Cleavage of SPACA1 regulates assembly of sperm–egg membrane fusion machinery in mature spermatozoa†

2019 ◽  
Vol 102 (3) ◽  
pp. 750-757
Author(s):  
Kenji Yamatoya ◽  
Marika Kousaka ◽  
Chizuru Ito ◽  
Kazuya Nakata ◽  
Masahiko Hatano ◽  
...  
Keyword(s):  
Membrane Fusion ◽  
Acrosome Reaction ◽  
Molecular Mechanisms ◽  
Morphological Changes ◽  
Reaction Process ◽  
Related Factor ◽  
Gamete Fusion ◽  
C Terminus ◽  
Egg Membrane ◽  
Mature Spermatozoa

Abstract The acrosome reaction is a multi-step event essential for physiological fertilization. During the acrosome reaction, gamete fusion-related factor IZUMO1 translocates from the anterior acrosome to the equatorial segment and assembles the gamete fusion machinery. The morphological changes in the acrosome reaction process have been well studied, but little is known about the molecular mechanisms of acrosome reorganization essential for physiological gamete membrane fusion. To elucidate the molecular mechanisms of IZUMO1 translocation, the steps of the acrosome reaction during that process must be clarified. In this study, we established a method to detect the early steps of the acrosome reaction and subdivided the process into seven populations through the use of two epitope-defined antibodies, anti-IZUMO1 and anti-SPACA1, a fertilization-inhibiting antibody. We found that part of the SPACA1 C-terminus in the periacrosomal space was cleaved and had begun to disappear when the vesiculation of the anterior acrosome occurred. The IZUMO1 epitope externalized from the acrosomal lumen before acrosomal vesiculation and phosphorylation of IZUMO1 occurred during the translocation to the equatorial segment. IZUMO1 circumvented the area of the equatorial segment where the SPACA1C-terminus was still localized. We therefore propose an IZUMO1 translocation model and involvement of SPACA1.

scholarly journals Isoliquiritigenin Confers Neuroprotection and Alleviates Amyloid-β42-Induced Neuroinflammation in Microglia by Regulating the Nrf2/NF-κB Signaling

2021 ◽  
Vol 15 ◽  
Author(s):  
Yue Fu ◽  
Jianping Jia
Keyword(s):  
Oxidative Stress ◽  
Inflammatory Response ◽  
Molecular Mechanisms ◽  
Morphological Changes ◽  
No Production ◽  
Related Factor ◽  
Nuclear Factor ◽  
Qrt Pcr ◽  
Bv2 Cells ◽  
And Oxidative Stress

BackgroundNeuroinflammation and oxidative stress are two major pathological characteristics of Alzheimer’s disease (AD). Amyloid-β oligomers (AβO), a toxic form of Aβ, promote the neuroinflammation and oxidative stress in the development of AD. Isoliquiritigenin (ISL), a natural flavonoid isolated from the root of liquorice, has been shown to exert inhibitory effects on inflammatory response and oxidative stress.ObjectivesThe main purpose of this study is to assess the influence of ISL on inflammatory response and oxidative stress in BV2 cells stimulated with AβO, and to explore the underlying molecular mechanisms.Methods3-(4,5-dimethyl-2-thiazolyl)-2, 5-diphenyl-2-H- tetrazolium bromide (MTT) and lactate dehydrogenase (LDH) cytotoxicity assays were used to assess the toxic or protective effects of ISL. The expression levels of interleukin-1β, interleukin-6, and tumor necrosis factor-α were assessed by quantitative real-time polymerase chain reaction (qRT-PCR) and enzyme-linked immunosorbent assays. Morphological changes in BV2 cells were assessed by immunofluorescence method. Nitric oxide (NO) assay kit was used to determinate the NO production. Western blot, qRT-PCR and immunofluorescence were used to explore the underlying molecular mechanisms.ResultsISL treatment reduced the production of inflammatory cytokines and NO, and alleviated the morphological changes in BV2 cells induced by AβO. ISL treatment further protected N2a cells from the toxic medium of AβO-stimulated BV2 cells. ISL activated nuclear factor erythroid-2 related factor 2 (Nrf2) signaling and suppressed nuclear factor-κB (NF-κB) signaling in BV2 cells.ConclusionISL suppresses AβO-induced inflammation and oxidative stress in BV2 cells via the regulation of Nrf2/NF-κB signaling. Therefore, ISL indirectly protects neurons from the damage of toxic conditioned media.

scholarly journals Evidence for involvement of metalloendoproteases in a step in sea urchin gamete fusion.

1988 ◽  
Vol 107 (2) ◽  
pp. 539-544 ◽  
Author(s):  
J L Roe ◽  
H A Farach ◽  
W J Strittmatter ◽  
W J Lennarz
Keyword(s):  
Plasma Membrane ◽  
Membrane Fusion ◽  
Sea Urchin ◽  
Acrosome Reaction ◽  
Fusion Process ◽  
Cortical Granule ◽  
Cortical Granules ◽  
Gamete Fusion ◽  
Cortical Granule Exocytosis ◽  
Cytoplasmic Continuity

Membrane fusion events are required in three steps in sea urchin fertilization: the acrosome reaction in sperm, fusion of the plasma membrane of acrosome-reacted sperm with the plasma membrane of the egg, and exocytosis of the contents of the egg cortical granules. We recently reported the involvement of a Zn2+-dependent metalloendoprotease in the acrosome reaction (Farach, H. C., D. I. Mundy, W. J. Strittmatter, and W. J. Lennarz. 1987. J. Biol. Chem. 262:5483-5487). In the current study, we investigated the possible involvement of metalloendoproteases in the two other fusion events of fertilization. The use of inhibitors of metalloendoproteases provided evidence that at least one of the fusion events subsequent to the acrosome reaction requires such enzymes. These inhibitors did not block the binding of sperm to egg or the process of cortical granule exocytosis. However, sperm-egg fusion, assayed by the ability of the bound sperm to establish cytoplasmic continuity with the egg, was inhibited by metalloendoprotease substrate. Thus, in addition to the acrosome reaction, an event in the gamete fusion process requires a metalloendoprotease.

Fertilization in ascidians: apical processes and gamete fusion in Ciona intestinalis spermatozoa

1988 ◽  
Vol 89 (2) ◽  
pp. 189-196
Author(s):  
MAKOTO FUKUMOTO
Keyword(s):  
Marine Invertebrates ◽  
Morphological Changes ◽  
Ciona Intestinalis ◽  
Sperm Head ◽  
The Other ◽  
Perivitelline Space ◽  
Apical Process ◽  
Electron Dense Material ◽  
Gamete Fusion ◽  
Egg Membrane

This paper describes the fine structure of the spermatozoan apex and its morphological changes during the process of fertilization in Ciona intestinalis. At the apex of the head an acrosome is present in the form of a flattened vesicle containing moderately electron-dense material with an electron-dense plate in its centre. Contiguous to the acrosome, an apical substance is located in the anterior-most tip of the sperm head. The spermatozoa that bind to and penetrate the chorion of both normal and caffeine-treated eggs have an intact acrosome and do not show an observable alteration of the plasmalemma enclosing the apex of the head. On the other hand, spermatozoa in the perivitelline space of both normal and caffeine-treated eggs lack an acrosome. Instead of an acrosome, apical processes are observed at the apex of the spermatozoa in the perivitelline space of both normal and caffeine-treated eggs. Gamete fusion seems to occur between the egg membrane and some of these apical processes at the tip of the sperm head. The apical process reported here is thought to be analogous to the acrosomal process in other marine invertebrates. The apical processes that are induced in the perivitelline space have not been previously described in ascidians.

Residues SFQ (173-175) in the large extracellular loop of CD9 are required for gamete fusion

Development ◽  
2002 ◽  
Vol 129 (8) ◽  
pp. 1995-2002 ◽  
Author(s):  
Guo-Zhang Zhu ◽  
Brent J. Miller ◽  
Claude Boucheix ◽  
Eric Rubinstein ◽  
Christopher C. Liu ◽  
...  
Keyword(s):  
Membrane Fusion ◽  
Point Mutation ◽  
Active Site ◽  
Gene Knockout ◽  
Extracellular Loop ◽  
Gamete Fusion ◽  
Extracellular Loops ◽  
Egg Membrane ◽  
Large Extracellular Loop ◽  
Fusion Ability

Gamete fusion is the fundamental first step initiating development of a new organism. Female mice with a gene knockout for the tetraspanin CD9 (CD9 KO mice) produce mature eggs that cannot fuse with sperm. However, nothing is known about how egg surface CD9 functions in the membrane fusion process. We found that constructs including CD9’s large extracellular loop significantly inhibited gamete fusion when incubated with eggs but not when incubated with sperm, suggesting that CD9 acts by interaction with other proteins in the egg membrane. We also found that injecting developing CD9 KO oocytes with CD9 mRNA restored fusion competence to the resulting CD9 KO eggs. Injecting mRNA for either mouse CD9 or human CD9, whose large extracellular loops differ in 18 residues, rescued fusion ability of the injected CD9 KO eggs. However, when the injected mouse CD9 mRNA contained a point mutation (F174 to A) the gamete fusion level was reduced fourfold, and a change of three residues (173-175, SFQ to AAA) abolished CD9’s activity in gamete fusion. These results suggest that SFQ in the CD9 large extracellular loop may be an active site which associates with and regulates the egg fusion machinery.

Trilobatin Protects Cardiomyocytes from Hypoxia/ Reperfusion Injury by Regulating the Nuclear Factor Erythroid 2-Related Factor 2/Heme Oxygenase-1 Pathway

2020 ◽  
Vol 19 (2) ◽  
pp. 133-138
Author(s):  
Wenyu Chen ◽  
Hui He
Keyword(s):  
Heme Oxygenase ◽  
Molecular Mechanisms ◽  
Heme Oxygenase 1 ◽  
Related Factor ◽  
Cellular Injury ◽  
Nuclear Factor ◽  
Oxygen Species ◽  
H9c2 Cells ◽  
Natural Plant ◽  
Induced Changes

Trilobatin is a natural plant-derived glycosylated flavonoid that has been shown to exhibit multiple beneficial pharmacologic activities including protection of heart against H/R-induced cardiomyocyte injury. However, the molecular mechanisms underlying protection from H/R-induced cardiomyocyte injury remain unknown. Using H9C2 cells as a model, we examined the effect of trilobatin on H/R-induced cellular injury, apoptosis, and generation of reactive oxygen species. The results showed that trilobatin protected H9C2 cells not only from cell death and apoptosis, but also counteracted H/R-induced changes in malondialdehyde, superoxide dismutase, glutathione, and glutathione peroxidase. The evaluation of the mechanism underlying the effect of trilobatin on protection from H/R-induced cellular injury suggested changes in the regulation of nuclear factor erythroid 2-related factor 2/heme oxygenase-1 pathway.

The Protective Effects of Green Tea Catechins in the Management of Neurodegenerative Diseases: A Review

2019 ◽  
Vol 16 (1) ◽  
pp. 57-65 ◽  
Author(s):  
Tahereh Farkhondeh ◽  
Hanieh Shaterzadeh Yazdi ◽  
Saeed Samarghandian
Keyword(s):  
Neurodegenerative Diseases ◽  
Signaling Pathways ◽  
Green Tea ◽  
Molecular Mechanisms ◽  
Main Role ◽  
Related Factor ◽  
Protective Effects ◽  
Tea Catechins ◽  
Green Tea Catechins ◽  
And Control

Background: The therapeutic strategies to manage neurodegenerative diseases remain limited and it is necessary to discover new agents for their prevention and control. Oxidative stress and inflammation play a main role in the pathogenesis of neurodegenerative diseases. The aim of this study is to review the effects of green tea catechins against the Neurodegenerative Diseases. Methods: In this study, we extensively reviewed all articles on the terms of Green tea, catechins, CNS disorders, and different diseases in PubMed, Science Direct, Scopus, and Google Scholar databases between the years 1990 and 2017. Results: The present study found that catechins, the major flavonoids in green tea, are powerful antioxidants and radical scavengers which possess the potential roles in the management of neurodegenerative diseases. Catechins modulate the cellular and molecular mechanisms through the inflammation-related NF-&amp;#954;B and the nuclear factor erythroid 2-related factor 2 (Nrf2) signaling pathways. Conclusion: The findings of the present review shows catechins could be effective against neurodegenerative diseases due to their antioxidation and anti-inflammation effects and the involved biochemical pathways including Nrf2 and NF-kB signaling pathways.<P&gt;

scholarly journals Platelet-derived lysophosphatidic acid mediated LPAR1 activation as a therapeutic target for osteosarcoma metastasis

Oncogene ◽  
2021 ◽  
Author(s):  
Satoshi Takagi ◽  
Yuki Sasaki ◽  
Sumie Koike ◽  
Ai Takemoto ◽  
Yosuke Seto ◽  
...  
Keyword(s):  
Platelet Activation ◽  
Pulmonary Metastasis ◽  
Lysophosphatidic Acid ◽  
Therapeutic Intervention ◽  
Molecular Mechanisms ◽  
Morphological Changes ◽  
Osteosarcoma Cells ◽  
Invasion And Metastasis ◽  
Platelet Releasate

AbstractOsteosarcoma is the most common primary malignant bone cancer, with high rates of pulmonary metastasis. Osteosarcoma patients with pulmonary metastasis have worse prognosis than those with localized disease, leading to dramatically reduced survival rates. Therefore, understanding the biological characteristics of metastatic osteosarcoma and the molecular mechanisms of invasion and metastasis of osteosarcoma cells will lead to the development of innovative therapeutic intervention for advanced osteosarcoma. Here, we identified that osteosarcoma cells commonly exhibit high platelet activation-inducing characteristics, and molecules released from activated platelets promote the invasiveness of osteosarcoma cells. Given that heat-denatured platelet releasate maintained the ability to promote osteosarcoma invasion, we focused on heat-tolerant molecules, such as lipid mediators in the platelet releasate. Osteosarcoma-induced platelet activation leads to abundant lysophosphatidic acid (LPA) release. Exposure to LPA or platelet releasate induced morphological changes and increased invasiveness of osteosarcoma cells. By analyzing publicly available transcriptome datasets and our in-house osteosarcoma patient-derived xenograft tumors, we found that LPA receptor 1 (LPAR1) is notably upregulated in osteosarcoma. LPAR1 gene KO in osteosarcoma cells abolished the platelet-mediated osteosarcoma invasion in vitro and the formation of early pulmonary metastatic foci in experimental pulmonary metastasis models. Of note, the pharmacological inhibition of LPAR1 by the orally available LPAR1 antagonist, ONO-7300243, prevented pulmonary metastasis of osteosarcoma in the mouse models. These results indicate that the LPA–LPAR1 axis is essential for the osteosarcoma invasion and metastasis, and targeting LPAR1 would be a promising therapeutic intervention for advanced osteosarcoma.

scholarly journals Crosstalk between Long-Term Sublethal Oxidative Stress and Detrimental Inflammation as Potential Drivers for Age-Related Retinal Degeneration

Antioxidants ◽  
2020 ◽  
Vol 10 (1) ◽  
pp. 25
Author(s):  
Lara Macchioni ◽  
Davide Chiasserini ◽  
Letizia Mezzasoma ◽  
Magdalena Davidescu ◽  
Pier Luigi Orvietani ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Molecular Mechanisms ◽  
Age Related Macular Degeneration ◽  
Inflammasome Activation ◽  
Related Factor ◽  
Nuclear Factor ◽  
Rpe Cells ◽  
Age Related ◽  
Oxidative Insult

Age-related retinal degenerations, including age-related macular degeneration (AMD), are caused by the loss of retinal pigmented epithelial (RPE) cells and photoreceptors. The pathogenesis of AMD, deeply linked to the aging process, also involves oxidative stress and inflammatory responses. However, the molecular mechanisms contributing to the shift from healthy aging to AMD are still poorly understood. Since RPE cells in the retina are chronically exposed to a pro-oxidant microenvironment throughout life, we simulated in vivo conditions by growing ARPE-19 cells in the presence of 10 μM H2O2 for several passages. This long-term oxidative insult induced senescence in ARPE-19 cells without affecting cell proliferation. Global proteomic analysis revealed a dysregulated expression in proteins involved in antioxidant response, mitochondrial homeostasis, and extracellular matrix organization. The analyses of mitochondrial functionality showed increased mitochondrial biogenesis and ATP generation and improved response to oxidative stress. The latter, however, was linked to nuclear factor-κB (NF-κB) rather than nuclear factor erythroid 2–related factor 2 (Nrf2) activation. NF-κB hyperactivation also resulted in increased pro-inflammatory cytokines expression and inflammasome activation. Moreover, in response to additional pro-inflammatory insults, senescent ARPE-19 cells underwent an exaggerated inflammatory reaction. Our results indicate senescence as an important link between chronic oxidative insult and detrimental chronic inflammation, with possible future repercussions for therapeutic interventions.

scholarly journals SNARE phosphorylation: a control mechanism for insulin-stimulated glucose transport and other regulated exocytic events

2017 ◽  
Vol 45 (6) ◽  
pp. 1271-1277 ◽  
Author(s):  
Kamilla M.E. Laidlaw ◽  
Rachel Livingstone ◽  
Mohammed Al-Tobi ◽  
Nia J. Bryant ◽  
Gwyn W. Gould
Keyword(s):  
Membrane Fusion ◽  
Molecular Mechanisms ◽  
Membrane Receptors ◽  
Multivesicular Bodies ◽  
Attachment Protein ◽  
Protein Receptor ◽  
Sensitive Factor ◽  
Plasma Membrane Receptors ◽  
Regulated Trafficking ◽  
Receptor Family

Trafficking within eukaryotic cells is a complex and highly regulated process; events such as recycling of plasma membrane receptors, formation of multivesicular bodies, regulated release of hormones and delivery of proteins to membranes all require directionality and specificity. The underpinning processes, including cargo selection, membrane fusion, trafficking flow and timing, are controlled by a variety of molecular mechanisms and engage multiple families of lipids and proteins. Here, we will focus on control of trafficking processes via the action of the SNARE (soluble N-ethylmaleimide-sensitive factor attachment protein receptor) family of proteins, in particular their regulation by phosphorylation. We will describe how these proteins are controlled in a range of regulated trafficking events, with particular emphasis on the insulin-stimulated delivery of glucose transporters to the surface of adipose and muscle cells. Here, we focus on a few examples of SNARE phosphorylation which exemplify distinct ways in which SNARE machinery phosphorylation may regulate membrane fusion.

scholarly journals Palmitoylation of Sindbis Virus TF Protein Regulates Its Plasma Membrane Localization and Subsequent Incorporation into Virions

2016 ◽  
Vol 91 (3) ◽  
Author(s):  
Jolene Ramsey ◽  
Emily C. Renzi ◽  
Randy J. Arnold ◽  
Jonathan C. Trinidad ◽  
Suchetana Mukhopadhyay
Keyword(s):  
Plasma Membrane ◽  
Sindbis Virus ◽  
Molecular Mechanisms ◽  
Wild Type Virus ◽  
Membrane Localization ◽  
Clear Understanding ◽  
Wild Type ◽  
Plasma Membrane Localization ◽  
C Terminus ◽  
N Terminus

ABSTRACT Palmitoylation is a reversible, posttranslational modification that helps target proteins to cellular membranes. The alphavirus small membrane proteins 6K and TF have been reported to be palmitoylated and to positively regulate budding. 6K and TF are isoforms that are identical in their N termini but unique in their C termini due to a −1 ribosomal frameshift during translation. In this study, we used cysteine (Cys) mutants to test differential palmitoylation of the Sindbis virus 6K and TF proteins. We modularly mutated the five Cys residues in the identical N termini of 6K and TF, the four additional Cys residues in TF's unique C terminus, or all nine Cys residues in TF. Using these mutants, we determined that TF palmitoylation occurs primarily in the N terminus. In contrast, 6K is not palmitoylated, even on these shared residues. In the C-terminal Cys mutant, TF protein levels increase both in the cell and in the released virion compared to the wild type. In viruses with the N-terminal Cys residues mutated, TF is much less efficiently localized to the plasma membrane, and it is not incorporated into the virion. The three Cys mutants have minor defects in cell culture growth but a high incidence of abnormal particle morphologies compared to the wild-type virus as determined by transmission electron microscopy. We propose a model where the C terminus of TF modulates the palmitoylation of TF at the N terminus, and palmitoylated TF is preferentially trafficked to the plasma membrane for virus budding. IMPORTANCE Alphaviruses are a reemerging viral cause of arthritogenic disease. Recently, the small 6K and TF proteins of alphaviruses were shown to contribute to virulence in vivo. Nevertheless, a clear understanding of the molecular mechanisms by which either protein acts to promote virus infection is missing. The TF protein is a component of budded virions, and optimal levels of TF correlate positively with wild-type-like particle morphology. In this study, we show that the palmitoylation of TF regulates its localization to the plasma membrane, which is the site of alphavirus budding. Mutants in which TF is not palmitoylated display drastically reduced plasma membrane localization, which effectively prevents TF from participating in budding or being incorporated into virus particles. Investigation of the regulation of TF will aid current efforts in the alphavirus field searching for approaches to mitigate alphaviral disease in humans.

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