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Effect of hypoxia-inducible factor-1alpha on transcription of survivin in non-small cell lung cancer.

Chen YQ, Zhao CL, Li W - J. Exp. Clin. Cancer Res. (2009)

Bottom Line: Site directed mutagenesis of the putative binding site for HIF-1alpha in the survivin promoter significantly decreased the activity of the survivin promoter in A549 cells.Inhibition of HIF-1alpha by RNAi decreased the expression of survivin in A549 cell lines.Our results indicate that the binding of HIF-1alpha to the survivin promoter increases transcription of the survivin gene.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Respiration, First Affiliated Hospital of Bengbu Medical College, Bengbu, Anhui, PR China. bbmccyq@126.com

ABSTRACT

Background: Survivin is a structurally and functionally unique member of the inhibitor of apoptosis protein (IAP) family. It plays an important role, not only in regulating mitosis but also in inhibiting apoptosis. The current literature contains few reports on the transcriptional regulation of survivin expression in lung cancer.

Methods: In this study, we investigated the effect of hypoxia-inducible factor-1alpha (HIF-1alpha) on the transcriptional activity of the survivin promoter in non-small cell lung cancer (NSCLC). Immunohistochemical staining was used to detect the expression of survivin and HIF-1alpha in the lung tissue of 120 patients with non-small cell lung cancer (NSCLC) and 40 patients with benign pulmonary disease. We also performed experiments with the lung adenocarcinoma cell line A549 cells, which were cultured under hypoxic conditions. The expression of survivin and HIF-1alpha was detected by real-time RT-PCR and Western blotting. In the survivin promoter the putative binding-site for HIF-1alpha, is -19 bpapproximately -16 bp upstream of TSS. We performed site-directed mutagenesis of this binding site, and used luciferase reporter plasmids to determine the relative activity of the survivin promoter in A549 cells. We also studied the effect of HIF-1alpha on the expression of survivin by dsRNA targeting of HIF-1alpha mRNA.

Results: HIF-1alpha (58.33%) and survivin (81.60%) were both over-expressed in NSCLC and their expressions correlated with one another. They were also expressed in A549 cells under normal and hypoxic conditions, with a significant increase under hypoxic conditions. Site directed mutagenesis of the putative binding site for HIF-1alpha in the survivin promoter significantly decreased the activity of the survivin promoter in A549 cells. Inhibition of HIF-1alpha by RNAi decreased the expression of survivin in A549 cell lines.

Conclusion: Our results indicate that the binding of HIF-1alpha to the survivin promoter increases transcription of the survivin gene. Thus, HIF-1alpha is an important transcriptional regulator of survivin expression.

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Expression of survivin and HIF-1α in NSCLC and benign lung disease tissues. Survivin and HIF-lα protein were detected and localised within paraffin-embedded human lung tissue using immunohistochemistry. A and B represent the negative expression of survivin protein and HIF-1α protein, respectively, in benign lung disease tissues. C and D represent the positive expression (arrow) of survivin protein and HIF-1α protein, respectively, in NSCLC,. E: The graph shows the statistical results. 81.60% (98/120) of lung cancer tissue samples were positive for survivin staining, and 58.33% (70/120_) of lung cancer tissue samples were positive for HIF-1α staining. ** p < 0.01.
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Figure 1: Expression of survivin and HIF-1α in NSCLC and benign lung disease tissues. Survivin and HIF-lα protein were detected and localised within paraffin-embedded human lung tissue using immunohistochemistry. A and B represent the negative expression of survivin protein and HIF-1α protein, respectively, in benign lung disease tissues. C and D represent the positive expression (arrow) of survivin protein and HIF-1α protein, respectively, in NSCLC,. E: The graph shows the statistical results. 81.60% (98/120) of lung cancer tissue samples were positive for survivin staining, and 58.33% (70/120_) of lung cancer tissue samples were positive for HIF-1α staining. ** p < 0.01.

Mentions: Survivin and HIF-lα proteins were detected and localised in paraffin-embedded human lung tissue sections using immunohistochemistry. Survivin was predominantly expressed in the cytosol of the tumour cells with some nuclear staining (Fig. 1C). Survivin was exclusively expressed in lung cancer tissue (Fig. 1C, E, 81.60%,) and not in benign lung disease tissue (Fig. 1A, E, 18.4%). The specificity of survivin protein in lung cancer was 100%. HIF-lα was found primarily in the cytosol of lung cancer cells, with some nuclear staining (Fig. 1D). Positive rate of HIF-lα in lung cancer tissue samples was 58.33% (70/120), higher than that in tissue samples from benign lung disease (10%, 4/40) (Fig. 1B, E, p < 0. 01). The expression of survivin or HIF-1α in NSCLC was not correlated with age or sex, but with differentiation grade, lymph node metastasis and disease stages (Table 1). Spearman correlation analysis showed a correlation between the expression of survivin and the expression of HIF-1α in (rs = 0.255, p = 0.005) (Table 1).


Effect of hypoxia-inducible factor-1alpha on transcription of survivin in non-small cell lung cancer.

Chen YQ, Zhao CL, Li W - J. Exp. Clin. Cancer Res. (2009)

Expression of survivin and HIF-1α in NSCLC and benign lung disease tissues. Survivin and HIF-lα protein were detected and localised within paraffin-embedded human lung tissue using immunohistochemistry. A and B represent the negative expression of survivin protein and HIF-1α protein, respectively, in benign lung disease tissues. C and D represent the positive expression (arrow) of survivin protein and HIF-1α protein, respectively, in NSCLC,. E: The graph shows the statistical results. 81.60% (98/120) of lung cancer tissue samples were positive for survivin staining, and 58.33% (70/120_) of lung cancer tissue samples were positive for HIF-1α staining. ** p < 0.01.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2663545&req=5

Figure 1: Expression of survivin and HIF-1α in NSCLC and benign lung disease tissues. Survivin and HIF-lα protein were detected and localised within paraffin-embedded human lung tissue using immunohistochemistry. A and B represent the negative expression of survivin protein and HIF-1α protein, respectively, in benign lung disease tissues. C and D represent the positive expression (arrow) of survivin protein and HIF-1α protein, respectively, in NSCLC,. E: The graph shows the statistical results. 81.60% (98/120) of lung cancer tissue samples were positive for survivin staining, and 58.33% (70/120_) of lung cancer tissue samples were positive for HIF-1α staining. ** p < 0.01.
Mentions: Survivin and HIF-lα proteins were detected and localised in paraffin-embedded human lung tissue sections using immunohistochemistry. Survivin was predominantly expressed in the cytosol of the tumour cells with some nuclear staining (Fig. 1C). Survivin was exclusively expressed in lung cancer tissue (Fig. 1C, E, 81.60%,) and not in benign lung disease tissue (Fig. 1A, E, 18.4%). The specificity of survivin protein in lung cancer was 100%. HIF-lα was found primarily in the cytosol of lung cancer cells, with some nuclear staining (Fig. 1D). Positive rate of HIF-lα in lung cancer tissue samples was 58.33% (70/120), higher than that in tissue samples from benign lung disease (10%, 4/40) (Fig. 1B, E, p < 0. 01). The expression of survivin or HIF-1α in NSCLC was not correlated with age or sex, but with differentiation grade, lymph node metastasis and disease stages (Table 1). Spearman correlation analysis showed a correlation between the expression of survivin and the expression of HIF-1α in (rs = 0.255, p = 0.005) (Table 1).

Bottom Line: Site directed mutagenesis of the putative binding site for HIF-1alpha in the survivin promoter significantly decreased the activity of the survivin promoter in A549 cells.Inhibition of HIF-1alpha by RNAi decreased the expression of survivin in A549 cell lines.Our results indicate that the binding of HIF-1alpha to the survivin promoter increases transcription of the survivin gene.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Respiration, First Affiliated Hospital of Bengbu Medical College, Bengbu, Anhui, PR China. bbmccyq@126.com

ABSTRACT

Background: Survivin is a structurally and functionally unique member of the inhibitor of apoptosis protein (IAP) family. It plays an important role, not only in regulating mitosis but also in inhibiting apoptosis. The current literature contains few reports on the transcriptional regulation of survivin expression in lung cancer.

Methods: In this study, we investigated the effect of hypoxia-inducible factor-1alpha (HIF-1alpha) on the transcriptional activity of the survivin promoter in non-small cell lung cancer (NSCLC). Immunohistochemical staining was used to detect the expression of survivin and HIF-1alpha in the lung tissue of 120 patients with non-small cell lung cancer (NSCLC) and 40 patients with benign pulmonary disease. We also performed experiments with the lung adenocarcinoma cell line A549 cells, which were cultured under hypoxic conditions. The expression of survivin and HIF-1alpha was detected by real-time RT-PCR and Western blotting. In the survivin promoter the putative binding-site for HIF-1alpha, is -19 bpapproximately -16 bp upstream of TSS. We performed site-directed mutagenesis of this binding site, and used luciferase reporter plasmids to determine the relative activity of the survivin promoter in A549 cells. We also studied the effect of HIF-1alpha on the expression of survivin by dsRNA targeting of HIF-1alpha mRNA.

Results: HIF-1alpha (58.33%) and survivin (81.60%) were both over-expressed in NSCLC and their expressions correlated with one another. They were also expressed in A549 cells under normal and hypoxic conditions, with a significant increase under hypoxic conditions. Site directed mutagenesis of the putative binding site for HIF-1alpha in the survivin promoter significantly decreased the activity of the survivin promoter in A549 cells. Inhibition of HIF-1alpha by RNAi decreased the expression of survivin in A549 cell lines.

Conclusion: Our results indicate that the binding of HIF-1alpha to the survivin promoter increases transcription of the survivin gene. Thus, HIF-1alpha is an important transcriptional regulator of survivin expression.

Show MeSH
Related in: MedlinePlus