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Functional interaction between retinoblastoma protein and stress-activated protein kinase in multiple myeloma cells
D Chauhan1, T Hideshima, S Treon
1Department of Adult Oncology, Dana Farber Cancer Institute, Harvard Medical School, Boston, Massachusetts 02115, USA.
Abstract:
Previous studies have demonstrated that gamma-irradiation (IR)-induced apoptosis in multiple myeloma (MM) is associated with activation of stress-activated protein kinase (SAPK). In the present study, we examined the molecules downstream of SAPK/C-Jun N-terminal kinase (JNK), focusing on the role of retinoblastoma protein (Rb) during IR-induced MM cell apoptosis. The results demonstrate that IR activates SAPK/JNK, which associates with Rb both in vivo and in vitro. Far Western blot analysis confirms that SAPK/JNK binds directly to Rb. IR-activated SAPK/JNK phosphorylates Rb, and deletion of the phosphorylation site in the COOH terminus domain of Rb abrogates phosphorylation of Rb by SAPK/JNK. Taken together, our results suggest that Rb is a target protein of SAPK/JNK and that the association of SAPK/JNK and Rb mediates IR-induced apoptosis in MM cells.
Insights
Gamma irradiation triggers apoptosis in multiple myeloma cells by activating stress-activated protein kinase/c-Jun N-terminal kinase (SAPK/JNK). This kinase directly binds and phosphorylates retinoblastoma protein (Rb), mediating cell death.
Area of Science:
- Oncology
- Molecular Biology
- Cellular Biology
Background:
- Gamma irradiation (IR) induces apoptosis in multiple myeloma (MM).
- Stress-activated protein kinase/c-Jun N-terminal kinase (SAPK/JNK) pathway activation is linked to IR-induced apoptosis in MM.
- The precise downstream targets of SAPK/JNK in this process remain under investigation.
Purpose of the Study:
- To investigate the role of retinoblastoma protein (Rb) as a downstream target of SAPK/JNK in IR-induced MM apoptosis.
- To elucidate the molecular interactions between SAPK/JNK and Rb during gamma irradiation.
Main Methods:
- In vivo and in vitro association studies to confirm SAPK/JNK and Rb interaction.
- Far Western blot analysis to verify direct binding of SAPK/JNK to Rb.
- Site-directed mutagenesis to examine the role of Rb phosphorylation in apoptosis.
Main Results:
- Gamma irradiation activates SAPK/JNK in MM cells.
- Activated SAPK/JNK directly binds to Rb both in vivo and in vitro.
- SAPK/JNK phosphorylates Rb at its COOH terminus domain.
- Abrogation of the Rb phosphorylation site prevents SAPK/JNK-mediated phosphorylation and IR-induced apoptosis.
Conclusions:
- Retinoblastoma protein (Rb) is a direct target of SAPK/JNK in multiple myeloma cells.
- The interaction and phosphorylation of Rb by SAPK/JNK are critical mediators of gamma irradiation-induced apoptosis in MM.