Browse
Submit Data
Databases
API
Help

Dataset Information

6 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Expression of Autophagy-Related Factors LC3A and Beclin 1 and Apoptosis-Related Factors Bcl-2 and BAX in Osteoblasts Treated With Sodium Fluoride.

ABSTRACT:

Objective

This study aims to analyze the expressions of autophagy-related factors light chain 3 alpha (LC3A) and Beclin 1 and apoptosis-related factors B-cell lymphoma 2 (Bcl-2) and Bcl-2-associated X (BAX) in primary osteoblasts treated with sodium fluoride (NaF).

Methods

Osteoblasts were extracted from Sprague-Dawley rats and treated with 0, 2.5, 5, and 10 mg/L NaF solutions, followed by 10 mmol/L 3-methyladenine (3-MA) for 24 h. The apoptotic rate was determined by flow cytometry, and the expressions of the autophagy- and apoptosis-related factors were measured by western blotting and real-time quantitative polymerase chain reaction.

Results

The mRNA expressions of LC3A, Beclin 1, and BAX in the NaF-treated osteoblast group were higher than those in the control group, while the protein expressions of these factors in the NaF-treated group were significantly higher than those in the control group. However, the Bcl-2 protein expression in the NaF-treated osteoblasts was significantly decreased compared to that in the control cells. After the 3-MA treatment, the protein expressions of LC3A, Beclin 1, and Bcl-2 were significantly decreased compared with those of the NaF-treated group, whereas the expression of BAX increased. Moreover, the apoptosis rate was increased after the addition of the 3-MA inhibitor.

Conclusion

NaF stimulation promoted autophagy and apoptosis of the osteoblasts, suggesting the involvement of fluoride damage in these processes.

SUBMITTER: Xu L

PROVIDER: S-EPMC8281676 | biostudies-literature |

REPOSITORIES: biostudies-literature

ACCESS DATA

Json Xml

Similar Datasets

Expression of SDF-1/CXCR4 and related inflammatory factors in sodium fluoride-treated hepatocytes.

Project description:At present, the mechanism of fluorosis-induced damage to the hepatic system is unclear. Studies have shown that excess fluoride causes some degree of damage to the liver, including inflammation. The SDF-1/CXCR4 signaling axis has been reported to have an impact on the regulation of inflammation in human cells. In this study, we investigated the role of the SDF-1/CXCR4 signaling axis and related inflammatory factors in fluorosis through in vitro experiments on human hepatic astrocytes (LX-2) cultured with sodium fluoride. CCK-8 assays showed that the median lethal dose at 24 h was 2 mmol/l NaF, and these conditions were used for subsequent enzyme-linked immunosorbent assays (ELISAs) and quantitative real-time polymerase chain reaction (qPCR) analysis. The protein expression levels of SDF-1/CXCR4 and the related inflammatory factors nuclear factor-κB (NF-κB), interleukin-6 (IL-6), tumor necrosis factor-α (TNF-α) and interleukin 1β (IL-1β) were detected by ELISAs from the experimental and control groups. The mRNA expression levels of these inflammatory indicators were also determined by qPCR in both groups. Moreover, the expression levels of these factors were significantly higher in the experimental group than in the control group at both the protein and mRNA levels (P < 0.05). Excess fluorine may stimulate the SDF-1/CXCR4 signaling axis, activating the inflammatory NF-κB signaling pathway and increasing the expression levels of the related inflammatory factors IL-6, TNF-α and IL-1β. Identification of this mechanism is important for elucidating the pathogenesis of fluorosis-induced liver injury.

| S-EPMC11192373 | biostudies-literature

The Bcl-2 homology domain 3 mimetic gossypol induces both Beclin 1-dependent and Beclin 1-independent cytoprotective autophagy in cancer cells.

Project description:Gossypol, a natural Bcl-2 homology domain 3 mimetic compound isolated from cottonseeds, is currently being evaluated in clinical trials. Here, we provide evidence that gossypol induces autophagy followed by apoptotic cell death in both the MCF-7 human breast adenocarcinoma and HeLa cell lines. We first show that knockdown of the Bcl-2 homology domain 3-only protein Beclin 1 reduces gossypol-induced autophagy in MCF-7 cells, but not in HeLa cells. Gossypol inhibits the interaction between Beclin 1 and Bcl-2 (B-cell leukemia/lymphoma 2), antagonizes the inhibition of autophagy by Bcl-2, and hence stimulates autophagy. We then show that knockdown of Vps34 reduces gossypol-induced autophagy in both cell lines, and consistent with this, the phosphatidylinositol 3-phosphate-binding protein WIPI-1 is recruited to autophagosomal membranes. Further, Atg5 knockdown also reduces gossypol-mediated autophagy. We conclude that gossypol induces autophagy in both a canonical and a noncanonical manner. Notably, we found that gossypol-mediated apoptotic cell death was potentiated by treatment with the autophagy inhibitor wortmannin or with small interfering RNA against essential autophagy genes (Vps34, Beclin 1, and Atg5). Our findings support the notion that gossypol-induced autophagy is cytoprotective and not part of the cell death process induced by this compound.

| S-EPMC2919121 | biostudies-literature

DAP-kinase-mediated phosphorylation on the BH3 domain of beclin 1 promotes dissociation of beclin 1 from Bcl-XL and induction of autophagy.

Project description:Autophagy, an evolutionarily conserved process, has functions both in cytoprotective and programmed cell death mechanisms. Beclin 1, an essential autophagic protein, was recently identified as a BH3-domain-only protein that binds to Bcl-2 anti-apoptotic family members. The dissociation of beclin 1 from its Bcl-2 inhibitors is essential for its autophagic activity, and therefore should be tightly controlled. Here, we show that death-associated protein kinase (DAPK) regulates this process. The activated form of DAPK triggers autophagy in a beclin-1-dependent manner. DAPK phosphorylates beclin 1 on Thr 119 located at a crucial position within its BH3 domain, and thus promotes the dissociation of beclin 1 from Bcl-XL and the induction of autophagy. These results reveal a substrate for DAPK that acts as one of the core proteins of the autophagic machinery, and they provide a new phosphorylation-based mechanism that reduces the interaction of beclin 1 with its inhibitors to activate the autophagic machinery.

| S-EPMC2658558 | biostudies-literature

LC3A-mediated autophagy regulates lung cancer cell plasticity.

Project description:AbbreviationsATG14: autophagy related 14; CDH2: cadherin 2; ChIP-qPCR: chromatin immunoprecipitation quantitative polymerase chain reaction; CQ: chloroquine; ECAR: extracellular acidification rate; EMT: epithelial-mesenchymal transition; EPCAM: epithelial cell adhesion molecule; MAP1LC3A/LC3A: microtubule associated protein 1 light chain 3 alpha; MAP1LC3B/LC3B: microtubule associated protein 1 light chain 3 beta; MAP1LC3C/LC3C: microtubule associated protein 1 light chain 3 gamma; NDUFV2: NADH:ubiquinone oxidoreductase core subunit V2; OCR: oxygen consumption rate; ROS: reactive oxygen species; RT-qPCR: reverse-transcriptase quantitative polymerase chain reaction; SC: scrambled control; shRNA: short hairpin RNA; SNAI2: snail family transcriptional repressor 2; SOX2: SRY-box transcription factor 2; SQSTM1/p62: sequestosome 1; TGFB/TGF-β: transforming growth factor beta; TOMM20: translocase of outer mitochondrial membrane 20; ZEB1: zinc finger E-box binding homeobox 1.

| S-EPMC9037431 | biostudies-literature

Prosurvival Bcl-2 family members affect autophagy only indirectly, by inhibiting Bax and Bak.

Project description:Antiapoptotic B-cell lymphoma 2 (Bcl-2) family members such as Bcl-2, myeloid cell leukemia 1 (Mcl-1), and B-cell lymphoma-X large (Bcl-xL) are proposed to inhibit autophagy by directly binding to the BH3 domain of Beclin 1/Atg6. However, these Bcl-2 family proteins also block the proapoptotic activity of Bcl-2-associated X (Bax) and Bcl-2 homologous antagonist/killer (Bak), and many inducers of autophagy also cause cell death. Therefore, when the mitochondrial-mediated apoptosis pathway is functional, interpretation of such experiments is complicated. To directly test the impact of the endogenous antiapoptotic Bcl-2 family members on autophagy in the absence of apoptosis, we inhibited their activity in cells lacking the essential cell death mediators Bax and Bak. We also used inducible lentiviral vectors to overexpress Bcl-2, Bcl-xL, or Mcl-1 in cells and subjected them to treatments that promote autophagy. In the absence of Bax and Bak, Bcl-2, Bcl-xL, and Mcl-1 had no detectable effect on autophagy or cell death in myeloid or fibroblast cell lines. On the other hand, when Bax and Bak were present, inhibiting the prosurvival Bcl-2 family members stimulated autophagy, but this correlated with increased cell death. In addition, inhibition of autophagy induced by amino acid starvation, etoposide, or interleukin-3 withdrawal did not affect cell death in the absence of Bax and Bak. These results demonstrate that the antiapoptotic Bcl-2 family members do not directly inhibit components of the autophagic pathway but instead affect autophagy indirectly, owing to their inhibition of Bax and Bak.

| S-EPMC4060681 | biostudies-literature

High-Throughput Screens To Identify Autophagy Inducers That Function by Disrupting Beclin 1/Bcl-2 Binding.

Project description:Autophagy, a lysosomal degradation pathway, plays a crucial role in cellular homeostasis, development, immunity, tumor suppression, metabolism, prevention of neurodegeneration, and lifespan extension. Thus, pharmacological stimulation of autophagy may be an effective approach for preventing or treating certain human diseases and/or aging. We sought to establish a method for developing new chemical compounds that specifically induce autophagy. To do this, we developed two assays to identify compounds that target a key regulatory node of autophagy induction-specifically, the binding of Bcl-2 (a negative regulator of autophagy) to Beclin 1 (an allosteric modulator of the Beclin 1/VPS34 lipid kinase complex that functions in autophagy initiation). These assays use either a split-luciferase assay to measure Beclin 1/Bcl-2 binding in cells or an AlphaLISA assay to directly measure direct Beclin 1/Bcl-2 binding in vitro. We screened two different chemical compound libraries, comprising ?300 K compounds, to identify small molecules that disrupt Beclin 1/Bcl-2 binding and induce autophagy. Three novel compounds were identified that directly inhibit Beclin 1/Bcl-2 interaction with an IC50 in the micromolar range and increase autophagic flux. These compounds do not demonstrate significant cytotoxicity, and they exert selectivity for disruption of Bcl-2 binding to the BH3 domain of Beclin 1 compared with the BH3 domain of the pro-apoptotic Bcl-2 family members, Bax and Bim. Thus, we have identified candidate molecules that serve as lead templates for developing potent and selective Beclin 1/Bcl-2 inhibitors that may be clinically useful as autophagy-inducing agents.

| S-EPMC6250051 | biostudies-literature

Antagonism of Beclin 1-dependent autophagy by BCL-2 at the endoplasmic reticulum requires NAF-1.

Project description:In addition to mitochondria, BCL-2 is located at the endoplasmic reticulum (ER) where it is a constituent of several distinct complexes. Here, we identify the BCL-2-interacting protein at the ER, nutrient-deprivation autophagy factor-1 (NAF-1)-a bitopic integral membrane protein whose defective expression underlies the aetiology of the neurodegenerative disorder Wolfram syndrome 2 (WFS2). NAF-1 contains a two iron-two sulphur coordinating domain within its cytosolic region, which is necessary, but not sufficient for interaction with BCL-2. NAF-1 is displaced from BCL-2 by the ER-restricted BH3-only protein BIK and contributes to regulation of BIK-initiated autophagy, but not BIK-dependent activation of caspases. Similar to BCL-2, NAF-1 is found in association with the inositol 1,4,5-triphosphate receptor and is required for BCL-2-mediated depression of ER Ca(2+) stores. During nutrient deprivation as a physiological stimulus of autophagy, BCL-2 is known to function through inhibition of the autophagy effector and tumour suppressor Beclin 1. NAF-1 is required in this pathway for BCL-2 at the ER to functionally antagonize Beclin 1-dependent autophagy. Thus, NAF-1 is a BCL-2-associated co-factor that targets BCL-2 for antagonism of the autophagy pathway at the ER.

| S-EPMC2830692 | biostudies-literature

Differential dependence on Beclin 1 for the regulation of pro-survival autophagy by Bcl-2 and Bcl-xL in HCT116 colorectal cancer cells.

Project description:Autophagy is described to be involved in homeostasis, development and disease, both as a survival and a death process. Its involvement in cell death proceeds from interrelationships with the apoptotic pathway. We focused on survival autophagy and investigated its interplays with the apoptotic machinery. We found that while Mcl-1 remained ineffective, Bcl-2 and Bcl-xL were required for starved cells to display a fully functional autophagic pathway as shown by proteolysis activity and detection of autophagic vesicles. Such pro-autophagic functions of Bcl-2 and Bcl-xL were independent of Bax. However they appeared to operate through non redundant mechanisms as Bcl-xL wielded a tighter control than Bcl-2 over the regulation of autophagy: unlike Bcl-2, Bcl-xL and Atg7 manipulation yielded identical phenotypes suggesting they could be components of the same signalling pathway; Bcl-xL subcellular localisation was modified upon starvation, and importantly Bcl-xL acted independently of Beclin 1. Still an intact BH3-binding site was required for Bcl-xL to stimulate a fully functional autophagic pathway. This study highlights that, in addition to their well-established anti-death function during apoptosis, Bcl-2 and Bcl-xL have a broader role in cell survival. Should Bcl-2 and Bcl-xL stand at the cross-roads between pro-survival and pro-death autophagy, this study introduces the new concept that the regulation of autophagy by Bcl-2 and Bcl-xL is adjusted according to its survival or death outcome.

| S-EPMC2807451 | biostudies-literature

Molecular basis of the regulation of Beclin 1-dependent autophagy by the gamma-herpesvirus 68 Bcl-2 homolog M11.

Project description:Gamma-herpesviruses (gammaHVs), including important human pathogens such as Epstein Barr virus, Kaposi's sarcoma-associated HV, and the murine gammaHV68, encode homologs of the antiapoptotic, cellular Bcl-2 (cBcl-2) to promote viral replication and pathogenesis. The precise molecular details by which these proteins function in viral infection are poorly understood. Autophagy, a lysosomal degradation pathway, is inhibited by the interaction of cBcl-2s with a key autophagy effector, Beclin 1, and can also be inhibited by gammaHV Bcl-2s. Here we investigate the gammaHV68 M11-Beclin 1 interaction in atomic detail, using biochemical and structural approaches. We show that the Beclin 1 BH3 domain is the primary determinant of binding to M11 and other Bcl-2s, and this domain binds in a hydrophobic groove on M11, reminiscent of the binding of different BH3 domains to other Bcl-2s. Unexpectedly, regions outside of, but contiguous with, the Beclin 1 BH3 domain also contribute to this interaction. We find that M11 binds to Beclin 1 more strongly than do KSHV Bcl-2 or cBcl-2. Further, the differential affinity of M11 for different BH3 domains is caused by subtle, yet significant, variations in the atomic details of each interaction. Consistent with our structural analysis, we find that Beclin 1 residues L116 and F123, and M11 residue pairs G86 + R87 and Y60 + L74, are required for M11 to bind to Beclin 1 and downregulate autophagy. Thus, our results suggest that M11 inhibits autophagy through a mechanism that involves the binding of the Beclin 1 BH3 domain in the M11 hydrophobic surface groove.

| S-EPMC2755179 | biostudies-literature

Regulation of the autophagic bcl-2/beclin 1 interaction.

Project description:Autophagy is an intracellular degradation process responsible for the delivery of cellular material to the lysosomes. One of the key mechanisms for control of autophagy is the modulation of the interaction between the autophagic protein Beclin 1 and the members of the anti-apoptotic Bcl-2 family (e.g., Bcl-2, Bcl-XL and Mcl-1). This binding is regulated by a variety of proteins and compounds that are able to enhance or inhibit the Bcl-2/Beclin 1 interaction in order to repress or activate autophagy, respectively. In this review we will focus on this interaction and discuss its characteristics, relevance and regulation.

| S-EPMC3901098 | biostudies-literature

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

Tweets

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data