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Transcriptome analysis of human OXR1 depleted cells reveals its role in regulating the p53 signaling pathway.

Yang M, Lin X, Rowe A, Rognes T, Eide L, Bjørås M - Sci Rep (2015)

Bottom Line: The oxidation resistance gene 1 (OXR1) is crucial for protecting against oxidative stress; however, its molecular function is unknown.In total, in non-treated and hydrogen peroxide exposed HeLa cells, OXR1 depletion resulted in down-regulation of 554 genes and up-regulation of 253 genes.In summary, OXR1 may act as a sensor of cellular oxidative stress to regulate the transcriptional networks required to detoxify reactive oxygen species and modulate cell cycle and apoptosis.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology, Oslo University Hospital and University of Oslo, Norway.

ABSTRACT
The oxidation resistance gene 1 (OXR1) is crucial for protecting against oxidative stress; however, its molecular function is unknown. We employed RNA sequencing to examine the role of human OXR1 for genome wide transcription regulation. In total, in non-treated and hydrogen peroxide exposed HeLa cells, OXR1 depletion resulted in down-regulation of 554 genes and up-regulation of 253 genes. These differentially expressed genes include transcription factors (i.e. HIF1A, SP6, E2F8 and TCF3), antioxidant genes (PRDX4, PTGS1 and CYGB) and numerous genes of the p53 signaling pathway involved in cell-cycle arrest (i.e. cyclin D, CDK6 and RPRM) and apoptosis (i.e. CytC and CASP9). We demonstrated that OXR1 depleted cells undergo cell cycle arrest in G2/M phase during oxidative stress and increase protein expression of the apoptosis initiator protease CASP9. In summary, OXR1 may act as a sensor of cellular oxidative stress to regulate the transcriptional networks required to detoxify reactive oxygen species and modulate cell cycle and apoptosis.

No MeSH data available.


Related in: MedlinePlus

HOXR1 depletion caused cell cycle arrest in G2/M phase.The proportion of cells decreased in G1, but increased in G2/M phase in hOXR1-depleted cells (siOXR1) as compared to control (siCon). Upper panel: one representative set of flow cytometry data. The HeLa cells were exposed to 0.25 mM H2O2 for 1 h or non-treated (NT), recovered in normal medium for 24 h, fixed, stained by propidium iodide (PI) and subjected to flow cytometry. Bottom panel: Data quantification, n = 4. *p < 0.01.
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f6: HOXR1 depletion caused cell cycle arrest in G2/M phase.The proportion of cells decreased in G1, but increased in G2/M phase in hOXR1-depleted cells (siOXR1) as compared to control (siCon). Upper panel: one representative set of flow cytometry data. The HeLa cells were exposed to 0.25 mM H2O2 for 1 h or non-treated (NT), recovered in normal medium for 24 h, fixed, stained by propidium iodide (PI) and subjected to flow cytometry. Bottom panel: Data quantification, n = 4. *p < 0.01.

Mentions: In response to DNA damage the p53 signaling pathway arrests the cell cycle. In our transcriptome analysis, we find four differentially expressed genes involved in regulating cell cycle progression via the p53 pathway; CDKN1A (p21), CCND1, CDK6 and reprimo (RPRM). In hOXR1 depleted cells under both normal conditions and oxidative stress, the cell cycle inhibitor p21 was down-regulated, the G2 arrest mediator RPRM was up-regulated and the G1/S transmission stimulators CDK6 and cyclin D were up- and down-regulated, respectively. To examine the role of hOXR1 in cell cycle regulation, we measured the distribution of hOXR1 depleted HeLa and control cells in G1, S and G2/M phase by flow cytometry. First, control cells were tested at 0.25 or 0.5 mM H2O2 exposure (1 h) and 24 h recovery time, showing that 17.2% or 36.5% of the cells were enriched in G2/M, respectively. It thus appears that the cells arrest in G2/M in a dose dependent manner at these concentrations of H2O2. Next, we exposed hOXR1 depleted cells and control to the lowest dose of H2O2 (0.25 mM) to avoid cell death (more than 95% survival). Non-treated hOXR1depleted cells showed a significant reduction in number of cells in G1 phase, but increased number of cells in S and G2/M phases in comparison to control cells (Fig. 6). After exposure to peroxide, the cell numbers in both G1 and S phase decreased (Fig. 6). As expected, the population of cells in G2/M phase increased in both control and hOXR1 silenced cells as compared to non-treated cells, confirming that the cells were mainly arrested in G2/M in response to hydrogen peroxide exposure. Importantly, the cell population in G1 was significantly lower in hOXR1 depleted cells as compared to control cells, while cell numbers in G2/M were significantly higher in hOXR1-depleted cells than control cells after hydrogen peroxide treatment. Thus it appears that hOXR1 plays an important role in cell cycle progression by regulating the p53 pathway via p21, CCND1, CDK6 and RPRM.


Transcriptome analysis of human OXR1 depleted cells reveals its role in regulating the p53 signaling pathway.

Yang M, Lin X, Rowe A, Rognes T, Eide L, Bjørås M - Sci Rep (2015)

HOXR1 depletion caused cell cycle arrest in G2/M phase.The proportion of cells decreased in G1, but increased in G2/M phase in hOXR1-depleted cells (siOXR1) as compared to control (siCon). Upper panel: one representative set of flow cytometry data. The HeLa cells were exposed to 0.25 mM H2O2 for 1 h or non-treated (NT), recovered in normal medium for 24 h, fixed, stained by propidium iodide (PI) and subjected to flow cytometry. Bottom panel: Data quantification, n = 4. *p < 0.01.
© Copyright Policy - open-access
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4663793&req=5

f6: HOXR1 depletion caused cell cycle arrest in G2/M phase.The proportion of cells decreased in G1, but increased in G2/M phase in hOXR1-depleted cells (siOXR1) as compared to control (siCon). Upper panel: one representative set of flow cytometry data. The HeLa cells were exposed to 0.25 mM H2O2 for 1 h or non-treated (NT), recovered in normal medium for 24 h, fixed, stained by propidium iodide (PI) and subjected to flow cytometry. Bottom panel: Data quantification, n = 4. *p < 0.01.
Mentions: In response to DNA damage the p53 signaling pathway arrests the cell cycle. In our transcriptome analysis, we find four differentially expressed genes involved in regulating cell cycle progression via the p53 pathway; CDKN1A (p21), CCND1, CDK6 and reprimo (RPRM). In hOXR1 depleted cells under both normal conditions and oxidative stress, the cell cycle inhibitor p21 was down-regulated, the G2 arrest mediator RPRM was up-regulated and the G1/S transmission stimulators CDK6 and cyclin D were up- and down-regulated, respectively. To examine the role of hOXR1 in cell cycle regulation, we measured the distribution of hOXR1 depleted HeLa and control cells in G1, S and G2/M phase by flow cytometry. First, control cells were tested at 0.25 or 0.5 mM H2O2 exposure (1 h) and 24 h recovery time, showing that 17.2% or 36.5% of the cells were enriched in G2/M, respectively. It thus appears that the cells arrest in G2/M in a dose dependent manner at these concentrations of H2O2. Next, we exposed hOXR1 depleted cells and control to the lowest dose of H2O2 (0.25 mM) to avoid cell death (more than 95% survival). Non-treated hOXR1depleted cells showed a significant reduction in number of cells in G1 phase, but increased number of cells in S and G2/M phases in comparison to control cells (Fig. 6). After exposure to peroxide, the cell numbers in both G1 and S phase decreased (Fig. 6). As expected, the population of cells in G2/M phase increased in both control and hOXR1 silenced cells as compared to non-treated cells, confirming that the cells were mainly arrested in G2/M in response to hydrogen peroxide exposure. Importantly, the cell population in G1 was significantly lower in hOXR1 depleted cells as compared to control cells, while cell numbers in G2/M were significantly higher in hOXR1-depleted cells than control cells after hydrogen peroxide treatment. Thus it appears that hOXR1 plays an important role in cell cycle progression by regulating the p53 pathway via p21, CCND1, CDK6 and RPRM.

Bottom Line: The oxidation resistance gene 1 (OXR1) is crucial for protecting against oxidative stress; however, its molecular function is unknown.In total, in non-treated and hydrogen peroxide exposed HeLa cells, OXR1 depletion resulted in down-regulation of 554 genes and up-regulation of 253 genes.In summary, OXR1 may act as a sensor of cellular oxidative stress to regulate the transcriptional networks required to detoxify reactive oxygen species and modulate cell cycle and apoptosis.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology, Oslo University Hospital and University of Oslo, Norway.

ABSTRACT
The oxidation resistance gene 1 (OXR1) is crucial for protecting against oxidative stress; however, its molecular function is unknown. We employed RNA sequencing to examine the role of human OXR1 for genome wide transcription regulation. In total, in non-treated and hydrogen peroxide exposed HeLa cells, OXR1 depletion resulted in down-regulation of 554 genes and up-regulation of 253 genes. These differentially expressed genes include transcription factors (i.e. HIF1A, SP6, E2F8 and TCF3), antioxidant genes (PRDX4, PTGS1 and CYGB) and numerous genes of the p53 signaling pathway involved in cell-cycle arrest (i.e. cyclin D, CDK6 and RPRM) and apoptosis (i.e. CytC and CASP9). We demonstrated that OXR1 depleted cells undergo cell cycle arrest in G2/M phase during oxidative stress and increase protein expression of the apoptosis initiator protease CASP9. In summary, OXR1 may act as a sensor of cellular oxidative stress to regulate the transcriptional networks required to detoxify reactive oxygen species and modulate cell cycle and apoptosis.

No MeSH data available.


Related in: MedlinePlus