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Molecular & Cellular Proteomics : MCP Sep 2023Cholesteatoma is a chronic inflammatory ear disease with abnormal keratinized epithelium proliferation and tissue damage. However, the mechanism of keratinized...
The Hyperproliferation Mechanism of Cholesteatoma Based on Proteomics: SNCA Promotes Autophagy-Mediated Cell Proliferation Through the PI3K/AKT/CyclinD1 Signaling Pathway.
Cholesteatoma is a chronic inflammatory ear disease with abnormal keratinized epithelium proliferation and tissue damage. However, the mechanism of keratinized epithelium hyperproliferation in cholesteatoma remains unknown. Hence, our study sought to shed light on mechanisms affecting the pathology and development of cholesteatoma, which could help develop adjunctive treatments. To investigate molecular changes in cholesteatoma pathogenesis, we analyzed clinical cholesteatoma specimens and paired ear canal skin with mass spectrometry-based proteomics and bioinformatics. From our screen, alpha-synuclein (SNCA) was overexpressed in middle ear cholesteatoma and might be a key hub protein associated with inflammation, proliferation, and autophagy in cholesteatoma. SNCA was more sensitive to lipopolysaccharide-induced inflammation, and autophagy marker increase was accompanied by autophagy activation in middle ear cholesteatoma tissues. Overexpression of SNCA activated autophagy and promoted cell proliferation and migration, especially under lipopolysaccharide inflammatory stimulation. Moreover, inhibiting autophagy impaired SNCA-mediated keratinocyte proliferation and corresponded with inhibition of the PI3K/AKT/CyclinD1 pathways. Also, 740Y-P, a PI3K activator reversed the suppression of autophagy and PI3K signaling by siATG5 in SNCA-overexpressing cells, which restored proliferative activity. Besides, knockdown of SNCA in RHEK-1 and HaCaT cells or knockdown of PI3K in RHEK-1 and HaCaT cells overexpressing SNCA both resulted in attenuated cell proliferation. Our studies indicated that SNCA overexpression in cholesteatoma might maintain the proliferative ability of cholesteatoma keratinocytes by promoting autophagy under inflammatory conditions. This suggests that dual inhibition of SNCA and autophagy may be a promising new target for treating cholesteatoma.
Topics: Humans; Cholesteatoma, Middle Ear; Proto-Oncogene Proteins c-akt; Phosphatidylinositol 3-Kinases; Lipopolysaccharides; Proteomics; Signal Transduction; Cell Proliferation; Autophagy; Inflammation; alpha-Synuclein
PubMed: 37532176
DOI: 10.1016/j.mcpro.2023.100628 -
Carcinogenesis Dec 2023Potassium Calcium-Activated Channel Subfamily N1 (KCNN1), an integral membrane protein, is thought to regulate neuronal excitability by contributing to the slow...
Potassium Calcium-Activated Channel Subfamily N1 (KCNN1), an integral membrane protein, is thought to regulate neuronal excitability by contributing to the slow component of synaptic after hyperpolarization. However, the role of KCNN1 in tumorigenesis has been rarely reported, and the underlying molecular mechanism remains unclear. Here, we report that KCNN1 functions as an oncogene in promoting breast cancer cell proliferation and metastasis. KCNN1 was overexpressed in breast cancer tissues and cells. The pro-proliferative and pro-metastatic effects of KCNN1 were demonstrated by CCK8, clone formation, Edu assay, wound healing assay and transwell experiments. Transcriptomic analysis using KCNN1 overexpressing cells revealed that KCNN1 could regulate key signaling pathways affecting the survival of breast cancer cells. KCNN1 interacts with ERLIN2 and enhances the effect of ERLIN2 on Cyclin B1 stability. Overexpression of KCNN1 promoted the protein expression of Cyclin B1, enhanced its stability and promoted its K63 dependent ubiquitination, while knockdown of KCNN1 had the opposite effects on Cyclin B1. Knockdown (or overexpression) ERLNI2 partially restored Cyclin B1 stability and K63 dependent ubiquitination induced by overexpression (or knockdown) of KCNN1. Knockdown (or overexpression) ERLIN2 also partially neutralizes the effects of overexpression (or knockdown) KCNN1-induced breast cancer cell proliferation, migration and invasion. In paired breast cancer clinical samples, we found a positive expression correlations between KCNN1 and ERLIN2, KCNN1 and Cyclin B1, as well as ERLIN2 and Cyclin B1. In conclusion, this study reveals, for the first time, the role of KCNN1 in tumorigenesis and emphasizes the importance of KCNN1/ERLIN2/Cyclin B1 axis in the development and metastasis of breast cancer.
Topics: Female; Humans; Breast Neoplasms; Carcinogenesis; Cell Line, Tumor; Cell Proliferation; Cyclin B1; Gene Expression Regulation, Neoplastic; Membrane Proteins; Small-Conductance Calcium-Activated Potassium Channels; Ubiquitination
PubMed: 37831636
DOI: 10.1093/carcin/bgad070 -
International Journal of Oncology Aug 2023The pathogenesis mechanism of lung cancer is very complex, with high incidence and mortality. Serpin family A member 3 (SERPINA3) expression levels were reduced in the...
The pathogenesis mechanism of lung cancer is very complex, with high incidence and mortality. Serpin family A member 3 (SERPINA3) expression levels were reduced in the sera of patients with lung cancer and may be a candidate diagnostic and prognostic survival biomarker in lung cancer, as previously reported. However, the detailed biological functions of SERPINA3 in the pathogenesis of lung cancer remain unknown. In the present study, it was aimed to explore the effects of SERPINA3 on the occurrence of lung cancer. SERPINA3 expression was assessed using bioinformatics database analysis and experimental detection. Then, the biological effects of SERPINA3 were investigated in a cell culture system and a xenograft model of human lung cancer. The potential regulatory mechanism of SERPINA3 in lung cancer was explored by data‑independent acquisition mass spectrometry (DIA‑MS) detection and further validated by western blotting (WB). The results indicated that SERPINA3 expression levels were significantly downregulated in lung cancer tissues and cell lines. At the cellular level, it was revealed that overexpressed SERPINA3 inhibited cell growth, proliferation, migration and invasion and promoted the apoptosis of lung cancer cells. Moreover, overexpressed SERPINA3 enhanced the sensitivity of lung cancer cells to osimertinib. , a xenograft model of human lung cancer was established with BALB/c nude mice. After the injection of A549 cells, the tumor growth of the tumor‑bearing mice in the SERPINA3‑overexpressing group increased more slowly, and the tumor volume was smaller than that in the empty‑vector group. Mechanistically, a total of 65 differentially expressed proteins were identified. It was found that the speckle‑type POZ protein (SPOP) was significantly upregulated in SERPINA3‑overexpressing H157 cells using DIA‑MS detection and analysis. WB validation showed that SPOP expression increased, and NF‑kappaB (NF‑κB) p65 was inhibited in cell lines and tumor tissues of mice when SERPINA3 was overexpressed. The present findings suggest that SERPINA3 is involved in the development of lung cancer and has an antineoplastic role in lung cancer.
Topics: Humans; Animals; Mice; NF-kappa B; Mice, Nude; Cell Line, Tumor; Lung Neoplasms; Cell Proliferation; Cell Movement; Gene Expression Regulation, Neoplastic; Serpins
PubMed: 37417362
DOI: 10.3892/ijo.2023.5544 -
Lipids in Health and Disease Oct 2023Ceramide, a bioactive signaling sphingolipid, has long been implicated in cancer. Members of the ceramide synthase (CerS) family determine the acyl chain lengths of...
BACKGROUND
Ceramide, a bioactive signaling sphingolipid, has long been implicated in cancer. Members of the ceramide synthase (CerS) family determine the acyl chain lengths of ceramides, with ceramide synthase 4 (CerS4) primarily generating C18-C20-ceramide. Although CerS4 is known to be overexpressed in breast cancer, its role in breast cancer pathogenesis is not well established.
METHODS
To investigate the role of CerS4 in breast cancer, public datasets, including The Cancer Genome Atlas (TCGA) and two Gene Expression Omnibus (GEO) datasets (GSE115577 and GSE96058) were analyzed. Furthermore, MCF-7 cells stably overexpressing CerS4 (MCF-7/CerS4) as a model for luminal subtype A (LumA) breast cancer were produced, and doxorubicin (also known as Adriamycin [AD])-resistant MCF-7/ADR cells were generated after prolonged treatment of MCF-7 cells with doxorubicin. Kaplan-Meier survival analysis assessed the clinical significance of CERS4 expression, while Student's t-tests or Analysis of Variance (ANOVA) compared gene expression and cell viability in different MCF-7 cell lines.
RESULTS
Analysis of the public datasets revealed elevated CERS4 expression in breast cancer, especially in the most common breast cancer subtype, LumA. Persistent CerS4 overexpression in MCF-7 cells activated multiple cancer-associated pathways, including pathways involving sterol regulatory element-binding protein, nuclear factor kappa B (NF-κB), Akt/mammalian target of rapamycin (mTOR), and β-catenin. Furthermore, MCF-7/CerS4 cells acquired doxorubicin, paclitaxel, and tamoxifen resistance, with concomitant upregulation of ATP-binding cassette (ABC) transporter genes, such as ABCB1, ABCC1, ABCC2, ABCC4, and ABCG2. MCF-7/CerS4 cells were characterized by increased cell migration and epithelial-mesenchymal transition (EMT). Finally, CERS4 knockdown in doxorubicin-resistant MCF-7/ADR cells resulted in reduced activation of cancer-associated pathways (NF-κB, Akt/mTOR, β-catenin, and EMT) and diminished chemoresistance, accompanied by ABCB1 and ABCC1 downregulation.
CONCLUSIONS
Chronic CerS4 overexpression may exert oncogenic effects in breast cancer via alterations in signaling, EMT, and chemoresistance. Therefore, CerS4 may represent an attractive target for anticancer therapy, especially in LumA breast cancer.
Topics: Female; Humans; ATP-Binding Cassette Transporters; beta Catenin; Breast Neoplasms; Doxorubicin; Drug Resistance, Neoplasm; NF-kappa B; Proto-Oncogene Proteins c-akt; TOR Serine-Threonine Kinases; Sphingosine N-Acyltransferase; MCF-7 Cells
PubMed: 37885013
DOI: 10.1186/s12944-023-01930-z -
Current Genetics Feb 2020Prions are self-propagating protein isoforms that are typically amyloid. In Saccharomyces cerevisiae, amyloid prion aggregates are fragmented by a trio involving three... (Review)
Review
Prions are self-propagating protein isoforms that are typically amyloid. In Saccharomyces cerevisiae, amyloid prion aggregates are fragmented by a trio involving three classes of chaperone proteins: Hsp40s, also known as J-proteins, Hsp70s, and Hsp104. Hsp104, the sole Hsp100-class disaggregase in yeast, along with the Hsp70 Ssa and the J-protein Sis1, is required for the propagation of all known amyloid yeast prions. However, when Hsp104 is ectopically overexpressed, only the prion [PSI] is efficiently eliminated from cell populations via a highly debated mechanism that also requires Sis1. Recently, we reported roles for two additional J-proteins, Apj1 and Ydj1, in this process. Deletion of Apj1, a J-protein involved in the degradation of sumoylated proteins, partially blocks Hsp104-mediated [PSI] elimination. Apj1 and Sis1 were found to have overlapping functions, as overexpression of one compensates for loss of function of the other. In addition, overexpression of Ydj1, the most abundant J-protein in the yeast cytosol, completely blocks Hsp104-mediated curing. Yeast prions exhibit structural polymorphisms known as "variants"; most intriguingly, these J-protein effects were only observed for strong variants, suggesting variant-specific mechanisms. Here, we review these results and present new data resolving the domains of Apj1 responsible, specifically implicating the involvement of Apj1's Q/S-rich low-complexity domain.
Topics: Animals; Fungal Proteins; HSP40 Heat-Shock Proteins; Humans; Molecular Chaperones; Prions; Protein Binding; Protein Interaction Domains and Motifs
PubMed: 31230108
DOI: 10.1007/s00294-019-01006-5 -
Biomolecules Apr 2022Human S100B is a small, multifunctional protein. Its activity, inside and outside cells, contributes to the biology of the brain, muscle, skin, and adipocyte tissues....
Human S100B is a small, multifunctional protein. Its activity, inside and outside cells, contributes to the biology of the brain, muscle, skin, and adipocyte tissues. Overexpression of S100B occurs in Down Syndrome, Alzheimer's disease, Creutzfeldt-Jakob disease, schizophrenia, multiple sclerosis, brain tumors, epilepsy, melanoma, myocardial infarction, muscle disorders, and sarcopenia. Modulating the activities of S100B, related to human diseases, without disturbing its physiological functions, is vital for drug and therapy design. This work focuses on the extracellular activity of S100B and one of its receptors, the Receptor for Advanced Glycation End products (RAGE). The functional outcome of extracellular S100B, partially, depends on the activation of intracellular signaling pathways. Here, we used Biotin Switch Technique enrichment and mass-spectrometry-based proteomics to show that the appearance of the S100B protein in the extracellular milieu of the mammalian Chinese Hamster Ovary (CHO) cells, and expression of the membrane-bound RAGE receptor, lead to changes in the intracellular S-nitrosylation of, at least, more than a hundred proteins. Treatment of the wild-type CHO cells with nanomolar or micromolar concentrations of extracellular S100B modulates the sets of S-nitrosylation targets inside cells. The cellular S-nitrosome is tuned differently, depending on the presence or absence of stable RAGE receptor expression. The presented results are a proof-of-concept study, suggesting that S-nitrosylation, like other post-translational modifications, should be considered in future research, and in developing tailored therapies for S100B and RAGE receptor-related diseases.
Topics: Animals; CHO Cells; Cricetinae; Cricetulus; Humans; Protein S; Receptor for Advanced Glycation End Products; Receptors, Immunologic; S100 Calcium Binding Protein beta Subunit
PubMed: 35625541
DOI: 10.3390/biom12050613 -
Journal of Thoracic Disease Jan 2021The sterile alpha motif (SAM) domain and histidine-aspartate (HD) domain-containing protein 1 (SAMHD1) has been specifically linked to lung cancer. However, the...
BACKGROUND
The sterile alpha motif (SAM) domain and histidine-aspartate (HD) domain-containing protein 1 (SAMHD1) has been specifically linked to lung cancer. However, the underlying mechanisms in regulating lung adenocarcinoma (LAC) are unclear. The aim of this study was to assess the specific regulation between SAMHD1 and LAC.
METHODS
We retrospectively reviewed 238 patients who underwent surgery for LAC between January 2018 and December 2019. The expression of SAMHD1 was detected by quantitative reverse-transcription polymerase chain reaction (RT-qPCR) in tumors and paired adjacent tissues. A lentivirus was used to overexpress SAMHD1 and stimulator of interferon genes (STING) in A549 cells; and RT-qPCR and western blot analysis were performed to verify their levels. Cell proliferation was evaluated via 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and Celigo imaging cytometry. Cell apoptosis was detected by Annexin V staining. Overexpressed SAMHD1 suppressed LAC progression in a xenograft model. The DNA damage response inhibitor (DDRi) was used to assess the cell proliferation and apoptosis rate in SAMHD1-overexpressing A549 cells and the control group. A rescue experiment was carried out to evaluate the potential influence of SAMHD1 and STING.
RESULTS
A low expression of SAMHD1 was associated with advanced disease. Overexpression of SAMHD1 decreased cell proliferation and invasion in A549 cells, and the apoptosis rate was significantly higher in the overexpressed SAMHD1 cells than those in the control group. The overexpression of SAMHD1 inhibited tumor progression in the xenograft model. The expression of STING was lower in SAMHD1-overexpressing A549 cells than those in the wild-type group. Furthermore, the inhibited cellular behaviors of LAC cells resulting from the stable SAMHD1 expression were partially reversed after STING overexpression. Treatment with DDRi could inhibit cancer cell progression.
CONCLUSIONS
Upregulation of SAMHD1 could suppress the progression of LAC and through the negative regulation of STING.
PubMed: 33569199
DOI: 10.21037/jtd-20-1889 -
Hepatology Communications Dec 2023Liver cancer is increasing due to the rise in metabolic dysfunction-associated steatohepatitis (MASH). High-mobility group box-1 (HMGB1) is involved in the pathogenesis...
BACKGROUND
Liver cancer is increasing due to the rise in metabolic dysfunction-associated steatohepatitis (MASH). High-mobility group box-1 (HMGB1) is involved in the pathogenesis of chronic liver disease, but its role in MASH-associated liver cancer is unknown. We hypothesized that an increase in hepatocyte-derived HMGB1 in a mouse model of inactivation of PTEN that causes MASH could promote MASH-induced tumorigenesis.
METHODS
We analyzed publicly available transcriptomics datasets, and to explore the effect of overexpressing HMGB1 in cancer progression, we injected 1.5-month-old Pten∆Hep mice with adeno-associated virus serotype-8 (AAV8) vectors to overexpress HMGB1-EGFP or EGFP, and sacrificed them at 3, 9 and 11 months of age.
RESULTS
We found that HMGB1 mRNA increases in human MASH and MASH-induced hepatocellular carcinoma (MASH-HCC) compared to healthy livers. Male and female Pten∆Hep mice overexpressing HMGB1 showed accelerated liver tumor development at 9 and 11 months, respectively, with increased tumor size and volume, compared to control Pten∆Hep mice. Moreover, Pten∆Hep mice overexpressing HMGB1, had increased incidence of mixed HCC-intrahepatic cholangiocarcinoma (iCCA). All iCCAs were positive for nuclear YAP and SOX9. Male Pten∆Hep mice overexpressing HMGB1 showed increased cell proliferation and F4/80+ cells at 3 and 9 months.
CONCLUSION
Overexpression of HMGB1 in hepatocytes accelerates liver tumorigenesis in Pten∆Hep mice, enhancing cell proliferation and F4/80+ cells to drive MASH-induced liver cancer.
Topics: Animals; Female; Humans; Infant; Male; Mice; Bile Duct Neoplasms; Bile Ducts, Intrahepatic; Carcinogenesis; Carcinoma, Hepatocellular; Fatty Liver; Hepatocytes; HMGB1 Protein; Liver Neoplasms; PTEN Phosphohydrolase
PubMed: 38055645
DOI: 10.1097/HC9.0000000000000311 -
Frontiers in Bioscience (Landmark... Aug 2023The role of Pin2 telomeric repeat factor 1-interacting telomerase inhibitor 1 (PinX1) in tumorigenesis and development has been extensively studied. As we previously...
BACKGROUND
The role of Pin2 telomeric repeat factor 1-interacting telomerase inhibitor 1 (PinX1) in tumorigenesis and development has been extensively studied. As we previously demonstrated, PinX1 plays an important role in modulating epithelial-mesenchymal transition (EMT), stemness, cell proliferation, and apoptosis in nasopharyngeal carcinoma (NPC). However, the relationship between PinX1, autophagy, and cell function in NPC remains unclear. This study aimed to investigate the mechanisms by which PinX1 regulates autophagy in NPC, and to explore its biological role and clinical significance in disease progression.
METHODS
The proliferative capacity of NPC cells was assessed by MTT and xenograft tumorigenicity assays. Autophagic flux was monitored using a tandem monomeric DAPI-FITC-LC3 reporter assay. The rates of apoptosis and the cell cycle in NPC cells were analyzed using flow cytometry. The activation of autophagy and the signaling status of the AKT/mTOR and NF-κB/p65 pathways were evaluated by Western blot analysis.
RESULTS
In addition to promoting autophagy and apoptosis, PinX1 overexpression suppressed proliferation, migration, invasion, and decelerated cell-cycle progression in NPC cells. These effects were reversed by inhibiting autophagy with 3-methyladenine. Mechanistic investigations clarified that PinX1 overexpression significantly reduced the expression of p-AKT, p-mTOR, p65, and p-p65. Chloroquine treatment in PinX1-overexpressing cells did not significantly alter p-AKT and p-mTOR levels, whereas 3-MA treatment in PinX1-overexpressing cells resulted in increased p65 and p-p65 expression, relative to untreated PinX1-overexpressing cells.
CONCLUSIONS
It appears that PinX1 promotes autophagy by inhibiting the AKT/mTOR signaling pathway, which then inhibits NF-κB/p65 pathways, and consequently inhibiting cell proliferation and causing cell apoptosis in NPC cells.
Topics: Humans; Apoptosis; Autophagy; Cell Proliferation; Nasopharyngeal Carcinoma; Nasopharyngeal Neoplasms; NF-kappa B; Proto-Oncogene Proteins c-akt; Signal Transduction; TOR Serine-Threonine Kinases; Animals
PubMed: 37664923
DOI: 10.31083/j.fbl2808170 -
Cellular & Molecular Biology Letters Aug 2023Desmocollin-1 (DSC1) is a desmosomal transmembrane glycoprotein that maintains cell-to-cell adhesion. DSC1 was previously associated with lymph node metastasis of...
BACKGROUND
Desmocollin-1 (DSC1) is a desmosomal transmembrane glycoprotein that maintains cell-to-cell adhesion. DSC1 was previously associated with lymph node metastasis of luminal A breast tumors and was found to increase migration and invasion of MCF7 cells in vitro. Therefore, we focused on DSC1 role in cellular and molecular mechanisms in luminal A breast cancer and its possible therapeutic modulation.
METHODS
Western blotting was used to select potential inhibitor decreasing DSC1 protein level in MCF7 cell line. Using atomic force microscopy we evaluated effect of DSC1 overexpression and modulation on cell morphology. The LC-MS/MS analysis of total proteome on Orbitrap Lumos and RNA-Seq analysis of total transcriptome on Illumina NextSeq 500 were performed to study the molecular mechanisms associated with DSC1. Pull-down analysis with LC-MS/MS detection was carried out to uncover DSC1 protein interactome in MCF7 cells.
RESULTS
Analysis of DSC1 protein levels in response to selected inhibitors displays significant DSC1 downregulation (p-value ≤ 0.01) in MCF7 cells treated with NF-κB inhibitor parthenolide. Analysis of mechanic cell properties in response to DSC1 overexpression and parthenolide treatment using atomic force microscopy reveals that DSC1 overexpression reduces height of MCF7 cells and conversely, parthenolide decreases cell stiffness of MCF7 cells overexpressing DSC1. The LC-MS/MS total proteome analysis in data-independent acquisition mode shows a strong connection between DSC1 overexpression and increased levels of proteins LACRT and IGFBP5, increased expression of IGFBP5 is confirmed by RNA-Seq. Pathway analysis of proteomics data uncovers enrichment of proliferative MCM_BIOCARTA pathway including CDK2 and MCM2-7 after DSC1 overexpression. Parthenolide decreases expression of LACRT, IGFBP5 and MCM_BIOCARTA pathway specifically in DSC1 overexpressing cells. Pull-down assay identifies DSC1 interactions with cadherin family proteins including DSG2, CDH1, CDH3 and tyrosine kinase receptors HER2 and HER3; parthenolide modulates DSC1-HER3 interaction.
CONCLUSIONS
Our systems biology data indicate that DSC1 is connected to mechanisms of cell cycle regulation in luminal A breast cancer cells, and can be effectively modulated by parthenolide.
Topics: Chromatography, Liquid; Desmocollins; Neoplasms; Proteome; Tandem Mass Spectrometry; Humans; MCF-7 Cells; Sesquiterpenes
PubMed: 37620794
DOI: 10.1186/s11658-023-00481-6