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BAD enables ceramide to signal apoptosis via Ras and Raf-1
1Laboratory of Signal Transduction, Sloan-Kettering Institute, Memorial Sloan-Kettering Cancer Center, New York, New York 10021, USA.
Abstract:
Prior investigations document that proliferative signaling cascades, under some circumstances, initiate apoptosis, although mechanisms that dictate the final outcome are largely unknown. In COS-7 cells, ceramide signals Raf-1 activation through Ras (Zhang, Y., Yao, B., Delikat, S., Bayoumy, S., Lin, X. H., Basu, S., McGinley, M., Chan-Hui, P. Y., Lichenstein, H., and Kolesnick, R. (1997) Cell 89, 63-72), but not apoptosis. However, expression of small amounts of the pro-apoptotic Bcl-2 family member, BAD, conferred ceramide-induced apoptosis onto COS-7 cells. Ceramide signaled apoptosis in BAD-expressing cells by a pathway involving sequentially kinase suppressor of Ras (KSR)/ceramide-activated protein kinase, Ras, c-Raf-1, and MEK1. Downstream, this pathway linked to BAD dephosphorylation at serine 136 by prolonged inactivation of Akt/PKB. Further, mutation of BAD at serine 136 abrogated ceramide signaling of apoptosis. The present study indicates that when ceramide signals through the Ras/Raf cascade, the availability of a single target, BAD, may dictate an apoptotic outcome.
Insights
Ceramide can trigger cell death (apoptosis) by activating the Ras/Raf pathway, but only if the pro-apoptotic protein BAD is present. BAD
Area of Science:
- Cellular signaling pathways
- Apoptosis regulation
- Molecular mechanisms of cell death
Background:
- Proliferative signaling can induce apoptosis, but the underlying mechanisms remain unclear.
- Ceramide activates Raf-1 via Ras in COS-7 cells, but this does not typically lead to apoptosis.
- The pro-apoptotic Bcl-2 family member BAD is crucial for mediating apoptosis in certain contexts.
Purpose of the Study:
- To investigate the role of BAD in ceramide-induced apoptosis.
- To elucidate the signaling pathway linking ceramide to apoptosis in BAD-expressing cells.
- To determine the critical molecular events downstream of ceramide signaling that dictate cell fate.
Main Methods:
- Utilized COS-7 cells engineered to express varying levels of the pro-apoptotic protein BAD.
- Investigated the signaling cascade initiated by ceramide, including Ras, Raf-1, MEK1, KSR, and Akt/PKB.
- Employed site-directed mutagenesis to assess the role of BAD phosphorylation at serine 136.
Main Results:
- Expression of BAD conferred ceramide-induced apoptosis in COS-7 cells.
- Ceramide signaled apoptosis via a pathway involving kinase suppressor of Ras (KSR), Ras, c-Raf-1, and MEK1.
- This pathway led to BAD dephosphorylation at serine 136 via Akt/PKB inactivation, and mutation of this site abrogated apoptosis.
Conclusions:
- The availability of BAD as a target is critical for ceramide to induce apoptosis through the Ras/Raf cascade.
- BAD dephosphorylation at serine 136 is a key event in ceramide-mediated apoptosis.
- This study highlights how a single protein's availability can determine the outcome of proliferative signaling.