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The selective activation of p53 target genes regulated by SMYD2 in BIX-01294 induced autophagy-related cell death.

Fan JD, Lei PJ, Zheng JY, Wang X, Li S, Liu H, He YL, Wang ZN, Wei G, Zhang X, Li LY, Wu M - PLoS ONE (2015)

Bottom Line: SMYD2 is a methyltransferase for p53 and regulates its transcription activity.Its deficiency enhances the BIX-01294-induced autophagy-related cell death through transcriptionally promoting the expression of p53 target genes.Taken together, our data suggest BIX-01294 induces autophagy-related cell death and selectively activates p53 target genes, which is repressed by SMYD2 methyltransferase.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry and Molecular Biology, College of Life Sciences, Wuhan University, Wuhan, Hubei 430072, China.

ABSTRACT
Transcription regulation emerged to be one of the key mechanisms in regulating autophagy. Inhibitors of H3K9 methylation activates the expression of LC3B, as well as other autophagy-related genes, and promotes autophagy process. However, the detailed mechanisms of autophagy regulated by nuclear factors remain elusive. In this study, we performed a drug screen of SMYD2-/- cells and discovered that SMYD2 deficiency enhanced the cell death induced by BIX01294, an inhibitor of histone H3K9 methylation. BIX-01294 induces accumulation of LC3 II and autophagy-related cell death, but not caspase-dependent apoptosis. We profiled the global gene expression pattern after treatment with BIX-01294, in comparison with rapamycin. BIX-01294 selectively activates the downstream genes of p53 signaling, such as p21 and DOR, but not PUMA, a typical p53 target gene inducing apoptosis. BIX-01294 also induces other autophagy-related genes, such as ATG4A and ATG9A. SMYD2 is a methyltransferase for p53 and regulates its transcription activity. Its deficiency enhances the BIX-01294-induced autophagy-related cell death through transcriptionally promoting the expression of p53 target genes. Taken together, our data suggest BIX-01294 induces autophagy-related cell death and selectively activates p53 target genes, which is repressed by SMYD2 methyltransferase.

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Genome wide gene expression profiles of the cells treated with BIX-01294 and rapamycin.HCT116 cells were transfected with control or SMYD2 siRNA, and treated with H2O, BIX-01294 and rapamycin respectively. (A) Clustering of the different expressed genes induced by rapamycin among the three samples showed that they can be divided into four groups based on their expression patterns. Each group was analyzed by GO analysis except group 1, which no significant biological process was found. (B) Venn diagram shows the numbers of gens regulated by BIX-01294, rapamycin alone or both. (C) GO analysis of signaling pathways enriched by different expressed genes uniquely activated by BIX-01294. (D) SMYD2+/+ and SMYD2-/- cells were treated with H2O, BIX-01294 and rapamycin respectively. Different expressed p53 target genes were shown in the heat map. (E) The mRNA level of the typical p53 target genes was examined by quantitative PCR in the indicated samples.
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pone.0116782.g005: Genome wide gene expression profiles of the cells treated with BIX-01294 and rapamycin.HCT116 cells were transfected with control or SMYD2 siRNA, and treated with H2O, BIX-01294 and rapamycin respectively. (A) Clustering of the different expressed genes induced by rapamycin among the three samples showed that they can be divided into four groups based on their expression patterns. Each group was analyzed by GO analysis except group 1, which no significant biological process was found. (B) Venn diagram shows the numbers of gens regulated by BIX-01294, rapamycin alone or both. (C) GO analysis of signaling pathways enriched by different expressed genes uniquely activated by BIX-01294. (D) SMYD2+/+ and SMYD2-/- cells were treated with H2O, BIX-01294 and rapamycin respectively. Different expressed p53 target genes were shown in the heat map. (E) The mRNA level of the typical p53 target genes was examined by quantitative PCR in the indicated samples.

Mentions: Our data suggested that BIX-01294 induce autophagy through transcription regulation (Fig. 3C). To further investigate the mechanisms of BIX-01294-induced autophagy, we compared the global gene expression profiles of BIX-01294 and rapamycin treatment, using high throughput mRNA sequencing. Before sample submission, we analyzed the cells by western blot to ensure the similar amount of LC3 II was induced by the above two drugs. BIX-01294 treatment changed the expression of 1950 genes (more than 1.5 folds), and 1494 genes for rapamycin (Fig. 5A and Tables B-C in S1 File). Among them, 535 different expressed genes (DEG) were overlapped (Fig. 5B). We took the DEGs between rapamycin and control, cluster them among all three samples and grouped into 4 groups (Fig. 5A). The genes in group 1 are activated by rapamycin but not BIX-01294 (Fig. 5A). We failed to link them with any biological process with GO analysis. The genes in group 2 are both activated by the two drugs and are mainly related with transcriptional regulation and RNA metabolism (Fig. 5A). The genes in group 3 are down regulated by both drugs and they are mostly related with processes involving mitochondria (Fig. 5A). Those in group 4 are down regulated by rapamycin, but did not change too much with BIX-01294. They are mostly related with cell cycle, metabolism and cell death (Fig. 5A).


The selective activation of p53 target genes regulated by SMYD2 in BIX-01294 induced autophagy-related cell death.

Fan JD, Lei PJ, Zheng JY, Wang X, Li S, Liu H, He YL, Wang ZN, Wei G, Zhang X, Li LY, Wu M - PLoS ONE (2015)

Genome wide gene expression profiles of the cells treated with BIX-01294 and rapamycin.HCT116 cells were transfected with control or SMYD2 siRNA, and treated with H2O, BIX-01294 and rapamycin respectively. (A) Clustering of the different expressed genes induced by rapamycin among the three samples showed that they can be divided into four groups based on their expression patterns. Each group was analyzed by GO analysis except group 1, which no significant biological process was found. (B) Venn diagram shows the numbers of gens regulated by BIX-01294, rapamycin alone or both. (C) GO analysis of signaling pathways enriched by different expressed genes uniquely activated by BIX-01294. (D) SMYD2+/+ and SMYD2-/- cells were treated with H2O, BIX-01294 and rapamycin respectively. Different expressed p53 target genes were shown in the heat map. (E) The mRNA level of the typical p53 target genes was examined by quantitative PCR in the indicated samples.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4285553&req=5

pone.0116782.g005: Genome wide gene expression profiles of the cells treated with BIX-01294 and rapamycin.HCT116 cells were transfected with control or SMYD2 siRNA, and treated with H2O, BIX-01294 and rapamycin respectively. (A) Clustering of the different expressed genes induced by rapamycin among the three samples showed that they can be divided into four groups based on their expression patterns. Each group was analyzed by GO analysis except group 1, which no significant biological process was found. (B) Venn diagram shows the numbers of gens regulated by BIX-01294, rapamycin alone or both. (C) GO analysis of signaling pathways enriched by different expressed genes uniquely activated by BIX-01294. (D) SMYD2+/+ and SMYD2-/- cells were treated with H2O, BIX-01294 and rapamycin respectively. Different expressed p53 target genes were shown in the heat map. (E) The mRNA level of the typical p53 target genes was examined by quantitative PCR in the indicated samples.
Mentions: Our data suggested that BIX-01294 induce autophagy through transcription regulation (Fig. 3C). To further investigate the mechanisms of BIX-01294-induced autophagy, we compared the global gene expression profiles of BIX-01294 and rapamycin treatment, using high throughput mRNA sequencing. Before sample submission, we analyzed the cells by western blot to ensure the similar amount of LC3 II was induced by the above two drugs. BIX-01294 treatment changed the expression of 1950 genes (more than 1.5 folds), and 1494 genes for rapamycin (Fig. 5A and Tables B-C in S1 File). Among them, 535 different expressed genes (DEG) were overlapped (Fig. 5B). We took the DEGs between rapamycin and control, cluster them among all three samples and grouped into 4 groups (Fig. 5A). The genes in group 1 are activated by rapamycin but not BIX-01294 (Fig. 5A). We failed to link them with any biological process with GO analysis. The genes in group 2 are both activated by the two drugs and are mainly related with transcriptional regulation and RNA metabolism (Fig. 5A). The genes in group 3 are down regulated by both drugs and they are mostly related with processes involving mitochondria (Fig. 5A). Those in group 4 are down regulated by rapamycin, but did not change too much with BIX-01294. They are mostly related with cell cycle, metabolism and cell death (Fig. 5A).

Bottom Line: SMYD2 is a methyltransferase for p53 and regulates its transcription activity.Its deficiency enhances the BIX-01294-induced autophagy-related cell death through transcriptionally promoting the expression of p53 target genes.Taken together, our data suggest BIX-01294 induces autophagy-related cell death and selectively activates p53 target genes, which is repressed by SMYD2 methyltransferase.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry and Molecular Biology, College of Life Sciences, Wuhan University, Wuhan, Hubei 430072, China.

ABSTRACT
Transcription regulation emerged to be one of the key mechanisms in regulating autophagy. Inhibitors of H3K9 methylation activates the expression of LC3B, as well as other autophagy-related genes, and promotes autophagy process. However, the detailed mechanisms of autophagy regulated by nuclear factors remain elusive. In this study, we performed a drug screen of SMYD2-/- cells and discovered that SMYD2 deficiency enhanced the cell death induced by BIX01294, an inhibitor of histone H3K9 methylation. BIX-01294 induces accumulation of LC3 II and autophagy-related cell death, but not caspase-dependent apoptosis. We profiled the global gene expression pattern after treatment with BIX-01294, in comparison with rapamycin. BIX-01294 selectively activates the downstream genes of p53 signaling, such as p21 and DOR, but not PUMA, a typical p53 target gene inducing apoptosis. BIX-01294 also induces other autophagy-related genes, such as ATG4A and ATG9A. SMYD2 is a methyltransferase for p53 and regulates its transcription activity. Its deficiency enhances the BIX-01294-induced autophagy-related cell death through transcriptionally promoting the expression of p53 target genes. Taken together, our data suggest BIX-01294 induces autophagy-related cell death and selectively activates p53 target genes, which is repressed by SMYD2 methyltransferase.

Show MeSH
Related in: MedlinePlus