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Src/STAT3-dependent heme oxygenase-1 induction mediates chemoresistance of breast cancer cells to doxorubicin by promoting autophagy.

Tan Q, Wang H, Hu Y, Hu M, Li X - Cancer Sci. (2015)

Bottom Line: In the present study, we found an induction of HO-1 expression in doxorubicin (DOX)-treated MDA-MB-231 human breast adenocarcinoma cells, which showed insensitivity to DOX treatment.Knockdown HO-1 expression dramatically upregulated the incidence of MDA-MB-231 cell death under DOX treatment, indicating that HO-1 functions as a critical contributor to drug resistance in MDA-MB-231 cells.Moreover, upregulation of HO-1 expression required the activation of both signal transducer and activator of transcription (STAT)3 and its upstream regulator, protein kinase Src.

View Article: PubMed Central - PubMed

Affiliation: Department of Stress Medicine, Beijing Institute of Basic Medical Sciences, Beijing, China.

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STAT-3 was responsible for heme oxygenase-1 (HO-1) induction under doxorubicin (DOX) exposure. (a) MDA-MB-231 and MDA-MB-453 cells were treated with DOX (0.2 μM) for the indicated time periods and then the induction of STAT3 activation was detected by western-blot assay. (b) MDA-MB-231 cells were transfected with STAT3-dependent luciferase reporter plasmid and then exposed to DOX (0.2 μM) 36 h after transfection. Then the induction of STAT3 luciferase activities was determined at the indicated time periods after DOX treatment. (c) MDA-MB-231 cells were transfected with STAT3 siRNA or control siRNA and then treated with DOX (0.2 μM) 36 h after transfection. Then the expressions of HO-1, Beclin-1 and LC3BI/II were determined 12 h after DOX exposure. To avoid the off-target effect, two different siRNA against STAT3 were purchased and tested. Data shown here were obtained from the most effective one. (d,e) MDA-MB-231 cells were transfected and treated as described in figure (c) and then autophagy was detected as described in Figure3(b,c). (f) MDA-MB-231 cells were transfected and treated as described in figure (c) and then cell death incidence was detected 48 h after DOX treatment.
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fig04: STAT-3 was responsible for heme oxygenase-1 (HO-1) induction under doxorubicin (DOX) exposure. (a) MDA-MB-231 and MDA-MB-453 cells were treated with DOX (0.2 μM) for the indicated time periods and then the induction of STAT3 activation was detected by western-blot assay. (b) MDA-MB-231 cells were transfected with STAT3-dependent luciferase reporter plasmid and then exposed to DOX (0.2 μM) 36 h after transfection. Then the induction of STAT3 luciferase activities was determined at the indicated time periods after DOX treatment. (c) MDA-MB-231 cells were transfected with STAT3 siRNA or control siRNA and then treated with DOX (0.2 μM) 36 h after transfection. Then the expressions of HO-1, Beclin-1 and LC3BI/II were determined 12 h after DOX exposure. To avoid the off-target effect, two different siRNA against STAT3 were purchased and tested. Data shown here were obtained from the most effective one. (d,e) MDA-MB-231 cells were transfected and treated as described in figure (c) and then autophagy was detected as described in Figure3(b,c). (f) MDA-MB-231 cells were transfected and treated as described in figure (c) and then cell death incidence was detected 48 h after DOX treatment.

Mentions: After revealing HO-1-dependent autophagy in mediating the cytoprotective effect, we next focused on elucidating the upstream signaling events leading to HO-1 induction in DOX-induced responses. According to previous reports, induction of HO-1 expression occurs exclusively at the transcriptional levels, which potentially involves the coordinated interaction of multiple transcription factors, such as Nrf2, NF-κB, Egr-1, AP-1 and STAT3.26–29 However, the functional significance of each factor is different under various stress conditions. Therefore, in the next study, we compared the activation status of the potential transcriptional factors involved in regulating HO-1 induction in DOX-treated MDA-MB-231 and MDA-MB-453 cells. We found that STAT3 was strongly activated in MDA-MB-231 cells after treatment with DOX, evidenced by dramatic upregulation of levels on tyrosine705-phosphorylated STAT3. No signaling indicating STAT3 activation was observed in MDA-MB-453 cells under the same DOX exposure conditions (Fig.4a). Furthermore, an enhancement of STAT3-dependent luciferase activities was readily detected in MDA-MB-231 cells after DOX stimulation (Fig.4b). These data indicate that although MDA-MB-231 cells are not sensitive to DOX treatment, DOX induces upregulation of STAT3 transcriptional activities in these cells.


Src/STAT3-dependent heme oxygenase-1 induction mediates chemoresistance of breast cancer cells to doxorubicin by promoting autophagy.

Tan Q, Wang H, Hu Y, Hu M, Li X - Cancer Sci. (2015)

STAT-3 was responsible for heme oxygenase-1 (HO-1) induction under doxorubicin (DOX) exposure. (a) MDA-MB-231 and MDA-MB-453 cells were treated with DOX (0.2 μM) for the indicated time periods and then the induction of STAT3 activation was detected by western-blot assay. (b) MDA-MB-231 cells were transfected with STAT3-dependent luciferase reporter plasmid and then exposed to DOX (0.2 μM) 36 h after transfection. Then the induction of STAT3 luciferase activities was determined at the indicated time periods after DOX treatment. (c) MDA-MB-231 cells were transfected with STAT3 siRNA or control siRNA and then treated with DOX (0.2 μM) 36 h after transfection. Then the expressions of HO-1, Beclin-1 and LC3BI/II were determined 12 h after DOX exposure. To avoid the off-target effect, two different siRNA against STAT3 were purchased and tested. Data shown here were obtained from the most effective one. (d,e) MDA-MB-231 cells were transfected and treated as described in figure (c) and then autophagy was detected as described in Figure3(b,c). (f) MDA-MB-231 cells were transfected and treated as described in figure (c) and then cell death incidence was detected 48 h after DOX treatment.
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fig04: STAT-3 was responsible for heme oxygenase-1 (HO-1) induction under doxorubicin (DOX) exposure. (a) MDA-MB-231 and MDA-MB-453 cells were treated with DOX (0.2 μM) for the indicated time periods and then the induction of STAT3 activation was detected by western-blot assay. (b) MDA-MB-231 cells were transfected with STAT3-dependent luciferase reporter plasmid and then exposed to DOX (0.2 μM) 36 h after transfection. Then the induction of STAT3 luciferase activities was determined at the indicated time periods after DOX treatment. (c) MDA-MB-231 cells were transfected with STAT3 siRNA or control siRNA and then treated with DOX (0.2 μM) 36 h after transfection. Then the expressions of HO-1, Beclin-1 and LC3BI/II were determined 12 h after DOX exposure. To avoid the off-target effect, two different siRNA against STAT3 were purchased and tested. Data shown here were obtained from the most effective one. (d,e) MDA-MB-231 cells were transfected and treated as described in figure (c) and then autophagy was detected as described in Figure3(b,c). (f) MDA-MB-231 cells were transfected and treated as described in figure (c) and then cell death incidence was detected 48 h after DOX treatment.
Mentions: After revealing HO-1-dependent autophagy in mediating the cytoprotective effect, we next focused on elucidating the upstream signaling events leading to HO-1 induction in DOX-induced responses. According to previous reports, induction of HO-1 expression occurs exclusively at the transcriptional levels, which potentially involves the coordinated interaction of multiple transcription factors, such as Nrf2, NF-κB, Egr-1, AP-1 and STAT3.26–29 However, the functional significance of each factor is different under various stress conditions. Therefore, in the next study, we compared the activation status of the potential transcriptional factors involved in regulating HO-1 induction in DOX-treated MDA-MB-231 and MDA-MB-453 cells. We found that STAT3 was strongly activated in MDA-MB-231 cells after treatment with DOX, evidenced by dramatic upregulation of levels on tyrosine705-phosphorylated STAT3. No signaling indicating STAT3 activation was observed in MDA-MB-453 cells under the same DOX exposure conditions (Fig.4a). Furthermore, an enhancement of STAT3-dependent luciferase activities was readily detected in MDA-MB-231 cells after DOX stimulation (Fig.4b). These data indicate that although MDA-MB-231 cells are not sensitive to DOX treatment, DOX induces upregulation of STAT3 transcriptional activities in these cells.

Bottom Line: In the present study, we found an induction of HO-1 expression in doxorubicin (DOX)-treated MDA-MB-231 human breast adenocarcinoma cells, which showed insensitivity to DOX treatment.Knockdown HO-1 expression dramatically upregulated the incidence of MDA-MB-231 cell death under DOX treatment, indicating that HO-1 functions as a critical contributor to drug resistance in MDA-MB-231 cells.Moreover, upregulation of HO-1 expression required the activation of both signal transducer and activator of transcription (STAT)3 and its upstream regulator, protein kinase Src.

View Article: PubMed Central - PubMed

Affiliation: Department of Stress Medicine, Beijing Institute of Basic Medical Sciences, Beijing, China.

Show MeSH
Related in: MedlinePlus