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BRCA1 regulates PIG3-mediated apoptosis in a p53-dependent manner.

Zhang W, Luo J, Chen F, Yang F, Song W, Zhu A, Guan X - Oncotarget (2015)

Bottom Line: Moreover, we reveal that overexpression of BRCA1 significantly increased expression of PIG3 in cells with intact p53, whereas no increase in PIG3 was observed in p53- MDA-MB-157 cells and p53-depleted HCT116p53-/- cells.Meanwhile, ectopic expression of BRCA1 could not lead to an increase expression level of prohibitin (PHB), which we have previously identified to induce PIG3-mediated apoptosis.Finally, ChIP analysis revealed that PHB can bind to the PIG3 promoter and activate PIG3 transcription independent of p53, although p53 presence did enhance this process.

View Article: PubMed Central - PubMed

Affiliation: Department of Medical Oncology, Jinling Hospital, Medical School of Nanjing University, Nanjing 210002, China.

ABSTRACT
BRCA1 plays a key role in the regulation of p53-dependent target gene transcription activation. Meanwhile, the p53 inducible gene 3 (PIG3) is a downstream target of p53 and is involved in p53-initiated apoptosis. However, little is known about whether BRCA1 can regulate PIG3-mediated apoptosis. Using a tissue microarray containing 149 breast cancer patient samples, we found that BRCA1 and PIG3 expression status were significantly positively correlated (r = 0.678, P < 0.001) and identified a significant positive correlation between high expression of BRCA1 and/or PIG3 and overall survival (OS). Moreover, we reveal that overexpression of BRCA1 significantly increased expression of PIG3 in cells with intact p53, whereas no increase in PIG3 was observed in p53- MDA-MB-157 cells and p53-depleted HCT116p53-/- cells. Meanwhile, ectopic expression of BRCA1 could not lead to an increase expression level of prohibitin (PHB), which we have previously identified to induce PIG3-mediated apoptosis. Finally, ChIP analysis revealed that PHB can bind to the PIG3 promoter and activate PIG3 transcription independent of p53, although p53 presence did enhance this process. Taken together, our findings suggest that BRCA1 regulates PIG3-mediated apoptosis in a p53-dependent manner, and that PIG3 expression is associated with a better OS in breast cancer patients.

No MeSH data available.


Related in: MedlinePlus

BRCA1 positively regulates PIG3 expression in a p53-dependent manner(A and B) Cell viability was measured by MTT assay, and apoptosis was detected by flow cytometry following transfection of plasmid encoding BRCA1 into MCF-7, T47D, SK-BR-3, and MDA-MB-157 cells. (C) BRCA1, p53, and PIG3 protein levels were determined by western blotting following transfection of plasmid encoding BRCA1 into MCF-7, T47D, SK-BR-3, and MDA-MB-157 cells, and normalized to GAPDH expression. (D) BRCA1, p53, and PIG3 mRNA expression levels were determined by RT-PCR following transfection of plasmid encoding BRCA1 into HCT116p53+/+ and HCT116p53−/− cells, and normalized to GAPDH expression. (E and F) Localization and expression of p53 and PIG3 were determined by fluorescence microscopy following the same treatment as (D), with DMSO treatment as a control. Nuclei were stained with DAPI. The high magnification (200×) regions were shown above. (G) Relative levels of PIG3 mRNA in 206 breast cancer cell samples with no BRCA1 mutation (WT) and 392 breast cancer cell samples with a BRCA1 mutation (MT). Data analyzed using Oncomine (www.oncomine.org) from original published data (Garnett et al., 2012). Data are the mean of three independent experiments. *P < 0.05, as compared with untreated cells.
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Figure 2: BRCA1 positively regulates PIG3 expression in a p53-dependent manner(A and B) Cell viability was measured by MTT assay, and apoptosis was detected by flow cytometry following transfection of plasmid encoding BRCA1 into MCF-7, T47D, SK-BR-3, and MDA-MB-157 cells. (C) BRCA1, p53, and PIG3 protein levels were determined by western blotting following transfection of plasmid encoding BRCA1 into MCF-7, T47D, SK-BR-3, and MDA-MB-157 cells, and normalized to GAPDH expression. (D) BRCA1, p53, and PIG3 mRNA expression levels were determined by RT-PCR following transfection of plasmid encoding BRCA1 into HCT116p53+/+ and HCT116p53−/− cells, and normalized to GAPDH expression. (E and F) Localization and expression of p53 and PIG3 were determined by fluorescence microscopy following the same treatment as (D), with DMSO treatment as a control. Nuclei were stained with DAPI. The high magnification (200×) regions were shown above. (G) Relative levels of PIG3 mRNA in 206 breast cancer cell samples with no BRCA1 mutation (WT) and 392 breast cancer cell samples with a BRCA1 mutation (MT). Data analyzed using Oncomine (www.oncomine.org) from original published data (Garnett et al., 2012). Data are the mean of three independent experiments. *P < 0.05, as compared with untreated cells.

Mentions: Mutations of the BRCA1 gene confer a high risk for breast cancers [14]. Here, we found that in breast cancer cells, overexpression of BRCA1 inhibited cell proliferation and induced apoptosis in p53-expressing cell lines, while the effect of BRCA1 was attenuated in p53- cells, MDA-MB-157 (Figure 2A and 2B) and validated in HCT116p53−/− cells (Supplementary Figure S1A–S1C).


BRCA1 regulates PIG3-mediated apoptosis in a p53-dependent manner.

Zhang W, Luo J, Chen F, Yang F, Song W, Zhu A, Guan X - Oncotarget (2015)

BRCA1 positively regulates PIG3 expression in a p53-dependent manner(A and B) Cell viability was measured by MTT assay, and apoptosis was detected by flow cytometry following transfection of plasmid encoding BRCA1 into MCF-7, T47D, SK-BR-3, and MDA-MB-157 cells. (C) BRCA1, p53, and PIG3 protein levels were determined by western blotting following transfection of plasmid encoding BRCA1 into MCF-7, T47D, SK-BR-3, and MDA-MB-157 cells, and normalized to GAPDH expression. (D) BRCA1, p53, and PIG3 mRNA expression levels were determined by RT-PCR following transfection of plasmid encoding BRCA1 into HCT116p53+/+ and HCT116p53−/− cells, and normalized to GAPDH expression. (E and F) Localization and expression of p53 and PIG3 were determined by fluorescence microscopy following the same treatment as (D), with DMSO treatment as a control. Nuclei were stained with DAPI. The high magnification (200×) regions were shown above. (G) Relative levels of PIG3 mRNA in 206 breast cancer cell samples with no BRCA1 mutation (WT) and 392 breast cancer cell samples with a BRCA1 mutation (MT). Data analyzed using Oncomine (www.oncomine.org) from original published data (Garnett et al., 2012). Data are the mean of three independent experiments. *P < 0.05, as compared with untreated cells.
© Copyright Policy - open-access
Related In: Results  -  Collection

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Figure 2: BRCA1 positively regulates PIG3 expression in a p53-dependent manner(A and B) Cell viability was measured by MTT assay, and apoptosis was detected by flow cytometry following transfection of plasmid encoding BRCA1 into MCF-7, T47D, SK-BR-3, and MDA-MB-157 cells. (C) BRCA1, p53, and PIG3 protein levels were determined by western blotting following transfection of plasmid encoding BRCA1 into MCF-7, T47D, SK-BR-3, and MDA-MB-157 cells, and normalized to GAPDH expression. (D) BRCA1, p53, and PIG3 mRNA expression levels were determined by RT-PCR following transfection of plasmid encoding BRCA1 into HCT116p53+/+ and HCT116p53−/− cells, and normalized to GAPDH expression. (E and F) Localization and expression of p53 and PIG3 were determined by fluorescence microscopy following the same treatment as (D), with DMSO treatment as a control. Nuclei were stained with DAPI. The high magnification (200×) regions were shown above. (G) Relative levels of PIG3 mRNA in 206 breast cancer cell samples with no BRCA1 mutation (WT) and 392 breast cancer cell samples with a BRCA1 mutation (MT). Data analyzed using Oncomine (www.oncomine.org) from original published data (Garnett et al., 2012). Data are the mean of three independent experiments. *P < 0.05, as compared with untreated cells.
Mentions: Mutations of the BRCA1 gene confer a high risk for breast cancers [14]. Here, we found that in breast cancer cells, overexpression of BRCA1 inhibited cell proliferation and induced apoptosis in p53-expressing cell lines, while the effect of BRCA1 was attenuated in p53- cells, MDA-MB-157 (Figure 2A and 2B) and validated in HCT116p53−/− cells (Supplementary Figure S1A–S1C).

Bottom Line: Moreover, we reveal that overexpression of BRCA1 significantly increased expression of PIG3 in cells with intact p53, whereas no increase in PIG3 was observed in p53- MDA-MB-157 cells and p53-depleted HCT116p53-/- cells.Meanwhile, ectopic expression of BRCA1 could not lead to an increase expression level of prohibitin (PHB), which we have previously identified to induce PIG3-mediated apoptosis.Finally, ChIP analysis revealed that PHB can bind to the PIG3 promoter and activate PIG3 transcription independent of p53, although p53 presence did enhance this process.

View Article: PubMed Central - PubMed

Affiliation: Department of Medical Oncology, Jinling Hospital, Medical School of Nanjing University, Nanjing 210002, China.

ABSTRACT
BRCA1 plays a key role in the regulation of p53-dependent target gene transcription activation. Meanwhile, the p53 inducible gene 3 (PIG3) is a downstream target of p53 and is involved in p53-initiated apoptosis. However, little is known about whether BRCA1 can regulate PIG3-mediated apoptosis. Using a tissue microarray containing 149 breast cancer patient samples, we found that BRCA1 and PIG3 expression status were significantly positively correlated (r = 0.678, P < 0.001) and identified a significant positive correlation between high expression of BRCA1 and/or PIG3 and overall survival (OS). Moreover, we reveal that overexpression of BRCA1 significantly increased expression of PIG3 in cells with intact p53, whereas no increase in PIG3 was observed in p53- MDA-MB-157 cells and p53-depleted HCT116p53-/- cells. Meanwhile, ectopic expression of BRCA1 could not lead to an increase expression level of prohibitin (PHB), which we have previously identified to induce PIG3-mediated apoptosis. Finally, ChIP analysis revealed that PHB can bind to the PIG3 promoter and activate PIG3 transcription independent of p53, although p53 presence did enhance this process. Taken together, our findings suggest that BRCA1 regulates PIG3-mediated apoptosis in a p53-dependent manner, and that PIG3 expression is associated with a better OS in breast cancer patients.

No MeSH data available.


Related in: MedlinePlus