asymmetry was introduced by partial, outside-only exchange of anionic phosphatidylglycerol (PG), mimicking this key asymmetry of bacterial membranes. Outer-leaflet and total fractions of PG were determined via ζ-potential (ζ) measurements after lipid exchange and after scrambling of asymmetry. ζ-Values were in good agreement with exclusive outside localization of PG. The electrogenic Na/H antiporter was active in asymmetric liposomes, and it can be concluded that reconstitution and generation of asymmetry were successful. Lipid asymmetry was stable for more than 7 days at 23°C and thus enabled characterization of the Na/H antiporter in an asymmetric lipid environment. We present and validate a simple five-step protocol that addresses key steps to be taken and pitfalls to be avoided for the preparation of asymmetric proteoliposomes: 1) optimization of desired lipid composition, 2) detergent-mediated protein reconstitution with subsequent detergent removal, 3) generation of lipid asymmetry by partial exchange of outer-leaflet lipid, 4) verification of lipid asymmetry and stability, and 5) determination of protein activity in the asymmetric lipid environment. This work offers guidance in designing asymmetric proteoliposomes that will enable researchers to compare functional and structural properties of membrane proteins in symmetric and asymmetric lipid environments.
Topics: Lipids; Proteolipids; Salmonella typhimurium; Unilamellar Liposomes
PubMed: 31843262
DOI: 10.1016/j.bpj.2019.10.043