scholarly journals Apoptotic endonuclease EndoG regulates alternative splicing of human telomerase catalytic subunit hTERT

2016 ◽  
Vol 62 (5) ◽  
pp. 544-554 ◽  
Author(s):  
D.D. Zhdanov ◽  
D.A. Vasina ◽  
E.V. Orlova ◽  
V.S. Orlova ◽  
M.V. Pokrovskaya ◽  
...  
Keyword(s):  
Alternative Splicing ◽  
Telomerase Activity ◽  
Catalytic Subunit ◽  
Over Expression ◽  
Htert Gene ◽  
Template Strand ◽  
Non Coding Rna ◽  
Dna Strand ◽  
Human Telomerase ◽  
Long Non Coding Rna

Human telomerase catalytic subunit hTERT is subjected to alternative splicing results in loss of its function and leads to decrease of telomerase activity. However, very little is known about the mechanism of hTERT pre-mRNA alternative splicing. Apoptotic endonuclease EndoG is known to participate this process. The aim of this study was to determine the role of EndoG in regulation of hTERT alternative splicing. Increased expression of b-deletion splice variant was determined during EndoG over-expression in CaCo-2 cell line, after EndoG treatment of cell cytoplasm and nuclei and after nuclei incubation with EndoG digested cell RNA. hTERT alternative splicing was induced by 47-mer RNA oligonucleotide in naked nuclei and in cells after transfection. Identified long non-coding RNA, that is the precursor of 47-mer RNA oligonucleotide. Its size is 1754 nucleotides. Based on the results the following mechanism was proposed. hTERT pre-mRNA is transcribed from coding DNA strand while long non-coding RNA is transcribed from template strand of hTERT gene. EndoG digests long non-coding RNA and produces 47-mer RNA oligonucleotide complementary to hTERT pre-mRNA exon 8 and intron 8 junction place. Interaction of 47-mer RNA oligonucleotide and hTERT pre-mRNA causes alternative splicing.

scholarly journals Induction of apoptotic endonuclease EndoG with DNA-damaging agents initiates alternative splicing of telomerase catalytic subunit hTERT and inhibition of telomerase activity hTERT in human CD4+ and CD8+ T-lymphocytes

2017 ◽  
Vol 63 (4) ◽  
pp. 296-305 ◽  
Author(s):  
D.D. Zhdanov ◽  
D.A. Vasina ◽  
V.S. Orlova ◽  
E.V. Orlova ◽  
D.V. Grishin ◽  
...  
Keyword(s):  
Alternative Splicing ◽  
T Lymphocytes ◽  
Splice Variants ◽  
Telomerase Activity ◽  
Catalytic Subunit ◽  
Human Telomerase Reverse Transcriptase ◽  
Dna Damages ◽  
Dna Damaging Agents ◽  
Cd8 T Lymphocytes ◽  
Human Telomerase

Activity of telomerase catalytic subunit hTERT (human Telomerase Reverse Transcriptase) can be regulated by alternative splicing of its mRNA. At present time exact mechanism of hTERT splicing is not fully understood. Apoptotic endonuclease EndoG is known to participate this process. EndoG expression is induced by DNA damages. The aim of this work was to investigate the ability of DNA-damaging agents with different mechanism of action to induce EndoG expression and inhibit telomerase activity due to the activation of hTERT alternative splicing in normal activated human CD4+ and CD8+ T-lymphocytes. All investigated DNA-damaging agents were able to induce EndoG expression. Cisplatin, a therapeutic compound, producing DNA cross-links induced the highest level of DNA damages and EndoG expression. Incubation of CD4+ and CD8+ T-cells with cisplatin caused the changes in proportion of hTERT splice variants and inhibition of telomerase activity.

scholarly journals Events in the immortalizing process of primary human mammary epithelial cells by the catalytic subunit of human telomerase

2002 ◽  
Vol 365 (3) ◽  
pp. 765-772 ◽  
Author(s):  
Hyunggee KIM ◽  
James FARRIS ◽  
Shelly A. CHRISTMAN ◽  
Byung-Whi KONG ◽  
Linda K. FOSTER ◽  
...  
Keyword(s):  
Single Copy ◽  
Telomerase Activity ◽  
Genetic Alterations ◽  
Mammary Epithelial ◽  
Catalytic Subunit ◽  
Early Passage ◽  
Htert Gene ◽  
Human Mammary Epithelial ◽  
Human Telomerase

The in vitro immortalization of primary human mammary epithelial (HME) cells solely by the exogenous introduction of the catalytic subunit of human telomerase (hTERT) has been achieved. Early passage hTERT-transfected HME (T-HME) cells continuously decreased the length and density of telomeres even in the presence of telomerase activity, with a significant number of cells staining positive for senescence-associated β-galactosidase (SA-β-gal). Subsequently, with the increase in cell passages, the copy number of the exogenously transfected hTERT gene and the percentage of SA-β-gal positive cells were found to decrease. Eventually, a single copy of the exogenous hTERT gene was observed in the relatively later passage T-HME cells in which telomere length was elongated and stabilized without obvious activation of endogenous hTERT and c-Myc expression. In T-HME cells, the expression of two p53 regulated genes p21WAF and HDM2 increased (as in primary senescent HME cells), and was found to be further elevated as the function of p53 was activated by treatment with DNA-damaging agents. p16INK4a was shown to be significantly higher in the primary senescent HME and the early passage T-HME cells when compared with the primary presenescent HME cells, with a dramatic repression of p16INK4a observed in the later passage T-HME cells. In addition, the expression of E2F1 and its transcription factor activity were found to be significantly higher in the later passage T-HME cells when compared with the earlier passage T-HME cells. Together, our results indicate that in vitro immortalization in HME cells may require the activation of the function of telomerase and other genetic alterations such as the spontaneous loss of p16INK4a expression.

scholarly journals Hypermethylation of the human telomerase catalytic subunit (hTERT) gene correlates with telomerase activity

2002 ◽  
Vol 101 (4) ◽  
pp. 335-341 ◽  
Author(s):  
Isabelle Guilleret ◽  
Pu Yan ◽  
Fabienne Grange ◽  
Richard Braunschweig ◽  
Fred T. Bosman ◽  
...  
Keyword(s):  
Telomerase Activity ◽  
Catalytic Subunit ◽  
Htert Gene ◽  
Human Telomerase

scholarly journals Apoptotic endonuclease EndoG induces alternative splicing of telomerase catalytic subunit hTERT and death of tumor cells

2016 ◽  
Vol 62 (3) ◽  
pp. 239-250 ◽  
Author(s):  
D.D. Zhdanov ◽  
D.A. Vasina ◽  
V.S. Orlova ◽  
V.Y. Gotovtseva ◽  
M.V. Bibikova ◽  
...  
Keyword(s):  
Alternative Splicing ◽  
Cell Fate ◽  
Replicative Senescence ◽  
Splice Variants ◽  
Telomerase Activity ◽  
Catalytic Subunit ◽  
Full Length ◽  
Human Telomerase Reverse Transcriptase ◽  
Mrna Induction ◽  
Human Telomerase

Telomerase activity is known to be regulated by alternative splicing of its catalytic subunit hTERT (human Telomerase Reverse Transcriptase) mRNA. Induction of non-active spliced hTERT leads to inhibition of telomerase activity. However, very little is known about the mechanism of hTERT mRNA alternative splicing. The aim of this study was to determine the role of apoptotic endonuclease EndoG in alternative splicing of hTERT and telomerase activity. Strong correlation was found between expression of EndoG and hTERT splice-variants in 12 colon cancer cell lines. Overexpression of EndoG in СаСо-2 cells downregulated the expression of active full-length hTERT variant and upregulated non-active spliced variant. Reduction of full-length hTERT caused downregulation of telomerase activity, dramatically shortening of telomeres length during cell divisions, converting cells to the replicative senescence state, activation of apoptosis and finally cell death. These data indicated the participation of EndoG in alternative splicing of mRNA of telomerase catalytic subunit, regulation of telomerase activity and cell fate.

scholarly journals PD-L1 lncRNA splice isoform promotes lung adenocarcinoma progression via enhancing c-Myc activity

2021 ◽  
Vol 22 (1) ◽  
Author(s):  
Shuang Qu ◽  
Zichen Jiao ◽  
Geng Lu ◽  
Bing Yao ◽  
Ting Wang ◽  
...  
Keyword(s):  
Alternative Splicing ◽  
Lung Adenocarcinoma ◽  
Solid Tumors ◽  
Immune Checkpoint ◽  
Checkpoint Inhibition ◽  
Immune Checkpoint Inhibition ◽  
Non Coding Rna ◽  
Adenocarcinoma Cells ◽  
Lung Adenocarcinoma Cells ◽  
Long Non Coding Rna

Abstract Background Although using a blockade of programmed death-ligand 1 (PD-L1) to enhance T cell immune responses shows great promise in tumor immunotherapy, the immune-checkpoint inhibition strategy is limited for patients with solid tumors. The mechanism and efficacy of such immune-checkpoint inhibition strategies in solid tumors remains unclear. Results Employing qRT-PCR, Sanger sequencing, and RNA BaseScope analysis, we show that human lung adenocarcinoma (LUAD) all produce a long non-coding RNA isoform of PD-L1 (PD-L1-lnc) by alternative splicing, regardless if the tumor is positive or negative for the protein PD-L1. Similar to PD-L1 mRNA, PD-L1-lnc in various lung adenocarcinoma cells is significantly upregulated by IFNγ. Both in vitro and in vivo studies demonstrate that PD-L1-lnc increases proliferation and invasion but decreases apoptosis of lung adenocarcinoma cells. Mechanistically, PD-L1-lnc promotes lung adenocarcinoma progression through directly binding to c-Myc and enhancing c-Myc transcriptional activity. Conclusions In summary, the PD-L1 gene can generate a long non-coding RNA through alternative splicing to promote lung adenocarcinoma progression by enhancing c-Myc activity. Our results argue in favor of investigating PD-L1-lnc depletion in combination with PD-L1 blockade in lung cancer therapy.

scholarly journals Silencing long non-coding RNA ROR improves sensitivity of non-small-cell lung cancer to cisplatin resistance by inhibiting PI3K/Akt/mTOR signaling pathway

Tumor Biology ◽  
2017 ◽  
Vol 39 (5) ◽  
pp. 101042831769756 ◽  
Author(s):  
Hui Shi ◽  
Jin Pu ◽  
Xiao-Li Zhou ◽  
Yun-Ye Ning ◽  
Chong Bai
Keyword(s):  
Cell Proliferation ◽  
Signaling Pathway ◽  
Negative Control ◽  
Mtor Signaling ◽  
Mtor Signaling Pathway ◽  
Control Groups ◽  
Over Expression ◽  
Non Coding Rna ◽  
Protein Expressions ◽  
Long Non Coding Rna

This study aimed to investigate the effects of long non-coding RNA ROR (regulator of reprogramming) on cisplatin (DDP) resistance in patients with non-small-cell lung cancer by regulating PI3K/Akt/mTOR signaling pathway. Human cisplatin-resistant A549/DDP cell lines were selected and divided into control group, negative control group, si-ROR group, ROR over-expression group, Wortmannin group, and ROR over-expression + Wortmannin group. MTT assay was used to determine the optimum inhibitory concentration of DDP. Quantitative real-time polymerase chain reaction and western blotting were applied to detect expressions of long non-coding RNA ROR, PI3K, Akt, and mTOR. Colony-forming assay, scratch test, Transwell assay, and flow cytometry were conducted to detect cell proliferation, migration, invasion, and apoptosis, respectively. Tumor-formation assay was performed to detect the growth of transplanted tumors. Long non-coding RNA ROR expression was high in human A549/DDP cell lines. Compared with the control and negative control groups, the mRNA and protein expressions of PI3K, Akt, mTOR, and bcl-2 decreased, whereas the mRNA and protein expression of bax and the sensitivity of cells to DDP significantly increased. Cell proliferation, migration, and invasion abilities decreased in the si-ROR and Wortmannin groups. In comparison with control and negative control groups, the mRNA and protein expressions of PI3K, Akt, mTOR, and bcl-2 increased, whereas the mRNA and protein expressions of bax decreased, the sensitivity of cells to DDP significantly increased, and cell proliferation, migration, and invasion abilities decreased in the ROR over-expression group. For nude mice in tumor-formation assay, compared with control and negative control groups, the tumor weight was found to be lighter (1.03 ± 0.15) g, the protein expressions of PI3K, Akt, mTOR, and bcl-2 decreased, and the protein expression of bax increased in the si-ROR group. Long non-coding RNA ROR may affect the sensitivity of lung adenocarcinoma cells to DDP by targeting PI3K/Akt/mTOR signaling pathway.

scholarly journals Overexpression of long non coding RNA OR3A4 is associated with altered p38 signalling, morphological and phenotypic changes, and reduced immunogenicity in Barretts Oesophagus

2021 ◽  
Author(s):  
Thomas Nieto ◽  
Yash Sinha ◽  
Qin Qin Zhuang ◽  
Mathew Coleman ◽  
Joanne D Stockton ◽  
...  
Keyword(s):  
T Cells ◽  
P38 Mapk ◽  
Significant Risk ◽  
Oesophageal Adenocarcinoma ◽  
Cellular Phenotype ◽  
Over Expression ◽  
Non Coding Rna ◽  
P38 Mapk Inhibitor ◽  
Long Non Coding Rna ◽  
Mapk Inhibitor

Background: Barretts Oesophagus (BO) presents a particular pathological dilemma, in that patients who have no dysplasia within their BO experience a small but significant risk of malignant progression each year. Screening programmes have attempted to reduce the mortality from BO associated oesophageal adenocarcinoma but cannot predict which BO patients will progress to invasive malignancy. We have previously identified the long non coding RNA, OR3A4, is differentially hypomethylated in progressive BO. We aimed to understand its role in BO pathogenicity Methods: The stable BO cell line CP-A, as well as the oesophageal adenocarcinoma cells line OE-33 was transfected with a lentiviral OR3A4 over-expression vector, and underwent high resolution microscopy, immunofluorescence, RT-qPCR, RNA sequencing, and targeted drug screening with the p38-MAPK inhibitor domipramod to understand the effects of OR3A4 expression on progression. We then compared progressive vs. non-progressive BO samples using quantitative multi-fluorophore (Vectra) immunohistochemistry. Results: Over-expression of OR3A4 in CP-A lines resulted in a hyperproliferative, dysplastic cellular phenotype, with strong over-expression of MAPK and anti-apoptotic pathways at the RNA and protein level, which was sensitive to the p38-MAPK inhibitor domipramod. Vectra immunohistochemistry demonstrated that progressive BO had reduced visibility associated with a reduction in CD8+ T-cells and CD68+ macrophages and reduced CD4+ T-cells in the stomal compartment. Conclusion: The overexpression of OR3A4, which we have previously shown is associated with progressive BO leads to a proliferative dysplastic cellular phenotype associated with increased, reversible MAPK signalling and loss of immune visibility.

scholarly journals UP-REGULATION OF LONG NON-CODING RNA MORBID ATTENUATES SENESCENCE

2019 ◽  
Vol 3 (Supplement_1) ◽  
pp. S90-S90
Author(s):  
Zheng Kuai ◽  
Meiting Chen ◽  
yang yu ◽  
Fan Yang ◽  
chunxiang Zhang
Keyword(s):  
Cardiovascular Disease ◽  
Noncoding Rna ◽  
Molecular Mechanisms ◽  
Cardiac Myocyte ◽  
Diagnostic Biomarker ◽  
Over Expression ◽  
Non Coding Rna ◽  
Organ Level ◽  
Long Non Coding Rna ◽  
Over Time

Abstract Aging is the inevitable, irreversible decline in function on the cellular and organ level leading to increased incidence of the most frequent diseases such as cancer and cardiovascular disease, that occurs over time. whereas the molecular mechanisms of senescence remain largely unknown. Here we identified that a novel long noncoding RNA, Morrbid was significantly decreased in different organs of aged mice, such as heart, liver, spleen, lung, kidney and brain. Interestingly, the telomeres length of Morrbid KO mice were significantly shorted than the WT mice at the same age. We also found that Morrbid was steeply decreased in a natural mouse cardiac myocyte senescence model. The senescence of mouse cardiac myocytes was effectively attenuated by Morrbid over-expression shown by the decreased β-galactosidase staining, increased telomere activity, decreased production of ROS and decreased cell apoptosis, but was enhanced by Morrbid knockdown. The results suggest that Morrbid is a critical regulator in senescence and could be used as a novel diagnostic biomarker for it, and a new therapeutic target for diverse diseases.

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