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Integrative genome-wide expression analysis bears evidence of estrogen receptor-independent transcription in heregulin-stimulated MCF-7 cells.

Nagashima T, Suzuki T, Kondo S, Kuroki Y, Takahashi K, Ide K, Yumoto N, Hasegawa A, Toyoda T, Kojima T, Konagaya A, Suzuki H, Hayashizaki Y, Sakaki Y, Hatakeyama M - PLoS ONE (2008)

Bottom Line: Through application of various threshold settings, 333 (326 up-regulated and 7 down-regulated) HRG-regulated genes were detected.However, with respect to enriched upstream TFs, 4 common TFs were identified in the 14 HRG-regulated (28.6%) and 13 E2-regulated (30.8%) gene groups.These results indicated that while E2 and HRG may induce common TFs, the regulatory mechanisms that govern HRG- and E2-induced gene expression differ.

View Article: PubMed Central - PubMed

Affiliation: Computational and Experimental Systems Biology Group, RIKEN Genomic Sciences Center, RIKEN Yokohama Institute, Yokohama, Japan.

ABSTRACT
Heregulin beta-1 (HRG) is an extracellular ligand that activates mitogen-activated protein kinase (MAPK) and phosphatidylinositol-3-OH kinase (PI3K)/Akt signaling pathways through ErbB receptors. MAPK and Akt have been shown to phosphorylate the estrogen receptor (ER) at Ser-118 and Ser-167, respectively, thereby mimicking the effects of estrogenic activity such as estrogen responsive element (ERE)-dependent transcription. In the current study, integrative analysis was performed using two tiling array platforms, comprising histone H3 lysine 9 (H3K9) acetylation and RNA mapping, together with array comparative genomic hybridization (CGH) analysis in an effort to identify HRG-regulated genes in ER-positive MCF-7 breast cancer cells. Through application of various threshold settings, 333 (326 up-regulated and 7 down-regulated) HRG-regulated genes were detected. Prediction of upstream transcription factors (TFs) and pathway analysis indicated that 21% of HRG-induced gene regulation may be controlled by the MAPK cascade, while only 0.6% of the gene expression is controlled by ERE. A comparison with previously reported estrogen (E2)-regulated gene expression data revealed that only 12 common genes were identified between the 333 HRG-regulated (3.6%) and 239 E2-regulated (5.0%) gene groups. However, with respect to enriched upstream TFs, 4 common TFs were identified in the 14 HRG-regulated (28.6%) and 13 E2-regulated (30.8%) gene groups. These results indicated that while E2 and HRG may induce common TFs, the regulatory mechanisms that govern HRG- and E2-induced gene expression differ.

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Comparison of tiling array-inferred up-regulated genes and expression array results at various thresholds.The proportion of genes that were identified as up-regulated and also found in the expression array-based gene list at various thresholds of α and β ((A) and (B)). (C) and (D) represent the number of identified genes.
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pone-0001803-g003: Comparison of tiling array-inferred up-regulated genes and expression array results at various thresholds.The proportion of genes that were identified as up-regulated and also found in the expression array-based gene list at various thresholds of α and β ((A) and (B)). (C) and (D) represent the number of identified genes.

Mentions: The threshold setting used to identify regulated genes from the above two integrated tiling array data sets was then examined. In this analysis, genes that only satisfied both Ahrg−Acontrol≥α and Rhrg−Rcontrol≥β were compared with results of the expression arrays. Figure 3 represents the percentage (A and B) and number (C and D) of genes that were identified as being up-regulated from the combined tiling array data and which were also found in the expression array results under various threshold (α and β) settings. As shown, increasing α (Figure 3A) and β (Figure 3B) yielded higher overlapping rates from the two tiling array data and expression array data sets. In contrast, increasing the threshold resulted in the identification of fewer genes (Figure 3C and D). β has a higher contribution to the precision rate and number of genes identified compared with α. For specific combinations of α and β (e.g. α = 4 and β = 4), 100% of the tiling array inferred that up-regulated genes were found in the expression array results. However, only 19 genes were identified in that setting. There is a trade-off between precision rate and the number of identified genes. Two parameters, rmin (minimum precision rate) and N (number of extracted genes), need to be determined in order to arrive at a reasonable threshold setting. Here, rmin and N were set to 70 and 300, respectively. Under these conditions, the largest β value was searched for and α was then determined since a larger threshold results in a higher precision rate. Finally, α = 4 and β = 2 was obtained and these values were used as thresholds. As a result, 326 and 7 genes were identified as being up- and down-regulated, respectively, following HRG treatment of cells (Table S1). With this setting, the up- and down-regulation of 72.4% of these transcripts (241 out of 333 genes) was confirmed with the expression array data.


Integrative genome-wide expression analysis bears evidence of estrogen receptor-independent transcription in heregulin-stimulated MCF-7 cells.

Nagashima T, Suzuki T, Kondo S, Kuroki Y, Takahashi K, Ide K, Yumoto N, Hasegawa A, Toyoda T, Kojima T, Konagaya A, Suzuki H, Hayashizaki Y, Sakaki Y, Hatakeyama M - PLoS ONE (2008)

Comparison of tiling array-inferred up-regulated genes and expression array results at various thresholds.The proportion of genes that were identified as up-regulated and also found in the expression array-based gene list at various thresholds of α and β ((A) and (B)). (C) and (D) represent the number of identified genes.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2266794&req=5

pone-0001803-g003: Comparison of tiling array-inferred up-regulated genes and expression array results at various thresholds.The proportion of genes that were identified as up-regulated and also found in the expression array-based gene list at various thresholds of α and β ((A) and (B)). (C) and (D) represent the number of identified genes.
Mentions: The threshold setting used to identify regulated genes from the above two integrated tiling array data sets was then examined. In this analysis, genes that only satisfied both Ahrg−Acontrol≥α and Rhrg−Rcontrol≥β were compared with results of the expression arrays. Figure 3 represents the percentage (A and B) and number (C and D) of genes that were identified as being up-regulated from the combined tiling array data and which were also found in the expression array results under various threshold (α and β) settings. As shown, increasing α (Figure 3A) and β (Figure 3B) yielded higher overlapping rates from the two tiling array data and expression array data sets. In contrast, increasing the threshold resulted in the identification of fewer genes (Figure 3C and D). β has a higher contribution to the precision rate and number of genes identified compared with α. For specific combinations of α and β (e.g. α = 4 and β = 4), 100% of the tiling array inferred that up-regulated genes were found in the expression array results. However, only 19 genes were identified in that setting. There is a trade-off between precision rate and the number of identified genes. Two parameters, rmin (minimum precision rate) and N (number of extracted genes), need to be determined in order to arrive at a reasonable threshold setting. Here, rmin and N were set to 70 and 300, respectively. Under these conditions, the largest β value was searched for and α was then determined since a larger threshold results in a higher precision rate. Finally, α = 4 and β = 2 was obtained and these values were used as thresholds. As a result, 326 and 7 genes were identified as being up- and down-regulated, respectively, following HRG treatment of cells (Table S1). With this setting, the up- and down-regulation of 72.4% of these transcripts (241 out of 333 genes) was confirmed with the expression array data.

Bottom Line: Through application of various threshold settings, 333 (326 up-regulated and 7 down-regulated) HRG-regulated genes were detected.However, with respect to enriched upstream TFs, 4 common TFs were identified in the 14 HRG-regulated (28.6%) and 13 E2-regulated (30.8%) gene groups.These results indicated that while E2 and HRG may induce common TFs, the regulatory mechanisms that govern HRG- and E2-induced gene expression differ.

View Article: PubMed Central - PubMed

Affiliation: Computational and Experimental Systems Biology Group, RIKEN Genomic Sciences Center, RIKEN Yokohama Institute, Yokohama, Japan.

ABSTRACT
Heregulin beta-1 (HRG) is an extracellular ligand that activates mitogen-activated protein kinase (MAPK) and phosphatidylinositol-3-OH kinase (PI3K)/Akt signaling pathways through ErbB receptors. MAPK and Akt have been shown to phosphorylate the estrogen receptor (ER) at Ser-118 and Ser-167, respectively, thereby mimicking the effects of estrogenic activity such as estrogen responsive element (ERE)-dependent transcription. In the current study, integrative analysis was performed using two tiling array platforms, comprising histone H3 lysine 9 (H3K9) acetylation and RNA mapping, together with array comparative genomic hybridization (CGH) analysis in an effort to identify HRG-regulated genes in ER-positive MCF-7 breast cancer cells. Through application of various threshold settings, 333 (326 up-regulated and 7 down-regulated) HRG-regulated genes were detected. Prediction of upstream transcription factors (TFs) and pathway analysis indicated that 21% of HRG-induced gene regulation may be controlled by the MAPK cascade, while only 0.6% of the gene expression is controlled by ERE. A comparison with previously reported estrogen (E2)-regulated gene expression data revealed that only 12 common genes were identified between the 333 HRG-regulated (3.6%) and 239 E2-regulated (5.0%) gene groups. However, with respect to enriched upstream TFs, 4 common TFs were identified in the 14 HRG-regulated (28.6%) and 13 E2-regulated (30.8%) gene groups. These results indicated that while E2 and HRG may induce common TFs, the regulatory mechanisms that govern HRG- and E2-induced gene expression differ.

Show MeSH
Related in: MedlinePlus