(Frontiers in Bioscience 14, 2514-2521, January 1, 2009)

Silencing of TGase 2 sensitizes breast cancer cells to apoptosis by regulation of survival factors

Dae-Seok Kim1, Kang-Seo Park1, Soo-Youl Kim1

1Molecular Oncology Branch, Division of Basic and Applied Science, Research Institute, National Cancer Center, Goyang, Kyonggi-do, Republic of Korea

TABLE OF CONTENTS

1. Abstract
2. Introduction
3. Materials and methods 3.1. Antibodies and reagents
3.2. Cell culture and sub-cellular preparation
3.3. Transient transfection
3.4. NF-kB activity assay
3.5. TGase 2 gene silencing by small interfering RNA
3.6. Western blotting
3.7. TGase activity assay
3.8. Apoptosis assay
4. Results and Discussion
4.1. TGase 2 expression and enzyme activity in two breast cancer cell lines
4.2. NF-kB activity is decreased by TGase 2 gene silencing
4.3. TGase 2 gene silencing reduces the level of nuclear NF-kB in the drug-resistant MDA231 cell line
4.4. TGase 2 gene silencing increases sensitivity to doxorubicin-induced apoptosis in MDA231 cells
4.5. TGase 2 gene silencing reduces the levels of survival factors BCl2 and BClXL
5. Discussion
6. Acknowledgement
7. Reference

1. ABSTRACT

The cross-linking enzyme, Transglutaminase 2 (TGase 2), contributes to physiological homeostasis and plays a role in cell death and survival. We previously showed that down-regulation of TGase 2 by cystamine or synthetic peptide R2 promotes apoptosis in drug-resistant cancer cells by restoring the level of I-kBa, leading to inactivation of NF-kB. To better define the action of TGase 2, its expression was blocked by small interfering RNA. This interference rendered, the doxorubicin-resistant breast cancer cells, highly susceptible to doxorubicin-induced apoptosis. This susceptibility, was associated with decreased levels of the cell-survival factors BCl2 and BCLXL whereas the level of BAX remained un-changed. Together, the findings support the view that TGase 2 leads to drug-resistance by up-regulating the level of survival factors via NF-kB activation.