scholarly journals Phosphatidylcholine Synthesis Influences the Diacylglycerol Homeostasis Required for Sec14p-dependent Golgi Function and Cell Growth

2001 ◽  
Vol 12 (3) ◽  
pp. 511-520 ◽  
Author(s):  
Annette L. Henneberry ◽  
Thomas A. Lagace ◽  
Neale D. Ridgway ◽  
Christopher R. McMaster
Keyword(s):  
Cell Death ◽  
Cell Growth ◽  
Phosphatidic Acid ◽  
Metabolic Pathways ◽  
Eukaryotic Cells ◽  
Kennedy Pathway ◽  
Phosphatidylcholine Synthesis ◽  
Long Chain

Phosphatidylcholine and phosphatidylethanolamine are the most abundant phospholipids in eukaryotic cells and thus have major roles in the formation and maintenance of vesicular membranes. In yeast, diacylglycerol accepts a phosphocholine moiety through aCPT1-derived cholinephosphotransferase activity to directly synthesize phosphatidylcholine. EPT1-derived activity can transfer either phosphocholine or phosphoethanolamine to diacylglcyerol in vitro, but is currently believed to primarily synthesize phosphatidylethanolamine in vivo. In this study we report that CPT1- and EPT1-derived cholinephosphotransferase activities can significantly overlap in vivo such that EPT1 can contribute to 60% of net phosphatidylcholine synthesis via the Kennedy pathway. Alterations in the level of diacylglycerol consumption through alterations in phosphatidylcholine synthesis directly correlated with the level of SEC14-dependent invertase secretion and affected cell viability. Administration of synthetic di8:0 diacylglycerol resulted in a partial rescue of cells fromSEC14-mediated cell death. The addition of di8:0 diacylglycerol increased di8:0 diacylglycerol levels 20–40-fold over endogenous long-chain diacylglycerol levels. Di8:0 diacylglcyerol did not alter endogenous phospholipid metabolic pathways, nor was it converted to di8:0 phosphatidic acid.

Synergistic and selective inhibition of NSCLC cell growth via a caspase-independent cell death pathway by tumor suppressor 101F6 nanoparticles plus vitamin C in vitro and in vivo

2006 ◽  
Vol 24 (18_suppl) ◽  
pp. 13086-13086
Author(s):  
S. Ohtani ◽  
K. Ueda ◽  
G. Jayanchandran ◽  
K. Xu ◽  
J. D. Minna ◽  
...  
Keyword(s):  
Cell Death ◽  
Cell Growth ◽  
Tumor Suppressor ◽  
Vitamin C ◽  
Tumor Cells ◽  
Nsclc Cell ◽  
Normal Lung ◽  
Human Nsclc

13086 Background: 101F6 is a candidate tumor suppressor gene on chromosome 3p21.3, a site of allele loss and genomic alterations were frequently found in many human cancers. We previously showed that enforced expression of wt-101F6 by adenoviral virus significantly inhibited tumor cell growth in 3p21.3-deficient NSCLC cells in vitro and in vivo. How 101F6 exerts this effect is largely unknown. Using a computer-aided structural and functional modeling, we recently identified 101F6 as a member of cytochrome b-561 protein family, which is involved in the regeneration of vitamin C. We hypothesized that under normal physiologic conditions, 101F6 protects cells from oxidative damage by regenerating antioxidant vitamin C and that in 101F6-deficient tumor cells, exogenous 101F6 facilitates vitamin C-mediated cytotoxic H2O2 formation. Methods and Results: We examined endogenous 101F6 expression in human NSCLC cell lines and tissue samples. All normal lung bronchial epithelial cells and fibroblasts but few lung cancers expressed 101F6. We investigated the combined effect of 101F6 and vitamin C on the cell growth: a nanoparticle-mediated wt-101F6 gene transfer plus a sub-pharmacologic concentration of vitamin C synergistically inhibited 3p21.3-deficient NSCLC cell growth but did not affect normal cell growth. We also used a human NSCLC H322 orthotopic lung tumor xenograft mouse model to evaluate the therapeutic efficacy of systemic injection of 101F6 nanoparticles and intraperitoneal injection of vitamin C. The growth of lung tumors was synergistically inhibited by the combination treatment (p<0.001). Furthermore, exogenous 101F6 promoted intracellular vitamin C uptake, leading to the vitamin C-mediated accumulation of H2O2 in the tumor cells, and these two agents synergistically killed the cells through caspase-independent apoptosis and autophagy cell death pathways. Conclusions: The synergistic and selective antitumor effect of 101F6 nanoparticles plus vitamin C may offer a useful tool for lung cancer prevention and intervention. This abstract is supported by grants from NCI (SPORE P50CA70907) and DOD (TARGET, DAMD17002–1-0706). No significant financial relationships to disclose.

scholarly journals C1orf109L promote R-loop accumulation induced DNA damage to inhibit cell growth

2019 ◽  
Author(s):  
Dou Peng ◽  
Li Yiqun ◽  
Xie Wanqiu ◽  
Zhang Xiaoqing ◽  
Zhang Dandan ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Dna Damage ◽  
Cell Death ◽  
Cell Growth ◽  
Genome Instability ◽  
Cell Death Pathway ◽  
Unknown Gene ◽  
Lower Expression

AbstractAs a function unknown gene, C1orf109 is lower expression in various cells. Here, we reported that C1orf109L, the longest variant of C1orf109, which interacted with R-loop-regulating proteins to trigger R-loop, a three-stranded nucleic acid structure frequently mediated genome instability, accumulation. C1orf109L induce chronic DNA damage to promote P21 upregulation and strongly inhibits cell growth in vitro and in vivo by arresting the cell cycle in the G2 phase. With camptothecin (CPT), an R-loop activator, treatment, C1orf109L further triggers R-loop accumulation-induced DNA damage and promotes cell death by activating cell-death pathway. Furthermore, CPT treatment increases C1orf109L ubiquitination and turnover, which inhibits cell death and promotes the G0/G1 phase of the cell cycle. Therefore, our data illustrated the mechanisms underlying C1orf109L-related cell growth inhibition and provide feasibility and limitations for C1orf109L as a potential target for cancer therapy.

CSIG-22. Cd95l/Cd95 GENE DELETION DECREASES MOUSE GLIOMA TUMORIGENESIS THROUGH THE INHIBITION OF NON-CANONICAL CD95L/CD95-MEDIATED CELL GROWTH AND IMMUNOSUPPRESSION

2021 ◽  
Vol 23 (Supplement_6) ◽  
pp. vi37-vi38
Author(s):  
Clara Quijano-Rubio ◽  
Michael Weller
Keyword(s):  
Cell Death ◽  
Cell Growth ◽  
Cell Lines ◽  
Cancer Cell ◽  
Apoptotic Cell ◽  
Immune Effector ◽  
Immunodeficient Mice ◽  
Murine Glioma

Abstract CD95 is a transmembrane receptor with potential to promote both cell death and growth. Initially described to trigger apoptosis upon ligand (CD95L) engagement, CD95 may also prompt cell proliferation, invasion and stemness. CD95 stimulation to induce cancer cell apoptosis has been proved clinically impracticable. However, in tumors expressing both CD95 and CD95L, strategically inhibiting CD95-CD95L interactions to simultaneously block cancer cell growth and apoptotic cell death in tumor microenvironment components, including CD95-expressing antitumor immune effector cells, may represent an alternative therapeutic strategy. Here we characterized the expression of CD95 and CD95L in murine glioma models in vitro and in vivo. To fully disrupt CD95-CD95L interactions, we deleted Cd95 or Cd95l by CRISPR-Cas9-mediated knockout (KO) and assessed the consequences on cell growth and tumorigenicity in immunocompetent and immunocompromised mice. CD95 expression was identified in selected murine glioma cell lines. In vitro, expression of the canonical, membrane-bound, form of CD95L was not detected but cell lines expressed a shorter non-canonical, soluble, Cd95l variant. Tumors generated upon implantation of the same cells in vivo expressed both Cd95l variants. Upon Cd95l KO, all investigated cell lines exhibited reduced growth in vitro. Cell growth reduction upon Cd95 KO in SMA-497 murine glioma cells was rescued upon Cd95 re-transfection, validating CD95 specificity of the phenotype. Cd95-overexpression in Cd95-expressing cells did not increase growth. In vivo, Cd95 or Cd95l KO cell implantation in syngeneic mice generated smaller tumors than wildtype cells, resulting in prolonged survival. While 40% Cd95l KO cell-implanted immunocompetent mice did not develop tumors, all immunodeficient mice did. Altogether, these data reveal a growth-promoting role of non-canonical CD95L-CD95 interactions in murine gliomas, which blockade through gene KO results in decreased tumorigenicity. Furthermore, our data suggest the contribution of CD95L-mediated immunosuppression to the reduction of Cd95l KO-associated tumorigenicity.

GRN163L, a Novel and Potent Telomerase Inhibitor, Inhibits Myeloma Cell Growth In Vitro and In Vivo.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 639-639
Author(s):  
Masood A. Shammas ◽  
Hemanta Koley ◽  
Pierfrancesco Tassone ◽  
Paola Neri ◽  
Alexei Protopopov ◽  
...  
Keyword(s):  
Cell Death ◽  
Cell Growth ◽  
Apoptotic Cell ◽  
Myeloma Cell ◽  
Telomerase Activity ◽  
Hsp90 Inhibitor ◽  
Telomere Shortening ◽  
Myeloma Cells

Abstract Telomerase activity is either low or completely absent in most normal somatic cells; while it is elevated in most cancer cells providing unlimited proliferative potential by preventing telomere shortening. The inhibitors of telomerase, therefore, induce telomere shortening leading to apoptotic cell death in tumor cells while having little or no effect on normal diploid cells. We have evaluated the in vitro and in vivo efficacy of thio-phosphoramidate oligonucleotide specifically targeting the RNA component of telomerase (GRN163L) with demonstrated nuclear uptake by &gt;99% cells without the transfection enhancer. Delivery of GRN163L (1 μM) to MM cells (INA6 and ARP) was specifically associated with complete loss of telomerase activity as early as 6 hrs following exposure and was accompanied by a reduction in myeloma cell growth and survival. Treatment of INA6 cells with GRN163L for three weeks induced 96±4% and 100% cell death at 0.5 and 1 μM concentrations, respectively. ARP cells, which express higher levels of telomerase activity and have longer telomeres, showed 67±4% cell death at 5 weeks with 0.5 μM inhibitor and 82±3% and 100% cell death at 4 and 5 weeks, respectively, with 2 μM GRN163L. The apoptotic cell death was confirmed in 51% INA6 cells at two weeks and in &gt;80% ARP cells at four weeks. Apoptosis was associated with reduction in mean Telomere Fluorescence Intensity (TFI) on interphase chromosomes from 87.1±6.2 in control oligo treated INA6 cells to 36.2±2 (2.4 fold) in GRN163L treated cells. Moreover, GRN163L treatment was also associated with a similar reduction in number of chromosomes with detectable telomeres, indicating development of telomere-free ends. We have confirmed in vivo efficacy of GRN163L in a SCID-hu murine model of multiple myeloma. Following growth of GFP-transduced myeloma cells in the fetal bone chip introduced into the mice, GRN163L was injected on alternate days. In two independent experiments significant reduction in tumor cell growth, as measured by reduction in human myeloma related protein, and better survival than mice injected with control oligo was observed. We have now evaluated efficacy of combination of GRN163L with other novel agents. We have observed synergistic activity with Hsp90 inhibitor 17AAG on myeloma cell death. Addition of 17AAG (0.05 μM) to myeloma cells pre-treated with GRN163L (1 μM) for one week led to complete growth arrest within four days compared to continued growth of cells not pre-treated with GRN-163. These data provide the preclinical rationale for clinical evaluation of GRN163L in myeloma and in combination with Hsp90 inhibitor.

scholarly journals EXTH-44. INHIBITION OF MerTK ACTIVATES GLIOBLASTOMA-ASSOCIATED MACROPHAGES AND INDUCES TUMOR CELL DEATH IN GLIOMA MICROENVIRONMENT

2019 ◽  
Vol 21 (Supplement_6) ◽  
pp. vi91-vi91
Author(s):  
Yu-Ting Su ◽  
Madison Butler ◽  
Lee Hwang ◽  
Dragan Maric ◽  
Shelton Earp ◽  
...  
Keyword(s):  
Cell Death ◽  
Cell Growth ◽  
Tumor Microenvironment ◽  
Tumor Cell ◽  
Colony Formation ◽  
Glioblastoma Cells ◽  
Tumor Cell Death ◽  
Glioma Microenvironment

Abstract BACKGROUND Glioblastoma-associated macrophages and microglia (GAMs) are the predominant immune cells in the tumor microenvironment. Activation of MerTK, a receptor tyrosine kinase, triggers efferocytosis and polarizes GAMs to an immunosuppressive phenotype, promoting glioma growth. Our previous findings showed that UNC2371, a small-molecule inhibitor of MerTK, induced a less immunosuppressive phenotype of GAMs. Here, we investigate the role of MerTK inhibition on glioblastoma cells in the tumor microenvironment in vitro and in vivo. METHODS Cytotoxicity of UNC2371 in glioblastoma cells was determined by cell viability and colony formation assays. The protein expression of MerTK, AKT, and Erk were quantified by Western blotting in UNC2371-treated glioblastoma cells. A syngeneic GL261 mouse orthotopic glioblastoma model was used to evaluate the survival benefit of UNC2371 treatment. Fluorescent multiplex immunohistochemistry (IHC) was used to evaluate the expression of CD206, an anti-inflammatory marker on GAMs in murine brain tumor tissues. RESULTS UNC2371 inhibited GBM cell growth with an EC50 < 100 nM in both human U251 and mouse GL261 glioma cells, but not in GAMs. UNC2371-induced cell death and decreased cell proliferation were demonstrated by colony formation assays. UNC2371 decreased protein expression of phosphorylated MerTK, AKT, and Erk, which are essential for cell survival signaling, in U251 and GL261 cells. Furthermore, UNC2371 treatment prolonged survival in the mouse orthotopic GL261 glioblastoma model, suggesting that UNC2371 induces glioma cell death. A decreased of CD206+ GAMs was found in mice glioma tissues by fluorescent multiplex IHC, consistent with our previous findings in the in vitro cell-based assays. These data suggest that in addition to alleviate immunosuppression in the glioma microenvironment, UNC2371 directly inhibits GBM cell growth in vitro and in vivo. CONCLUSION Our findings suggest that UNC2371 has a therapeutic benefit via promoting GAM polarization towards proinflammatory status in the glioblastoma microenvironment and unexpectedly, inducing tumor cell death.

scholarly journals Oxidative Medicine and Cellular Longevity Hsa_circ_0013731 mediated by E2F1 inhibits ferroptosis in hepatocellular carcinoma cells by sponging miR-877-3p and targeting SLC7A11

2021 ◽  
Author(s):  
Shiji Fang ◽  
Weiqian Chen ◽  
Jiayi Ding ◽  
Dengke Zhang ◽  
Liyun Zheng ◽  
...  
Keyword(s):  
Hepatocellular Carcinoma ◽  
Cell Death ◽  
Cell Growth ◽  
Vital Role ◽  
Expression Level ◽  
Luciferase Reporter ◽  
Circular Rnas ◽  
Regulatory Loop

Abstract Background: The regulatory loop between circular RNAs and microRNAs has a vital role in cell death. Ferroptosis is the form of iron-dependent cell death, which is distinct from necroptosis and apoptosis. Increasing evidences showed that ferroptosis is an important form of cell death in hepatocellular carcinoma.Methods: Real-time PCR were used to examine the expression level of circ_0013731 in hepatocellular carcinoma tissues. Edu and colony formation were performed to detect the cell proliferation. A luciferase reporter assay was used to determine the relationship between circ_0013731, miR-877-3p and SLC7A11. ChIP-qPCR assays were performed to examine the potential binding of E2F1 to the circ_0013731 promoter. Iron Assay Kit (Sigma Aldrich) was used to detect total iron or Fe2+. C11 BODIPY 581/591 staining and flow cytometer were used to examine the Lipid ROS. The role of circ_0013731 was examined in xenograft tumors model. Results: We revealed that the expression level of circ_0013731 was elevated in hepatocellular carcinoma. Moreover, E2F1 promote the transcription of circ_0013731. Overexpression of circ_0013731 promoted cell growth and inhibited ferroptosis in SMMC-7721 and QGY-7703 in vitro. miR-877-3p was proved as the direct target of circ_0013731. Then, inhibition of miR-877-3p enhanced cell growth and inhibited ferroptosis. Further mechanism research demonstrated that circ_0013731 upregulated the expression level of SLC7A11 by sponging miR-877-3p. Finally, circ_0013731 promoted HCC growth via miR-877-3p/ SLC7A11 axis in vivo.Conclusions: Our data reveal that circ_0013731 mediated by E2F1 facilitates cell growth and suppressed the ferroptosis via miR-877-3p/ SLC7A11 axis in hepatocellular carcinoma. Therefore, circ_0013731 could be acted as potential therapeutical target for hepatocellular carcinoma.

Hsa_circ_0013731 Mediated by E2F1 Inhibits Ferroptosis in Hepatocellular Carcinoma Cells by Sponging miR-877-3p and Targeting SLC7A11

2020 ◽  
Author(s):  
Zhongwei Zhao ◽  
Jingjing Song ◽  
Dengke Zhang ◽  
Fazong Wu ◽  
Jianfei Tu ◽  
...  
Keyword(s):  
Hepatocellular Carcinoma ◽  
Cell Death ◽  
Cell Growth ◽  
Vital Role ◽  
Expression Level ◽  
Luciferase Reporter ◽  
Circular Rnas ◽  
Regulatory Loop

Abstract Background: The regulatory loop between circular RNAs and microRNAs has a vital role in cell death. Ferroptosis is the form of iron-dependent cell death, which is distinct from necroptosis and apoptosis. Increasing evidences showed that ferroptosis is an important form of cell death in hepatocellular carcinoma.Methods: Real-time PCR were used to examine the expression level of circ_0013731 in hepatocellular carcinoma tissues. Edu and colony formation were performed to detect the cell proliferation. A luciferase reporter assay was used to determine the relationship between circ_0013731, miR-877-3p and SLC7A11. ChIP-qPCR assays were performed to examine the potential binding of E2F1 to the circ_0013731 promoter. Iron Assay Kit (Sigma Aldrich) was used to detect total iron or Fe2+. C11 BODIPY 581/591 staining and flow cytometer were used to examine the Lipid ROS. The role of circ_0013731 was examined in xenograft tumors model. Results: We revealed that the expression level of circ_0013731 was elevated in hepatocellular carcinoma. Moreover, E2F1 promote the transcription of circ_0013731. Overexpression of circ_0013731 promoted cell growth and inhibited ferroptosis in SMMC-7721 and QGY-7703 in vitro. miR-877-3p was proved as the direct target of circ_0013731. Then, inhibition of miR-877-3p enhanced cell growth and inhibited ferroptosis. Further mechanism research demonstrated that circ_0013731 upregulated the expression level of SLC7A11 by sponging miR-877-3p. Finally, circ_0013731 promoted HCC growth via miR-877-3p/ SLC7A11 axis in vivo.Conclusions: Our data reveal that circ_0013731 mediated by E2F1 facilitates cell growth and suppressed the ferroptosis via miR-877-3p/ SLC7A11 axis in hepatocellular carcinoma. Therefore, circ_0013731 could be acted as potential therapeutical target for hepatocellular carcinoma.

scholarly journals In Vivo Assay Reveals Microbial OleA Thiolases Initiating Hydrocarbon and β-Lactone Biosynthesis

mBio ◽  
2020 ◽  
Vol 11 (2) ◽  
Author(s):  
Megan D. Smith ◽  
Serina L. Robinson ◽  
Mandkhai Molomjamts ◽  
Lawrence P. Wackett
Keyword(s):  
Metabolic Pathways ◽  
Xanthomonas Campestris ◽  
Membrane Lipids ◽  
Membrane Lipid ◽  
Limited Information ◽  
Content Type ◽  
Bacterial Phyla ◽  
Long Chain

ABSTRACT OleA, a member of the thiolase superfamily, is known to catalyze the Claisen condensation of long-chain acyl coenzyme A (acyl-CoA) substrates, initiating metabolic pathways in bacteria for the production of membrane lipids and β-lactone natural products. OleA homologs are found in diverse bacterial phyla, but to date, only one homodimeric OleA has been successfully purified to homogeneity and characterized in vitro. A major impediment for the identification of new OleA enzymes has been protein instability and time-consuming in vitro assays. Here, we developed a bioinformatic pipeline to identify OleA homologs and a new rapid assay to screen OleA enzyme activity in vivo and map their taxonomic diversity. The screen is based on the discovery that OleA displayed surprisingly high rates of p-nitrophenyl ester hydrolysis, an activity not shared by other thiolases, including FabH. The high rates allowed activity to be determined in vitro and with heterologously expressed OleA in vivo via the release of the yellow p-nitrophenol product. Seventy-four putative oleA genes identified in the genomes of diverse bacteria were heterologously expressed in Escherichia coli, and 25 showed activity with p-nitrophenyl esters. The OleA proteins tested were encoded in variable genomic contexts from seven different phyla and are predicted to function in distinct membrane lipid and β-lactone natural product metabolic pathways. This study highlights the diversity of unstudied OleA proteins and presents a rapid method for their identification and characterization. IMPORTANCE Microbially produced β-lactones are found in antibiotic, antitumor, and antiobesity drugs. Long-chain olefinic membrane hydrocarbons have potential utility as fuels and specialty chemicals. The metabolic pathway to both end products share bacterial enzymes denoted as OleA, OleC, and OleD that transform acyl-CoA cellular intermediates into β-lactones. Bacteria producing membrane hydrocarbons via the Ole pathway additionally express a β-lactone decarboxylase, OleB. Both β-lactone and olefin biosynthesis pathways are initiated by OleA enzymes that define the overall structure of the final product. There is currently very limited information on OleA enzymes apart from the single representative from Xanthomonas campestris. In this study, bioinformatic analysis identified hundreds of new, putative OleA proteins, 74 proteins were screened via a rapid whole-cell method, leading to the identification of 25 stably expressed OleA proteins representing seven bacteria phyla.

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