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DNA methylation and histone modifications regulate SOX11 expression in lymphoid and solid cancer cells.

Nordström L, Andersson E, Kuci V, Gustavsson E, Holm K, Ringnér M, Guldberg P, Ek S - BMC Cancer (2015)

Bottom Line: In contrast, SOX11 regulation in neoplastic tissues is more complex involving both DNA methylation and histone modifications.The possibility to re-express SOX11 in non-methylated tissue is of clinical relevance, and was successfully achieved in cell lines with low levels of SOX11 methylation.In breast cancer patients, methylation of the SOX11 promoter was shown to correlate with estrogen receptor status, suggesting that SOX11 may be functionally re-expressed during treatment with HDAC inhibitors in specific patient subgroups.

View Article: PubMed Central - PubMed

Affiliation: Department of Immunotechnology, CREATE Health, Lund University, Lund, Sweden. lena.nordstrom@immun.lth.se.

ABSTRACT

Background: The neural transcription factor SOX11 is present at specific stages during embryo development with a very restricted expression in adult tissue, indicating precise regulation of transcription. SOX11 is strongly up-regulated in some malignancies and have a functional role in tumorgenesis. With the aim to explore differences in epigenetic regulation of SOX11 expression in normal versus neoplastic cells, we investigated methylation and histone modifications related to the SOX11 promoter and the possibility to induce re-expression using histone deacetylase (HDAC) or EZH2 inhibitors.

Methods: The epigenetic regulation of SOX11 was investigated in distinct non-malignant cell populations (n = 7) and neoplastic cell-lines (n = 42) of different cellular origins. DNA methylation was assessed using bisulfite sequencing, methylation-specific melting curve analysis, MethyLight and pyrosequencing. The presence of H3K27me3 was assessed using ChIP-qPCR. The HDAC inhibitors Vorinostat and trichostatin A were used to induce SOX11 in cell lines with no endogenous expression.

Results: The SOX11 promoter shows a low degree of methylation and strong enrichment of H3K27me3 in non-malignant differentiated cells, independent of cellular origin. Cancers of the B-cell lineage are strongly marked by de novo methylation at the SOX11 promoter in SOX11 non-expressing cells, while solid cancer entities display a more varying degree of SOX11 promoter methylation. The silencing mark H3K27me3 was generally present at the SOX11 promoter in non-expressing cells, and an increased enrichment was observed in cancer cells with a low degree of SOX11 methylation compared to cells with dense methylation. Finally, we demonstrate that the HDAC inhibitors (vorinostat and trichostatin A) induce SOX11 expression in cancer cells with low levels of SOX11 methylation.

Conclusions: We show that SOX11 is strongly marked by repressive histone marks in non-malignant cells. In contrast, SOX11 regulation in neoplastic tissues is more complex involving both DNA methylation and histone modifications. The possibility to re-express SOX11 in non-methylated tissue is of clinical relevance, and was successfully achieved in cell lines with low levels of SOX11 methylation. In breast cancer patients, methylation of the SOX11 promoter was shown to correlate with estrogen receptor status, suggesting that SOX11 may be functionally re-expressed during treatment with HDAC inhibitors in specific patient subgroups.

No MeSH data available.


Related in: MedlinePlus

Enrichment of H3K27me3 within the SOX11 promoter. Histone methylation of lysine 27 on histone 3 (H3K27me3) was assessed using chromatin immunoprecipitation and RT-qPCR for GAPDH (negative control), TSH2B (positive control) and SOX11. (A)Enrichment of H3K27me3 in unmethylated cell lines lacking SOX11. (B) Enrichment of H3K27me3 in methylated cell lines lacking SOX11.
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Fig5: Enrichment of H3K27me3 within the SOX11 promoter. Histone methylation of lysine 27 on histone 3 (H3K27me3) was assessed using chromatin immunoprecipitation and RT-qPCR for GAPDH (negative control), TSH2B (positive control) and SOX11. (A)Enrichment of H3K27me3 in unmethylated cell lines lacking SOX11. (B) Enrichment of H3K27me3 in methylated cell lines lacking SOX11.

Mentions: As discussed above, normal cells have a strong enrichment of the silencing histone mark H3K27me3 on the promoter of SOX11 but show a low degree of promoter methylation (Figure 1). In contrast, many neoplastic cell lines show a high degree of SOX11 promoter methylation (Figure 3). To investigate if neoplastic cell lines with a low degree of methylation depend on H3K27me3 to silence SOX11, cell lines with a low or high degree of methylation were investigated to determine the enrichment of H3K27me3 at the SOX11 promoter. The biological variation was significant, exemplified by the major variation in enrichment of the positive control, TSH2B. In two cell lines, DMS-114 and KCN-69n, the positive control showed such low levels of enrichment that data on SOX11 cannot be interpreted. GAPDH was used as a negative control and background levels were set to the largest observed GAPDH value. We show that H3K27me3 at the SOX11 promoter is enriched in several cell lines, including JIMT-1, LN-18 and JVM-2. However, SK-BR-3 and HS683 show a low enrichment compared to the positive control (TSH2B) and are likely dependent on other epigenetic regulation than promoter methylation or H3K27me3 to silence SOX11 (Figure 5A). For comparison, three methylated cell-lines were analyzed and all three cell lines, DOHH-2, RAJI and A2780-CP7 showed low enrichment of H3K27me3 at the SOX11 promoter compared to the positive control, indicating that methylation of the promoter may correlate to loss of repressive histone marks (Figure 5B).Figure 5


DNA methylation and histone modifications regulate SOX11 expression in lymphoid and solid cancer cells.

Nordström L, Andersson E, Kuci V, Gustavsson E, Holm K, Ringnér M, Guldberg P, Ek S - BMC Cancer (2015)

Enrichment of H3K27me3 within the SOX11 promoter. Histone methylation of lysine 27 on histone 3 (H3K27me3) was assessed using chromatin immunoprecipitation and RT-qPCR for GAPDH (negative control), TSH2B (positive control) and SOX11. (A)Enrichment of H3K27me3 in unmethylated cell lines lacking SOX11. (B) Enrichment of H3K27me3 in methylated cell lines lacking SOX11.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4403777&req=5

Fig5: Enrichment of H3K27me3 within the SOX11 promoter. Histone methylation of lysine 27 on histone 3 (H3K27me3) was assessed using chromatin immunoprecipitation and RT-qPCR for GAPDH (negative control), TSH2B (positive control) and SOX11. (A)Enrichment of H3K27me3 in unmethylated cell lines lacking SOX11. (B) Enrichment of H3K27me3 in methylated cell lines lacking SOX11.
Mentions: As discussed above, normal cells have a strong enrichment of the silencing histone mark H3K27me3 on the promoter of SOX11 but show a low degree of promoter methylation (Figure 1). In contrast, many neoplastic cell lines show a high degree of SOX11 promoter methylation (Figure 3). To investigate if neoplastic cell lines with a low degree of methylation depend on H3K27me3 to silence SOX11, cell lines with a low or high degree of methylation were investigated to determine the enrichment of H3K27me3 at the SOX11 promoter. The biological variation was significant, exemplified by the major variation in enrichment of the positive control, TSH2B. In two cell lines, DMS-114 and KCN-69n, the positive control showed such low levels of enrichment that data on SOX11 cannot be interpreted. GAPDH was used as a negative control and background levels were set to the largest observed GAPDH value. We show that H3K27me3 at the SOX11 promoter is enriched in several cell lines, including JIMT-1, LN-18 and JVM-2. However, SK-BR-3 and HS683 show a low enrichment compared to the positive control (TSH2B) and are likely dependent on other epigenetic regulation than promoter methylation or H3K27me3 to silence SOX11 (Figure 5A). For comparison, three methylated cell-lines were analyzed and all three cell lines, DOHH-2, RAJI and A2780-CP7 showed low enrichment of H3K27me3 at the SOX11 promoter compared to the positive control, indicating that methylation of the promoter may correlate to loss of repressive histone marks (Figure 5B).Figure 5

Bottom Line: In contrast, SOX11 regulation in neoplastic tissues is more complex involving both DNA methylation and histone modifications.The possibility to re-express SOX11 in non-methylated tissue is of clinical relevance, and was successfully achieved in cell lines with low levels of SOX11 methylation.In breast cancer patients, methylation of the SOX11 promoter was shown to correlate with estrogen receptor status, suggesting that SOX11 may be functionally re-expressed during treatment with HDAC inhibitors in specific patient subgroups.

View Article: PubMed Central - PubMed

Affiliation: Department of Immunotechnology, CREATE Health, Lund University, Lund, Sweden. lena.nordstrom@immun.lth.se.

ABSTRACT

Background: The neural transcription factor SOX11 is present at specific stages during embryo development with a very restricted expression in adult tissue, indicating precise regulation of transcription. SOX11 is strongly up-regulated in some malignancies and have a functional role in tumorgenesis. With the aim to explore differences in epigenetic regulation of SOX11 expression in normal versus neoplastic cells, we investigated methylation and histone modifications related to the SOX11 promoter and the possibility to induce re-expression using histone deacetylase (HDAC) or EZH2 inhibitors.

Methods: The epigenetic regulation of SOX11 was investigated in distinct non-malignant cell populations (n = 7) and neoplastic cell-lines (n = 42) of different cellular origins. DNA methylation was assessed using bisulfite sequencing, methylation-specific melting curve analysis, MethyLight and pyrosequencing. The presence of H3K27me3 was assessed using ChIP-qPCR. The HDAC inhibitors Vorinostat and trichostatin A were used to induce SOX11 in cell lines with no endogenous expression.

Results: The SOX11 promoter shows a low degree of methylation and strong enrichment of H3K27me3 in non-malignant differentiated cells, independent of cellular origin. Cancers of the B-cell lineage are strongly marked by de novo methylation at the SOX11 promoter in SOX11 non-expressing cells, while solid cancer entities display a more varying degree of SOX11 promoter methylation. The silencing mark H3K27me3 was generally present at the SOX11 promoter in non-expressing cells, and an increased enrichment was observed in cancer cells with a low degree of SOX11 methylation compared to cells with dense methylation. Finally, we demonstrate that the HDAC inhibitors (vorinostat and trichostatin A) induce SOX11 expression in cancer cells with low levels of SOX11 methylation.

Conclusions: We show that SOX11 is strongly marked by repressive histone marks in non-malignant cells. In contrast, SOX11 regulation in neoplastic tissues is more complex involving both DNA methylation and histone modifications. The possibility to re-express SOX11 in non-methylated tissue is of clinical relevance, and was successfully achieved in cell lines with low levels of SOX11 methylation. In breast cancer patients, methylation of the SOX11 promoter was shown to correlate with estrogen receptor status, suggesting that SOX11 may be functionally re-expressed during treatment with HDAC inhibitors in specific patient subgroups.

No MeSH data available.


Related in: MedlinePlus