scholarly journals Fundc1 is necessary for proper body axis formation during embryogenesis in zebrafish

2019 ◽  
Vol 9 (1) ◽  
Author(s):  
Gongyu Xu ◽  
Hao Shen ◽  
Emile Nibona ◽  
Kongyue Wu ◽  
Xiaomei Ke ◽  
...  
Keyword(s):  
T Cells ◽  
Mammalian Cells ◽  
Mitochondrial Protein ◽  
Ectopic Expression ◽  
The Body ◽  
Body Axis ◽  
Terminal Deoxynucleotidyl Transferase ◽  
Mitochondrial Outer Membrane ◽  
Axis Formation ◽  
Rare Minnow

AbstractFUN14 domain-containing protein 1 (FUNDC1) is a mitochondrial outer membrane protein which is responsible for hypoxia-induced mitophagy in mammalian cells. Knockdown of fundc1 is known to cause severe defects in the body axis of a rare minnow. To understand the role of Fundc1 in embryogenesis, we used zebrafish in this study. We used bioimaging to locate zebrafish Fundc1 (DrFundc1) with MitoTracker, a marker of mitochondria, and/or CellLight Lysosomes-GFP, a label of lysosomes, in the transfected ovary cells of grass carp. The use of Western blotting detected DrFundc1 as a component of mitochondrial proteins with endogenous COX IV, LC3B, and FUNDC1 in transgenic human embryonic kidney 293 T cells. DrFundc1 induced LC3B activation. The ectopic expression of Drfundc1 caused cell death and apoptosis as well as impairing cell proliferation in the 293 T cell line, as detected by Trypan blue, terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) and incorporation of BrdU. DrFundc1 up-regulated expression of both autophagy- and apoptosis-related genes, including ATG5, ATG7, LC3B, BECLIN1, and BAX in transgenic 293 T cells. A knockdown of Drfundc1 using short hairpin RNA (shRNA) led to midline bifurcation with two notochords and two spinal cords in zebrafish embryos. Co-injection of Drfundc1 mRNA repaired defects resulting from shRNA. Knockdown of Drfundc1 resulted in up- or down-regulation of genes related to autophagy and apoptosis, as well as decreased expression of neural genes such as cyclinD1, pax2a, opl, and neuroD1. In summary, DrFundc1 is a mitochondrial protein which is involved in mitophagy and is critical for typical body axis development in zebrafish.

Activin receptor mRNA is expressed early in Xenopus embryogenesis and the level of the expression affects the body axis formation

1991 ◽  
Vol 181 (2) ◽  
pp. 684-690 ◽  
Author(s):  
Mariko Kondo ◽  
Kosuke Tashiro ◽  
Gen Fujii ◽  
Misaki Asano ◽  
Ryutaro Miyoshi ◽  
...  
Keyword(s):  
The Body ◽  
Body Axis ◽  
Axis Formation ◽  
Activin Receptor ◽  
Receptor Mrna ◽  
Body Axis Formation

scholarly journals Involvement of Noxa in Cellular Apoptotic Responses to Interferon, Double-stranded RNA, and Virus Infection

2005 ◽  
Vol 280 (16) ◽  
pp. 15561-15568 ◽  
Author(s):  
Yaping Sun ◽  
Douglas W. Leaman
Keyword(s):  
Mitochondrial Protein ◽  
Membrane Integrity ◽  
Ectopic Expression ◽  
Human Tumor ◽  
Mitochondrial Outer Membrane ◽  
Dependent Manner ◽  
Mitochondrial Targeting ◽  
Double Stranded Rna ◽  
Bh3 Domain ◽  
Apoptosis Protein

Double-stranded RNA (dsRNA) accumulates in virally infected cells, leading to induction of genes encoding proteins involved in signaling, apoptosis, protein synthesis/processing, and cell metabolism. Noxa is a BH3-containing mitochondrial protein that contributes to apoptosis by disrupting mitochondrial outer membrane integrity. Here we demonstrate potent induction of Noxa expression by exposure of cells to dsRNA, interferon (IFN), and virus. Noxa induction was confirmed by using reverse transcriptase-PCR and immunoblot analyses in multiple human tumor cell lines. Importantly, Noxa regulation by IFN and dsRNA was independent of p53, thereby identifying a novel mechanism of Noxa induction. Ectopic expression of Noxa in HT1080 fibrosarcoma cells enhanced cellular sensitivity to viral or dsRNA/actinomycin D-induced apoptosis, typified by enhanced cytochromecrelease from the mitochondrial to the cytosolic fraction and increased cleavage of caspases 3 and 9. Point and deletion mutations of Noxa confirmed that both the BH3 domain and the mitochondrial-targeting domain were necessary for enhanced cellular apoptotic responses to dsRNA, IFN, or virus. Treatment of cells with dsRNA or virus, but not etoposide, induced interaction between Noxa and Bax that required an intact Noxa BH3 domain. Interestingly, the Noxa mitochondrial-targeting domain deletion mutant interacted with Bax in a dsRNA-dependent manner and redirected Bax away from the mitochondria, thus acting as a dominant-negative protein. Together, these data suggest that Noxa is an important component of the innate immune response of cells to viral infection, leading to enhanced cellular apoptosis that may play a role in limiting viral dissemination.

Cytoplasmic localization and chordamesoderm induction in the frog embryo

Development ◽  
1985 ◽  
Vol 89 (Supplement) ◽  
pp. 89-111
Author(s):  
Robert L. Gimlich
Keyword(s):  
Central Nervous System ◽  
Nervous System ◽  
Normal Development ◽  
Early Embryo ◽  
The Body ◽  
Body Axis ◽  
Host Cells ◽  
Normal Embryo ◽  
Axis Formation ◽  
Cytoplasmic Localization

The experiments described here were designed to reveal the distribution in the frog early embryo of components which are sufficient for specification of the dorsal structures of the embryonic body axis. The approach was to allow cleavage planes to divide the embryo into various well-defined regions and to transplant cells from each region into recipient embryos which would otherwise fail to form axial structures. Partial or complete body axis development could then be scored by the use of external criteria or histological methods. Recipients were embryos which had been irradiated before first cleavage with ultraviolet light on the vegetal surface. Irradiated embryos display a well-characterized set of deficiencies in the dorsal structures of the body axis, but their development can be ‘rescued’ toward normalcy in several ways. In particular, transplantation of certain small groups of blastomeres from the normal 32- to 64-cell embryo into irradiated recipients was sufficient to cause partial or complete axis development. Cell groups which could cause rescue were located in the vegetal and equatorial levels of one quadrant of the normal embryo — the quadrant centered on the future dorsal midline. Clonal marking analysis showed that the vegetal-most cells of this quadrant contribute primarily to endodermal structures in normal development. In rescued recipient embryos, these cells also contributed only to the endoderm; the dorsal mesoderm and central nervous system were formed exclusively by host cells which originated near the transplant. Rescue could also result from transplantation of equatorial cells from the dorsal quadrant of the normal embryo. As in normal development, these cells formed primarily the chordamesoderm of the rescued embryo. Host cells were induced to contribute the somitic mesoderm, central nervous system, and other structures which would have been missing but for the presence of the transplanted cells. The frequency and degree of rescue caused by equatorial and vegetal transplants is variable. This was explained by the discovery that the location of components needed for rescue varies among individual embryos without regard to the positions of cleavage planes. This was true even when donor embryos were selected on the basis of a precisely regular pattern of cleavage. In such selected embryos, particular blastomeres make a predictable contribution of progeny to the body axis. Thus it may be that the positions of components which can cause axis formation vary without exact regard to the fate map of prospective areas. The implications of this for the study of cytoplasmic localization in the early embryo are discussed. In any case, it is likely that regional interactions and a degree of developmental autonomy in the area of the prospective chordamesoderm are both involved in formation of the dorsal structures of the embryonic body axis.

The role of an endogenous PKA inhibitor, PKIα, in organizing left-right axis formation

Development ◽  
2001 ◽  
Vol 128 (13) ◽  
pp. 2509-2515
Author(s):  
Minoru Kawakami ◽  
Nobuki Nakanishi
Keyword(s):  
Protein Kinase ◽  
Kinase Inhibitor ◽  
Ectopic Expression ◽  
The Body ◽  
Specific Gene ◽  
Axis Formation ◽  
Lateral Plate ◽  
Heart Looping ◽  
Dependent Protein ◽  
The Right

Protein kinase inhibitor (PKI) is an endogenous inhibitor of cAMP-dependent protein kinase A (PKA). We have found that the α-isoform of PKI (PKIα) is asymmetrically expressed along the left-right (L-R) axis in chick embryos. At stage 6, PKIα is expressed on the right side of the node, and this asymmetric expression continues until stage 7+. After stage 8, PKIα expression returns symmetric. Treatment of embryos with antisense PKIα oligonucleotides increased the incidence of reversed heart looping. Antisense oligonucleotides also induced ectopic expression of the left-specific genes Nodal and Pitx2, and suppressed the expression of the right-specific gene SnR in the right lateral plate mesoderm. Similarly, treatment with PKA activators forskolin and Sp-cAMPs resulted in both reversed heart looping and bilateral expression of Nodal. Ectopic activin induced PKIα on the left side of the node, while ectopic Shh and anti-Shh antibody had no effect on PKIα expression. Taken together, these data suggest that PKIα induced by an activin-like molecule, through the inhibition of PKA activity, suppresses the Nodal-Pitx2 pathway on the right side of the body.

scholarly journals Heterodimeric IL-15 delays tumor growth and promotes intratumoral CTL and dendritic cell accumulation by a cytokine network involving XCL1, IFN-γ, CXCL9 and CXCL10

2020 ◽  
Vol 8 (1) ◽  
pp. e000599 ◽  
Author(s):  
Cristina Bergamaschi ◽  
Hrishikesh Pandit ◽  
Bethany A Nagy ◽  
Dimitris Stellas ◽  
Shawn M Jensen ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
T Cells ◽  
T Cell ◽  
Antitumor Activity ◽  
Tumor Growth ◽  
Mammalian Cells ◽  
The Body ◽  
Mouse Tumor ◽  
Dependent Manner ◽  
Ifn Γ

BackgroundInterleukin-15 (IL-15) promotes growth and activation of cytotoxic CD8+T and natural killer (NK) cells. Bioactive IL-15 is produced in the body as a heterodimeric cytokine, comprising the IL-15 and IL-15 receptor alpha chains (hetIL-15). Several preclinical models support the antitumor activity of hetIL-15 promoting its application in clinical trials.MethodsThe antitumor activity of hetIL-15 produced from mammalian cells was tested in mouse tumor models (MC38 colon carcinoma and TC-1 epithelial carcinoma). The functional diversity of the immune infiltrate and the cytokine/chemokine network within the tumor was evaluated by flow cytometry, multicolor immunohistochemistry (IHC), gene expression profiling by Nanostring Technologies, and protein analysis by electrochemiluminescence and ELISA assays.ResultshetIL-15 treatment resulted in delayed primary tumor growth. Increased NK and CD8+T cell tumoral infiltration with an increased CD8+/Treg ratio were found by flow cytometry and IHC in hetIL-15 treated animals. Intratumoral NK and CD8+T cells showed activation features with enhanced interferon-γ (IFN-γ) production, proliferation (Ki67+), cytotoxic potential (Granzyme B+) and expression of the survival factor Bcl-2. Transcriptomics and proteomics analyses revealed complex effects on the tumor microenvironment triggered by hetIL-15 therapy, including increased levels of IFN-γ and XCL1 with intratumoral accumulation of XCR1+IRF8+CD103+conventional type 1 dendritic cells (cDC1). Concomitantly, the production of the chemokines CXCL9 and CXCL10 by tumor-localized myeloid cells, including cDC1, was boosted by hetIL-15 in an IFN-γ-dependent manner. An increased frequency of circulating CXCR3+NK and CD8+T cells was found, suggesting their ability to migrate toward the tumors following the CXCL9 and CXCL10 chemokine gradient.ConclusionsOur results show that hetIL-15 administration enhances T cell entry into tumors, increasing the success rate of immunotherapy interventions. Our study further supports the incorporation of hetIL-15 in tumor immunotherapy approaches to promote the development of antitumor responses by favoring effector over regulatory cells and by promoting lymphocyte and DC localization into tumors through the modification of the tumor chemokine and cytokine milieu.

scholarly journals Regulation of Mitochondrial Morphology by USP30, a Deubiquitinating Enzyme Present in the Mitochondrial Outer Membrane

2008 ◽  
Vol 19 (5) ◽  
pp. 1903-1911 ◽  
Author(s):  
Nobuhiro Nakamura ◽  
Shigehisa Hirose
Keyword(s):  
Outer Membrane ◽  
Mammalian Cells ◽  
Mitochondrial Dynamics ◽  
Ectopic Expression ◽  
Mitochondrial Outer Membrane ◽  
Mitochondrial Morphology ◽  
Deubiquitinating Enzyme ◽  
Human Ubiquitin ◽  
Specific Protease ◽  
Insight Into

Recent studies have suggested that ubiquitination of mitochondrial proteins participates in regulating mitochondrial dynamics in mammalian cells, but it is unclear whether deubiquitination is involved in this process. Here, we identify human ubiquitin-specific protease 30 (USP30) as a deubiquitinating enzyme that is embedded in the mitochondrial outer membrane. Depletion of USP30 expression by RNA interference induced elongated and interconnected mitochondria, depending on the activities of the mitochondrial fusion factors mitofusins, without changing the expression levels of the key regulators for mitochondrial dynamics. Mitochondria were rescued from this abnormal phenotype by ectopic expression of USP30 in a manner dependent on its enzymatic activity. Our findings reveal that USP30 participates in the maintenance of mitochondrial morphology, a finding that provides new insight into the cellular function of deubiquitination.

scholarly journals Expression patterns of signalling molecules and transcription factors in the early rabbit embryo and their significance for modelling amniote axis formation

2021 ◽  
Author(s):  
Ruben Plöger ◽  
Christoph Viebahn
Keyword(s):  
Transcription Factors ◽  
Expression Patterns ◽  
In Situ Hybridisation ◽  
Body Plan ◽  
The Body ◽  
Anchor Point ◽  
Axis Formation ◽  
Point Model ◽  
Signalling Molecules ◽  
Rabbit Embryo

AbstractThe anterior-posterior axis is a central element of the body plan and, during amniote gastrulation, forms through several transient domains with specific morphogenetic activities. In the chick, experimentally proven activity of signalling molecules and transcription factors lead to the concept of a ‘global positioning system’ for initial axis formation whereas in the (mammotypical) rabbit embryo, a series of morphological or molecular domains are part of a putative ‘three-anchor-point model’. Because circular expression patterns of genes involved in axis formation exist in both amniote groups prior to, and during, gastrulation and may thus be suited to reconcile these models, the expression patterns of selected genes known in the chick, namely the ones coding for the transcription factors eomes and tbx6, the signalling molecule wnt3 and the wnt inhibitor pkdcc, were analysed in the rabbit embryonic disc using in situ hybridisation and placing emphasis on their germ layer location. Peripheral wnt3 and eomes expression in all layers is found initially to be complementary to central pkdcc expression in the hypoblast during early axis formation. Pkdcc then appears — together with a posterior-anterior gradient in wnt3 and eomes domains — in the epiblast posteriorly before the emerging primitive streak is marked by pkdcc and tbx6 at its anterior and posterior extremities, respectively. Conserved circular expression patterns deduced from some of this data may point to shared mechanisms in amniote axis formation while the reshaping of localised gene expression patterns is discussed as part of the ‘three-anchor-point model’ for establishing the mammalian body plan.

scholarly journals METTL3-dependent m6A modification programs T follicular helper cell differentiation

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Yingpeng Yao ◽  
Ying Yang ◽  
Wenhui Guo ◽  
Lifan Xu ◽  
Menghao You ◽  
...  
Keyword(s):  
T Cells ◽  
Transcriptional Regulation ◽  
Cell Differentiation ◽  
Ectopic Expression ◽  
Signature Genes ◽  
Follicular Helper ◽  
Conditional Deletion ◽  
A Cell ◽  
A Site ◽  
Post Transcriptional Regulation

AbstractT follicular helper (TFH) cells are specialized effector CD4+ T cells critical to humoral immunity. Whether post-transcriptional regulation has a function in TFH cells is unknown. Here, we show conditional deletion of METTL3 (a methyltransferase catalyzing mRNA N6-methyladenosine (m6A) modification) in CD4+ T cells impairs TFH differentiation and germinal center responses in a cell-intrinsic manner in mice. METTL3 is necessary for expression of important TFH signature genes, including Tcf7, Bcl6, Icos and Cxcr5 and these effects depend on intact methyltransferase activity. m6A-miCLIP-seq shows the 3′ UTR of Tcf7 mRNA is subjected to METTL3-dependent m6A modification. Loss of METTL3 or mutation of the Tcf7 3′ UTR m6A site results in accelerated decay of Tcf7 transcripts. Importantly, ectopic expression of TCF-1 (encoded by Tcf7) rectifies TFH defects owing to METTL3 deficiency. Our findings indicate that METTL3 stabilizes Tcf7 transcripts via m6A modification to ensure activation of a TFH transcriptional program, indicating a pivotal function of post-transcriptional regulation in promoting TFH cell differentiation.

scholarly journals Selenium deficiency induces spleen pathological changes in pigs by decreasing selenoprotein expression, evoking oxidative stress, and activating inflammation and apoptosis

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Shuang Li ◽  
Wenjuan Sun ◽  
Kai Zhang ◽  
Jiawei Zhu ◽  
Xueting Jia ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Inflammatory Cytokines ◽  
The Body ◽  
Terminal Deoxynucleotidyl Transferase ◽  
Antioxidant Systems ◽  
Tunel Staining ◽  
White Pulp ◽  
Sibling Pairs ◽  
Se Deficiency ◽  
Spleen Index

Abstract Background The immune system is one aspect of health that is affected by dietary selenium (Se) levels and selenoprotein expression. Spleen is an important immune organ of the body, which is directly involved in cellular immunity. However, there are limited reports on Se levels and spleen health. Therefore, this study established a Se-deficient pig model to investigate the mechanism of Se deficiency-induced splenic pathogenesis. Methods Twenty-four pure line castrated male Yorkshire pigs (45 days old, 12.50 ± 1.32 kg, 12 full-sibling pairs) were divided into two equal groups and fed Se-deficient diet (0.007 mg Se/kg) or Se-adequate diet (0.3 mg Se/kg) for 16 weeks. At the end of the trial, blood and spleen were collected to assay for erythroid parameters, the osmotic fragility of erythrocytes, the spleen index, histology, terminal deoxynucleotidyl transferase nick-end labeling (TUNEL) staining, Se concentrations, the selenogenome, redox status, and signaling related inflammation and apoptosis. Results Dietary Se deficiency decreased the erythroid parameters and increased the number of osmotically fragile erythrocytes (P < 0.05). The spleen index did not change, but hematoxylin and eosin and TUNEL staining indicated that the white pulp decreased, the red pulp increased, and splenocyte apoptosis occurred in the Se deficient group. Se deficiency decreased the Se concentration and selenoprotein expression in the spleen (P < 0.05), blocked the glutathione and thioredoxin antioxidant systems, and led to redox imbalance. Se deficiency activated the NF-κB and HIF-1α transcription factors, thus increasing pro-inflammatory cytokines (IL-1β, IL-6, IL-8, IL-17, and TNF-α), decreasing anti-inflammatory cytokines (IL-10, IL-13, and TGF-β) and increasing expression of the downstream genes COX-2 and iNOS (P < 0.05), which in turn induced inflammation. In addition, Se-deficiency induced apoptosis through the mitochondrial pathway, upregulated apoptotic genes (Caspase3, Caspase8, and Bak), and downregulated antiapoptotic genes (Bcl-2) (P < 0.05) at the mRNA level, thus verifying the results of TUNEL staining. Conclusions These results indicated that Se deficiency induces spleen injury through the regulation of selenoproteins, oxidative stress, inflammation and apoptosis.

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