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Long non-coding RNA ROR decoys gene-specific histone methylation to promote tumorigenesis.

Fan J, Xing Y, Wen X, Jia R, Ni H, He J, Ding X, Pan H, Qian G, Ge S, Hoffman AR, Zhang H, Fan X - Genome Biol. (2015)

Bottom Line: Suppression of ROR in tumors results in silencing of TESC expression, and G9A-mediated histone H3K9 methylation in the TESC promoter is restored, which significantly reduces tumor growth and metastasis.Without ROR silencing, TESC knockdown presents consistent and significant reductions in tumor progression.Our results reveal a novel mechanism by which ROR may serve as a decoy oncoRNA that blocks binding surfaces, preventing the recruitment of histone modifying enzymes, thereby specifying a new pattern of histone modifications that promote tumorigenesis.

View Article: PubMed Central - PubMed

Affiliation: Department of Ophthalmology, Ninth People's Hospital, Shanghai JiaoTong University School of Medicine, Shanghai, 200025, P. R. China.

ABSTRACT

Background: Long non-coding RNAs (lncRNAs) are not translated into proteins and were initially considered to be part of the 'dark matter' of the genome. Recently, it has been shown that lncRNAs play a role in the recruitment of chromatin modifying complexes and can influence gene expression. However, it is unknown if lncRNAs function in a similar way in cancer.

Results: Here, we show that the lncRNA ROR occupies and activates the TESC promoter by repelling the histone G9A methyltransferase and promoting the release of histone H3K9 methylation. Suppression of ROR in tumors results in silencing of TESC expression, and G9A-mediated histone H3K9 methylation in the TESC promoter is restored, which significantly reduces tumor growth and metastasis. Without ROR silencing, TESC knockdown presents consistent and significant reductions in tumor progression.

Conclusions: Our results reveal a novel mechanism by which ROR may serve as a decoy oncoRNA that blocks binding surfaces, preventing the recruitment of histone modifying enzymes, thereby specifying a new pattern of histone modifications that promote tumorigenesis.

No MeSH data available.


Related in: MedlinePlus

TESC serves as a target of ROR. a Real-time PCR measurement of mRNA expression of TESC. The expression levels of TESC in tumor cells were significantly higher than normal. Once ROR was silenced in tumor cells, there was a remarkable decrease in TESC expression. NCM460: normal colon cell; GES-1: normal gastric cell. b Overlapping altered genes from lncRNA-ROR knockdown cells from AGS and HT29 cells. By using four-fold changes as baseline, there were approximately 58 altered genes after depletion of ROR in AGS and 125 altered genes in HT29. A total of six genes were changed in AGS and HT29 cells after ROR knockdown. c Western blot showing that the expression of TESC in ROR silenced AGS and HT29 tumor cells. The expression of TESC was greatly decreased in ROR-depleted tumor cells as well as in normal cells, including fibroblasts, normal colon cell NCM460, and normal gastric cell GES-1. d, e Silenced expression of TESC using siRNAs in AGS (d) and HT29 cells (e). Real-time PCR demonstrated siTESC provided the optimal deletion of TESC. Experiments were performed 48 h following siTESC (125 pmol) and control siRNA(125 pmol) treatment. *P <0.05: compared with the control and NC. NC: non-silencing control. f Western blot demonstrating that siTESC efficiently silenced TESC at the protein expression level in AGS and HT29 cells. All experiments were performed 48 h following siTESC (125 pmol) and control siRNA (125 pmol) treatment. NC: non-silencing control. g RT-PCR showing that ROR expression was not significantly changed when TESC was silenced in AGS and HT29 cells
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Fig3: TESC serves as a target of ROR. a Real-time PCR measurement of mRNA expression of TESC. The expression levels of TESC in tumor cells were significantly higher than normal. Once ROR was silenced in tumor cells, there was a remarkable decrease in TESC expression. NCM460: normal colon cell; GES-1: normal gastric cell. b Overlapping altered genes from lncRNA-ROR knockdown cells from AGS and HT29 cells. By using four-fold changes as baseline, there were approximately 58 altered genes after depletion of ROR in AGS and 125 altered genes in HT29. A total of six genes were changed in AGS and HT29 cells after ROR knockdown. c Western blot showing that the expression of TESC in ROR silenced AGS and HT29 tumor cells. The expression of TESC was greatly decreased in ROR-depleted tumor cells as well as in normal cells, including fibroblasts, normal colon cell NCM460, and normal gastric cell GES-1. d, e Silenced expression of TESC using siRNAs in AGS (d) and HT29 cells (e). Real-time PCR demonstrated siTESC provided the optimal deletion of TESC. Experiments were performed 48 h following siTESC (125 pmol) and control siRNA(125 pmol) treatment. *P <0.05: compared with the control and NC. NC: non-silencing control. f Western blot demonstrating that siTESC efficiently silenced TESC at the protein expression level in AGS and HT29 cells. All experiments were performed 48 h following siTESC (125 pmol) and control siRNA (125 pmol) treatment. NC: non-silencing control. g RT-PCR showing that ROR expression was not significantly changed when TESC was silenced in AGS and HT29 cells

Mentions: To determine the factors that coordinated these tumor variations after ROR knockdown, we examined gene expression by a genome-wide cDNA array. Compared with untreated tumor cells, the expression of at least 58 genes was significantly changed by more than four-fold (Additional file 2: Table S1, GEO accession number: GSE67416, fold >4), including genes that were both up- and downregulated in AGS cells. We then selected seven notable altered genes: AKR1C1, AKR1C3, LMO4, MGST1, LXN, TIMP3, and TESC. The results indicate that TESC was significantly decreased in ROR-depleted cells (Fig. 3a). Similarly, we also further confirmed the reduction of the other six candidate genes through real-time PCR (Additional file 3: Figure S2 a-f). Unfortunately, of these potential targets, altered tumor activity was not detected after silencing their expression by siRNA except for in TESC (Additional file 3: Figure S2 g, h). We also detected gene expression after ROR depletion in HT29 cells by genome-wide cDNA array. Intriguingly, in HT29 cells, TESC was also one of the most significantly altered genes (Additional file 4: Table S2, GEO accession number: GSE67416, fold >4), and its expression overlapped with that of AGS cells (Fig. 3b). Moreover, because the regulatory role for TESC in tumorigenesis was not indicated, we then explored whether TESC serves as a possible downstream ROR-targeting gene to modulate tumor activity.Fig. 3


Long non-coding RNA ROR decoys gene-specific histone methylation to promote tumorigenesis.

Fan J, Xing Y, Wen X, Jia R, Ni H, He J, Ding X, Pan H, Qian G, Ge S, Hoffman AR, Zhang H, Fan X - Genome Biol. (2015)

TESC serves as a target of ROR. a Real-time PCR measurement of mRNA expression of TESC. The expression levels of TESC in tumor cells were significantly higher than normal. Once ROR was silenced in tumor cells, there was a remarkable decrease in TESC expression. NCM460: normal colon cell; GES-1: normal gastric cell. b Overlapping altered genes from lncRNA-ROR knockdown cells from AGS and HT29 cells. By using four-fold changes as baseline, there were approximately 58 altered genes after depletion of ROR in AGS and 125 altered genes in HT29. A total of six genes were changed in AGS and HT29 cells after ROR knockdown. c Western blot showing that the expression of TESC in ROR silenced AGS and HT29 tumor cells. The expression of TESC was greatly decreased in ROR-depleted tumor cells as well as in normal cells, including fibroblasts, normal colon cell NCM460, and normal gastric cell GES-1. d, e Silenced expression of TESC using siRNAs in AGS (d) and HT29 cells (e). Real-time PCR demonstrated siTESC provided the optimal deletion of TESC. Experiments were performed 48 h following siTESC (125 pmol) and control siRNA(125 pmol) treatment. *P <0.05: compared with the control and NC. NC: non-silencing control. f Western blot demonstrating that siTESC efficiently silenced TESC at the protein expression level in AGS and HT29 cells. All experiments were performed 48 h following siTESC (125 pmol) and control siRNA (125 pmol) treatment. NC: non-silencing control. g RT-PCR showing that ROR expression was not significantly changed when TESC was silenced in AGS and HT29 cells
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Fig3: TESC serves as a target of ROR. a Real-time PCR measurement of mRNA expression of TESC. The expression levels of TESC in tumor cells were significantly higher than normal. Once ROR was silenced in tumor cells, there was a remarkable decrease in TESC expression. NCM460: normal colon cell; GES-1: normal gastric cell. b Overlapping altered genes from lncRNA-ROR knockdown cells from AGS and HT29 cells. By using four-fold changes as baseline, there were approximately 58 altered genes after depletion of ROR in AGS and 125 altered genes in HT29. A total of six genes were changed in AGS and HT29 cells after ROR knockdown. c Western blot showing that the expression of TESC in ROR silenced AGS and HT29 tumor cells. The expression of TESC was greatly decreased in ROR-depleted tumor cells as well as in normal cells, including fibroblasts, normal colon cell NCM460, and normal gastric cell GES-1. d, e Silenced expression of TESC using siRNAs in AGS (d) and HT29 cells (e). Real-time PCR demonstrated siTESC provided the optimal deletion of TESC. Experiments were performed 48 h following siTESC (125 pmol) and control siRNA(125 pmol) treatment. *P <0.05: compared with the control and NC. NC: non-silencing control. f Western blot demonstrating that siTESC efficiently silenced TESC at the protein expression level in AGS and HT29 cells. All experiments were performed 48 h following siTESC (125 pmol) and control siRNA (125 pmol) treatment. NC: non-silencing control. g RT-PCR showing that ROR expression was not significantly changed when TESC was silenced in AGS and HT29 cells
Mentions: To determine the factors that coordinated these tumor variations after ROR knockdown, we examined gene expression by a genome-wide cDNA array. Compared with untreated tumor cells, the expression of at least 58 genes was significantly changed by more than four-fold (Additional file 2: Table S1, GEO accession number: GSE67416, fold >4), including genes that were both up- and downregulated in AGS cells. We then selected seven notable altered genes: AKR1C1, AKR1C3, LMO4, MGST1, LXN, TIMP3, and TESC. The results indicate that TESC was significantly decreased in ROR-depleted cells (Fig. 3a). Similarly, we also further confirmed the reduction of the other six candidate genes through real-time PCR (Additional file 3: Figure S2 a-f). Unfortunately, of these potential targets, altered tumor activity was not detected after silencing their expression by siRNA except for in TESC (Additional file 3: Figure S2 g, h). We also detected gene expression after ROR depletion in HT29 cells by genome-wide cDNA array. Intriguingly, in HT29 cells, TESC was also one of the most significantly altered genes (Additional file 4: Table S2, GEO accession number: GSE67416, fold >4), and its expression overlapped with that of AGS cells (Fig. 3b). Moreover, because the regulatory role for TESC in tumorigenesis was not indicated, we then explored whether TESC serves as a possible downstream ROR-targeting gene to modulate tumor activity.Fig. 3

Bottom Line: Suppression of ROR in tumors results in silencing of TESC expression, and G9A-mediated histone H3K9 methylation in the TESC promoter is restored, which significantly reduces tumor growth and metastasis.Without ROR silencing, TESC knockdown presents consistent and significant reductions in tumor progression.Our results reveal a novel mechanism by which ROR may serve as a decoy oncoRNA that blocks binding surfaces, preventing the recruitment of histone modifying enzymes, thereby specifying a new pattern of histone modifications that promote tumorigenesis.

View Article: PubMed Central - PubMed

Affiliation: Department of Ophthalmology, Ninth People's Hospital, Shanghai JiaoTong University School of Medicine, Shanghai, 200025, P. R. China.

ABSTRACT

Background: Long non-coding RNAs (lncRNAs) are not translated into proteins and were initially considered to be part of the 'dark matter' of the genome. Recently, it has been shown that lncRNAs play a role in the recruitment of chromatin modifying complexes and can influence gene expression. However, it is unknown if lncRNAs function in a similar way in cancer.

Results: Here, we show that the lncRNA ROR occupies and activates the TESC promoter by repelling the histone G9A methyltransferase and promoting the release of histone H3K9 methylation. Suppression of ROR in tumors results in silencing of TESC expression, and G9A-mediated histone H3K9 methylation in the TESC promoter is restored, which significantly reduces tumor growth and metastasis. Without ROR silencing, TESC knockdown presents consistent and significant reductions in tumor progression.

Conclusions: Our results reveal a novel mechanism by which ROR may serve as a decoy oncoRNA that blocks binding surfaces, preventing the recruitment of histone modifying enzymes, thereby specifying a new pattern of histone modifications that promote tumorigenesis.

No MeSH data available.


Related in: MedlinePlus