Proteomic and phosphoproteomic landscape underlying DGAT1 inhibitor-ameliorated lipotoxicity in Min6 cells
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ABSTRACT:
ORGANISM(S): Mus Musculus
SUBMITTER: 
Hu Zhou
PROVIDER: PXD070881 | iProX | Tue Nov 18 00:00:00 GMT 2025
REPOSITORIES: iProX
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Project description:Protein phosphatase 2A (PP2A) is one of the most common serine/threonine phosphatases in mammalian cells and primarily functions to regulate cell signaling, glycolipid metabolism, and apoptosis. Its PP2A catalytic subunit (PP2Ac) plays an important role in its function. Nonetheless, at present, there are only a few reports on the regulatory role of PP2Ac in pancreatic β-cells under lipotoxicity. Mouse pancreatic insulinoma (MIN6) cells were transfected by lentiviruses to generate PP2Ac knockdown cells and incubated with palmitate (PA) to establish the lipotoxicity model. Serine/Threonine Phosphatase Assay System kit, Cell Counting Kit-8 (CCK-8), flow cytometry, enzyme-linked immunosorbent assay (ELISA), Western Blotting (WB) and other techniques were used to measure PP2A activity, cell viability, apoptosis, oxidative stress, and insulin secretion. An animal model of lipotoxicity with knockdown of PP2Ac was established by a high-fat diet (HFD) after using adeno-associated viruses (AAV) to interfere with PP2Ac expression in mouse pancreatic tissues. Serine/Threonine Phosphatase Assay System kit, ELISA, pancreatic tissue immunofluorescence and other techniques were used to measure PP2A activity in pancreatic tissue, serum insulin level and the proliferation of mouse pancreatic β-cells. We found that PP2Ac knockdown inhibited lipotoxicity-induced PP2A hyperactivation, increased the resistance of pancreatic β-cells to lipotoxicity, decreased PA-induced apoptosis in MIN6 cells, protected the function of both the endoplasmic reticulum (ER) and mitochondria, and improved insulin secretion. By mRNA sequencing and Western blotting analysis, we hypothesized that the protective effects of PP2Ac knockdown in MIN6 cells may be mediated by the MAPK pathway. Moreover, the results obtained from animal experiments suggest that the specific knockdown of pancreatic PP2Ac could effectively attenuate HFD-induced insulin resistance and reduce the compensatory proliferation of pancreatic β-cells in mice. The present study revealed the effects and mechanisms of interfering with PP2Ac gene expression on pancreatic β-cells in vivo and in vitro, which might provide insights for the treatment of type 2 diabetes in the clinic.
2023-09-13 | GSE242538 | GEO
Project description:To identify the genes induced by lipotoxicity and integrated stress response in pancreatic beta cells, we have employed whole genome microarray expression in panvreatic beta cell line MIN6m9 cells.
2015-08-01 | GSE49647 | GEO
Project description:The accumulation of lipid is a histological and biochemical hallmark of obesity-associated hepato-steatosis.Moreover, growing evidenceindicatesthathigher free fatty acids (FFAs) level in hepatocytes affects a myriad of biological processes leading to excessive metabolic imbalance, increased reactive oxygen species (ROS), deregulated autophagy, and impairment of mitochondrial and ER stress, that collectively drives cell death. Lipotoxicity and cell death mechanisms have been studied for many years. However, the molecular signals that link these two events during lipid stress remain poorly understood. From the very beginning, to systematically study hepato-lipotoxicity, HepG2 treated with PA providentially recapitulates the global lipotoxic responses, including insulin resistance to hepatocyte death. Therefore, using this cell-based model system, we pursued a comprehensive, differential quantitative approach where measurements of protein dynamics are analysed by mass spectrometry. Given that indispensable information, successive temporal phosphoproteomics dynamics are allowed us to in-depth analysis of lipotoxicity associated mechanistic network of cell death more precisely.
2022-07-16 | PXD029016 | Pride
Project description:The accumulation of lipid is a histological and biochemical hallmark of obesity-associated hepato-steatosis.Moreover, growing evidenceindicatesthathigher free fatty acids (FFAs) level in hepatocytes affects a myriad of biological processes leading to excessive metabolic imbalance, increased reactive oxygen species (ROS), deregulated autophagy, and impairment of mitochondrial and ER stress, that collectively drives cell death. Lipotoxicity and cell death mechanisms have been studied for many years. However, the molecular signals that link these two events during lipid stress remain poorly understood. From the very beginning, to systematically study hepato-lipotoxicity, HepG2 treated with PA providentially recapitulates the global lipotoxic responses, including insulin resistance to hepatocyte death. Therefore, using this cell-based model system, we pursued a comprehensive, differential quantitative approach where measurements of protein dynamics are analysed by mass spectrometry. Given that indispensable information, successive temporal phosphoproteomics dynamics are allowed us to in-depth analysis of lipotoxicity associated mechanistic network of cell death more precisely.
2022-07-16 | PXD029005 | Pride
Project description:The accumulation of lipid is a histological and biochemical hallmark of obesity-associated hepato-steatosis.Moreover, growing evidenceindicatesthathigher free fatty acids (FFAs) level in hepatocytes affects a myriad of biological processes leading to excessive metabolic imbalance, increased reactive oxygen species (ROS), deregulated autophagy, and impairment of mitochondrial and ER stress, that collectively drives cell death. Lipotoxicity and cell death mechanisms have been studied for many years. However, the molecular signals that link these two events during lipid stress remain poorly understood. From the very beginning, to systematically study hepato-lipotoxicity, HepG2 treated with PA providentially recapitulates the global lipotoxic responses, including insulin resistance to hepatocyte death. Therefore, using this cell-based model system, we pursued a comprehensive, differential quantitative approach where measurements of protein dynamics are analysed by mass spectrometry. Given that indispensable information, successive temporal phosphoproteomics dynamics are allowed us to in-depth analysis of lipotoxicity associated mechanistic network of cell death more precisely.
2022-07-16 | PXD029010 | Pride
Project description:To identify the genes induced by lipotoxicity and integrated stress response in pancreatic beta cells, we have employed whole genome microarray expression in panvreatic beta cell line MIN6m9 cells. First microarray analysis : Wild type MIN6m9 cells were treated with Palmitate, Oleate or negative control condition. Second microarray analysis : Wild type and Fv2E-PERK transgened MIN6m9 cells were treated with mock or artificial dimerizer AP20187(AP) in various time course.
2015-08-01 | E-GEOD-49647 | biostudies-arrayexpress
Project description:Successful early embryo development requires the correct reprogramming and configuration of gene networks by the timely and faithful execution of zygotic genome activation (ZGA). However, the regulatory principle of molecular elements and circuits fundamental to embryo development remains largely obscure. Here, we profiled the transcriptomes of single oocytes and blastomeres, obtained from in vitro fertilized (IVF) or parthenogenetically activated (PA) porcine early embryos (1- to 8-cell), focusing on the gene expression dynamics and regulatory networks associated with maternal-to-zygote transition (MZT) (e.g. maternal RNA clearance and ZGA). We found that minor and major ZGAs occur at 1-cell and 4-cell stages for both IVF and PA embryos, respectively. Maternal RNAs gradually decay from 1- to 8-cell embryos. Top abundantly expressed genes (CDV3, PCNA, CDR1, YWHAE, DNMT1, IGF2BP3, ARMC1, BTG4, UHRF2 and gametocyte-specific factor 1-like) in both IVF and PA early embryos identified are of vital roles for embryo development. Differently expressed genes within IVF groups are different from that within PA groups, indicating bi-parental and maternal-only embryos have specific sets of mRNAs distinctly decayed and activated. Pathways enriched from DEGs showed that RNA associated pathways (RNA biding, processing, transport and degradation) could be important. Moreover, mitochondrial RNAs were found to be actively transcribed and dynamically changed. DNA/H3K4 methylation and transcription factors also showed dynamic expression patterns. Taken together, our findings provide an important resource to investigate further the epigenetic and genome regulation of MZT events in early embryos of pigs.
2021-07-19 | GSE164812 | GEO
Project description:Metabolic adaptation enables cancer cell survival under nutrient stress, but how stromal cells in tumor microenvironment (TME) withstand similar metabolic change remains elusive. Here, we report that fatty acids (FAs), a nutrient with dual roles in energy provision and lipotoxicity, orchestrate a pro-metastatic adaptation program in cancer-associated fibroblasts (CAFs). In patients with oral squamous cell carcinoma (OSCC), the FA transporter CD36 was upregulated not only in cancer cells but also in myofibroblastic CAFs (myoCAFs), where it correlated with more advanced metastasis stage. Among 5 kinds of FAs enriched in OSCC tissues, palmitic acid (PA) potently activated myoCAFs phenotypes across 2D, 3D, and organoid co-culture models. In vivo, PA promoted lymph node metastasis in orthotopic tumors comprising OSCC cells and CAFs, an effect abolished by CD36 knockdown in CAFs. Mechanistically, PA engaged chromatin occupancy of H3K27ac at multiple genes including IRE1 and TMBIM6, two stress-adaptive regulators. Transcription of IRE1 and TMBIM6 in CAFs was regulated by CCAAT/enhancer-binding protein γ (CEBPG) through an enhancer-dependent manner. Disruption of the CEBPG–IRE1/TMBIM6 axis attenuated myoCAFs properties and abrogated PA-driven metastasis. Our results unveil a stromal metabolic checkpoint and establish CEBPG-enhanced stress resilience as a therapeutic target to curtail metastasis.
2025-12-26 | GSE314457 | GEO
Project description:Metabolic adaptation enables cancer cell survival under nutrient stress, but how stromal cells in tumor microenvironment (TME) withstand similar metabolic change remains elusive. Here, we report that fatty acids (FAs), a nutrient with dual roles in energy provision and lipotoxicity, orchestrate a pro-metastatic adaptation program in cancer-associated fibroblasts (CAFs). In patients with oral squamous cell carcinoma (OSCC), the FA transporter CD36 was upregulated not only in cancer cells but also in myofibroblastic CAFs (myoCAFs), where it correlated with more advanced metastasis stage. Among 5 kinds of FAs enriched in OSCC tissues, palmitic acid (PA) potently activated myoCAFs phenotypes across 2D, 3D, and organoid co-culture models. In vivo, PA promoted lymph node metastasis in orthotopic tumors comprising OSCC cells and CAFs, an effect abolished by CD36 knockdown in CAFs. Mechanistically, PA engaged chromatin occupancy of H3K27ac at multiple genes including IRE1 and TMBIM6, two stress-adaptive regulators. Transcription of IRE1 and TMBIM6 in CAFs was regulated by CCAAT/enhancer-binding protein γ (CEBPG) through an enhancer-dependent manner. Disruption of the CEBPG–IRE1/TMBIM6 axis attenuated myoCAFs properties and abrogated PA-driven metastasis. Our results unveil a stromal metabolic checkpoint and establish CEBPG-enhanced stress resilience as a therapeutic target to curtail metastasis.
2025-12-12 | GSE312946 | GEO
Project description:Background: Prolonged exposure to elevated free fatty acids induces β-cell failure (lipotoxicity) and contributes to the pathogenesis of type 2 diabetes. In vitro exposure of β-cells to the saturated free fatty acid palmitate is a valuable model of lipotoxicity, reproducing features of β-cell failure observed in type 2 diabetes. In order to map the β-cell response to lipotoxicity, we combined RNA-sequencing of palmitate-treated human islets with iTRAQ proteomics of insulin-secreting INS-1E cells following a time course exposure to palmitate. Results: Crossing transcriptome and proteome of palmitate-treated β-cells revealed 85 upregulated and 122 downregulated genes at both transcript and protein level. Pathway analysis identified lipid metabolism, oxidative stress, amino-acid metabolism and cell cycle pathways among the most enriched palmitate-modified pathways. Palmitate induced gene expression changes compatible with increased free fatty acid mitochondrial import and β-oxidation, decreased lipogenesis and modified cholesterol transport. Palmitate modified genes regulating endoplasmic reticulum (ER) function, ER-to-Golgi transport and ER stress pathways. Furthermore, palmitate modulated cAMP/protein kinase A (PKA) signaling, inhibiting expression of PKA anchoring proteins and downregulating the GLP-1 receptor. SLC7 family amino-acid transporters were upregulated in response to palmitate but this induction did not contribute to β-cell demise. To unravel critical mediators of lipotoxicity upstream of the palmitate-modified genes, we identified overrepresented transcription factor binding sites and performed network inference analysis. These identified LXR, PPARα, FOXO1 and BACH1 as key transcription factors orchestrating the metabolic and oxidative stress responses to palmitate. Conclusions: This is the first study to combine transcriptomic and sensitive time course proteomic profiling of palmitate-exposed β-cells. Our results provide comprehensive insight into gene and protein expression changes, corroborating and expanding beyond previous findings. The identification of critical drivers and pathways of the β-cell lipotoxic response points to novel therapeutic targets for type 2 diabetes.
2020-08-13 | GSE156109 | GEO