SAG/ROC-SCFβ-TrCP E3 ubiquitin ligase promotes pro-caspase-3 degradation as a mechanism of apoptosis protection

M Tan, JR Gallegos, Q Gu, Y Huang, J Li, Y Jin, H Lu… - Neoplasia, 2006 - Elsevier
M Tan, JR Gallegos, Q Gu, Y Huang, J Li, Y Jin, H Lu, Y Sun
Neoplasia, 2006Elsevier
Abstract Skp1-cullin-F-box protein (SCF) is a multicomponent E3 ubiquitin (Ub) ligase that
ubiquitinates a number of important biologic molecules such as p27, β-catenin, and lκB for
proteasomal degradation, thus regulating cell proliferation and survival. One SCF
component, SAG/ROC2/Rbx2/Hrt2, a RING finger protein, was first identified as a redox-
inducible protein, which, when overexpressed, inhibited apoptosis both in vitro and in vivo.
We report here that sensitive to apoptosis gene (SAG), as well as its family member …
Abstract
Skp1-cullin-F-box protein (SCF) is a multicomponent E3 ubiquitin (Ub) ligase that ubiquitinates a number of important biologic molecules such as p27, β-catenin, and lκB for proteasomal degradation, thus regulating cell proliferation and survival. One SCF component, SAG/ROC2/Rbx2/Hrt2, a RING finger protein, was first identified as a redox-inducible protein, which, when overexpressed, inhibited apoptosis both in vitro and in vivo. We report here that sensitive to apoptosis gene (SAG), as well as its family member ROC1/Rbxi, bound to the proinactive form of caspase-3 (pro-caspase-3). Binding was likely mediated through F-box protein, β-transducin repeat-containing protein (β-TrCP), which binds to the first 38 amino acids of pro-caspase-3. Importantly, β-TrCP1 expression significantly shortened the protein half-life of pro-caspase-3, whereas expression of a dominant-negative β-TrCP1 mutant with the F-box domain deleted extended it. An in vitro ubiquitination assay showed that SAG/ROC-SCF -Trcp promoted ubiquitination of pro-caspase-3. Furthermore, endogenous levels of pro-caspase-3 were decreased by overexpression of SAG/ROC-SCFβ-TrCP E3 Ub ligases, but increased on siRNA silencing of SAG, regulator of cullin-1 (ROC1), or β-TrCPs, leading to increased apoptosis by etoposide and TNF-related apoptosis-inducing ligand through increased activation of caspase-3. Thus, pro-caspase-3 appears to be a substrate of SAG/ROC-SCFβ-TrCP E3 Ub ligase, which protects cells from apoptosis through increased apoptosis threshold by reducing the basal level of pro-caspase-3.
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