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Down Regulation of CLDND1 Induces Apoptosis in Breast Cancer Cells.

Achari C, Winslow S, Larsson C - PLoS ONE (2015)

Bottom Line: Genes that were down regulated in all cell lines were further studied for survival-supporting effects.However, CLDND1 down regulation augmented ERK1/2 phosphorylation, which thereby may protect against the apoptosis inducing effects of CLDND1 down regulation.There is today little information on the function of CLDND1.

View Article: PubMed Central - PubMed

Affiliation: Translational Cancer Research, Department of Laboratory Medicine, Lund University, Lund, Sweden.

ABSTRACT
Identification of targets for apoptosis induction is important to provide novel therapeutic approaches in breast cancer. Our earlier studies showed that down regulation of protein kinase C δ (PKCδ) induces death in breast cancer cells. In this study we set out to identify previously unrecognized apoptosis regulators in breast cancer cells. To identify candidates, global expression analysis with microarray was performed after down regulation of PKCδ in the basal-like breast cancer cell lines MDA-MB-231, MDA-MB-468 and BT-549. Genes that were down regulated in all cell lines were further studied for survival-supporting effects. The claudin-like CLDND1 was singled out since several independent siRNAs targeting CLDND1 induced cell death in several cell lines. The cell death induced by CLDND1 knockdown was caspase-dependent, suggesting induction of apoptosis. Nuclear fragmentation, cleavage of caspase-3 and PARP and release of cytochrome C from the mitochondria upon CLDND1 depletion demonstrated involvement of the intrinsic apoptotic pathway. Inhibition of MEK1/2 and JNK further potentiated the cell death induction by CLDND1 knockdown. However, CLDND1 down regulation augmented ERK1/2 phosphorylation, which thereby may protect against the apoptosis inducing effects of CLDND1 down regulation. A concomitant inhibition of MEK1/2 suppresses the ERK1/2 phosphorylation and markedly potentiates the cell death following CLDND1 siRNA treatment. There is today little information on the function of CLDND1. These data provide novel information on CLDND1 and highlight it as a novel survival factor in basal-like breast cancer cell lines.

No MeSH data available.


Related in: MedlinePlus

Effects of silencing CLDND1, PLEKHA2, ACSL3, SLC30A9, MSN, CALM3 and PRKAR1A on breast cancer cell death.MDA-MB-231 (A), MDA-MB-468 (B) and BT-549 (C) breast cancer cells were transfected with siRNA oligonucleotides targeting CLDND1, PLEKHA2, ACSL3, SLC30A9, MSN, CALM3, PRKAR1A or with a control siRNA for 72 h. Cells were subjected to Annexin V-APC staining and flow cytometry analysis. Data (mean ± S.E.M., n = 3) represent percent Annexin V-positive cells related to control conditions. Asterisks indicate statistically significant differences (*, p<0.05; **, p<0.01) compared with control cells.
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pone.0130300.g001: Effects of silencing CLDND1, PLEKHA2, ACSL3, SLC30A9, MSN, CALM3 and PRKAR1A on breast cancer cell death.MDA-MB-231 (A), MDA-MB-468 (B) and BT-549 (C) breast cancer cells were transfected with siRNA oligonucleotides targeting CLDND1, PLEKHA2, ACSL3, SLC30A9, MSN, CALM3, PRKAR1A or with a control siRNA for 72 h. Cells were subjected to Annexin V-APC staining and flow cytometry analysis. Data (mean ± S.E.M., n = 3) represent percent Annexin V-positive cells related to control conditions. Asterisks indicate statistically significant differences (*, p<0.05; **, p<0.01) compared with control cells.

Mentions: To identify cell death regulators of breast cancer cell death, apoptosis was induced by treatment with a PKCδ siRNA, as previously shown [4], and the global gene expression was analyzed. Analysis of differential expression levels highlighted CLDND1, PLEKHA2, ACSL3, SLC30A9, MSN, CALM3 and PRKAR1A as potential regulators of breast cancer cell survival since they were affected by PKCδ siRNA in all cell lines (Table 3). To study their potential role in cell death the breast cancer cell lines were treated with siRNAs targeting the genes (Fig 1). CALM3 siRNA effectively increased cell death, measured as Annexin V positivity, in both MDA-MB-231 and MDA-MB-468 cells and had a similar tendency in BT-549 cells. The siRNA targeting CLDND1 also induced cell death in MDA-MB-468 cells and had a similar tendency in MDA-MB-231 and BT-549 cells. The siRNAs targeting the other candidate genes had less consistent cell-death inducing effects (Fig 1). Based on these data it is premature to conclusively exclude a role in cell survival for these genes. However, based on this initial screening CALM3 and CLDND1 were selected as the most promising candidates for further study.


Down Regulation of CLDND1 Induces Apoptosis in Breast Cancer Cells.

Achari C, Winslow S, Larsson C - PLoS ONE (2015)

Effects of silencing CLDND1, PLEKHA2, ACSL3, SLC30A9, MSN, CALM3 and PRKAR1A on breast cancer cell death.MDA-MB-231 (A), MDA-MB-468 (B) and BT-549 (C) breast cancer cells were transfected with siRNA oligonucleotides targeting CLDND1, PLEKHA2, ACSL3, SLC30A9, MSN, CALM3, PRKAR1A or with a control siRNA for 72 h. Cells were subjected to Annexin V-APC staining and flow cytometry analysis. Data (mean ± S.E.M., n = 3) represent percent Annexin V-positive cells related to control conditions. Asterisks indicate statistically significant differences (*, p<0.05; **, p<0.01) compared with control cells.
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4470986&req=5

pone.0130300.g001: Effects of silencing CLDND1, PLEKHA2, ACSL3, SLC30A9, MSN, CALM3 and PRKAR1A on breast cancer cell death.MDA-MB-231 (A), MDA-MB-468 (B) and BT-549 (C) breast cancer cells were transfected with siRNA oligonucleotides targeting CLDND1, PLEKHA2, ACSL3, SLC30A9, MSN, CALM3, PRKAR1A or with a control siRNA for 72 h. Cells were subjected to Annexin V-APC staining and flow cytometry analysis. Data (mean ± S.E.M., n = 3) represent percent Annexin V-positive cells related to control conditions. Asterisks indicate statistically significant differences (*, p<0.05; **, p<0.01) compared with control cells.
Mentions: To identify cell death regulators of breast cancer cell death, apoptosis was induced by treatment with a PKCδ siRNA, as previously shown [4], and the global gene expression was analyzed. Analysis of differential expression levels highlighted CLDND1, PLEKHA2, ACSL3, SLC30A9, MSN, CALM3 and PRKAR1A as potential regulators of breast cancer cell survival since they were affected by PKCδ siRNA in all cell lines (Table 3). To study their potential role in cell death the breast cancer cell lines were treated with siRNAs targeting the genes (Fig 1). CALM3 siRNA effectively increased cell death, measured as Annexin V positivity, in both MDA-MB-231 and MDA-MB-468 cells and had a similar tendency in BT-549 cells. The siRNA targeting CLDND1 also induced cell death in MDA-MB-468 cells and had a similar tendency in MDA-MB-231 and BT-549 cells. The siRNAs targeting the other candidate genes had less consistent cell-death inducing effects (Fig 1). Based on these data it is premature to conclusively exclude a role in cell survival for these genes. However, based on this initial screening CALM3 and CLDND1 were selected as the most promising candidates for further study.

Bottom Line: Genes that were down regulated in all cell lines were further studied for survival-supporting effects.However, CLDND1 down regulation augmented ERK1/2 phosphorylation, which thereby may protect against the apoptosis inducing effects of CLDND1 down regulation.There is today little information on the function of CLDND1.

View Article: PubMed Central - PubMed

Affiliation: Translational Cancer Research, Department of Laboratory Medicine, Lund University, Lund, Sweden.

ABSTRACT
Identification of targets for apoptosis induction is important to provide novel therapeutic approaches in breast cancer. Our earlier studies showed that down regulation of protein kinase C δ (PKCδ) induces death in breast cancer cells. In this study we set out to identify previously unrecognized apoptosis regulators in breast cancer cells. To identify candidates, global expression analysis with microarray was performed after down regulation of PKCδ in the basal-like breast cancer cell lines MDA-MB-231, MDA-MB-468 and BT-549. Genes that were down regulated in all cell lines were further studied for survival-supporting effects. The claudin-like CLDND1 was singled out since several independent siRNAs targeting CLDND1 induced cell death in several cell lines. The cell death induced by CLDND1 knockdown was caspase-dependent, suggesting induction of apoptosis. Nuclear fragmentation, cleavage of caspase-3 and PARP and release of cytochrome C from the mitochondria upon CLDND1 depletion demonstrated involvement of the intrinsic apoptotic pathway. Inhibition of MEK1/2 and JNK further potentiated the cell death induction by CLDND1 knockdown. However, CLDND1 down regulation augmented ERK1/2 phosphorylation, which thereby may protect against the apoptosis inducing effects of CLDND1 down regulation. A concomitant inhibition of MEK1/2 suppresses the ERK1/2 phosphorylation and markedly potentiates the cell death following CLDND1 siRNA treatment. There is today little information on the function of CLDND1. These data provide novel information on CLDND1 and highlight it as a novel survival factor in basal-like breast cancer cell lines.

No MeSH data available.


Related in: MedlinePlus