Abstract
Apoptosis mediated by Bax or Bak is usually thought to be triggered by BH3-only members of the Bcl-2 protein family. BH3-only proteins can directly bind to and activate Bax or Bak, or indirectly activate them by binding to anti-apoptotic Bcl-2 family members, thereby relieving their inhibition of Bax and Bak. Here we describe a third way of activation of Bax/Bak dependent apoptosis that does not require triggering by multiple BH3-only proteins. In factor dependent myeloid (FDM) cell lines, cycloheximide induced apoptosis by a Bax/Bak dependent mechanism, because Bax-/-Bak-/- lines were profoundly resistant, whereas FDM lines lacking one or more genes for BH3-only proteins remained highly sensitive. Addition of cycloheximide led to the rapid loss of Mcl-1 but did not affect the expression of other Bcl-2 family proteins. In support of these findings, similar results were observed by treating FDM cells with the CDK inhibitor, roscovitine. Roscovitine reduced Mcl-1 abundance and caused Bax/Bak dependent cell death, yet FDM lines lacking one or more genes for BH3-only proteins remained highly sensitive. Therefore Bax/Bak dependent apoptosis can be regulated by the abundance of anti-apoptotic Bcl-2 family members such as Mcl-1, independently of several known BH3-only proteins.
Highlights
The role of Bcl-2 as an inhibitor of cell death was first established in FDC-P1 cells, an IL-3 dependent mouse myeloid cell line [1]
CHX induced cell death by this time point was dependent on the expression of Bax or Bak because bax/bak deficient factor dependent myeloid (FDM) cells derived from bax/ bak double knockout (DKO) mice were profoundly resistant to CHX treatment (Fig 1A)
This resistance was confirmed by treating bax/bak DKO cells for 96 hours with CHX, at which the majority of the cells were still viable, in contrast to the rapid cell death of the WT cells (Fig 1B) From these experiments a concentration of 20 μg/ml CHX was chosen and was used in all subsequent experiments
Summary
The role of Bcl-2 as an inhibitor of cell death was first established in FDC-P1 cells, an IL-3 dependent mouse myeloid cell line [1]. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript
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