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  current news   Press   selected story    
     
  30th January 2009  
 

Baxβ: a constitutively active human Bax isoform that is under tight regulatory control by the proteasomal degradation mechanism

 
 




Authors
Nai Yang FU, Sunil SUKUMARAN, Sze Yen KERK, Victor C. YU.

Abstract
Although mRNAs of multiple isoforms of Bax, which encodes a central regulator of apoptosis signaling, have been reported, only Baxα protein has been well documented and studied. Baxα exists in latent form and is activated upon apoptosis induction through conformational changes. Here, we demonstrate that Baxβ protein is ubiquitously present among human cells, but its activity is restricted through stringent regulation by proteasomal degradation. In contrast to Baxα, native Baxβ spontaneously integrates into mitochondrial membrane and is highly potent in inducing cytochrome c release from mitochondria. Remarkably, Baxβ protein is up-regulated by apoptotic stimuli via inhibition of its ubiquitination process and stable expression of Baxβ in HCT116-Bax-/- cell restores its sensitivity to multiple stimuli. Bax▀ associates with and promotes Baxα activation. Moreover, selective knockdown of Baxβ desensitizes HCT116-Bax+/- cells to Bax-dependent apoptosis signaling. These observations underscore the plasticity of human Bax in serving its role as a "gatekeeper" for apoptosis.

 
 


 
 


FIGURE 1:
The Baxβ isoform is tightly regulated by the ubiquitin-proteasome system
(A) Illustration depicting primary structure of human Baxα and Baxβ proteins.
(B) The C-terminus of Baxβ mediates its degradation. 293T cells transfected with the plasmid encoding the indicated myc-tagged proteins were treated with vehicle or MG132 for 8 h, followed by either direct IB of total cell lysates (upper) or IP-Western using the indicated antibodies (lower). Heavy chain of IgG (*) is indicated.
(C) Endogenous Baxβ protein is up-regulated in the cells upon proteasome inhibition. Equivalent lysates from 293T cells treated as indicated for 14 hr were IP-Western with the indicated antibodies.

 
 


 
 


FIGURE 2:
Native Baxβ is constitutively in active form and is a potent inducer of Cyto c release from mitochondria
(A) Baxβ is constitutively in an active conformation. MCF-7 cells transfected with the plasmid encoding various GFP-tagged Bax proteins were immunostained for active form of Bax (Act Bax) with the anti-Bax conformation-specific antibody N20 (red).
(B) Baxβ triggers Cyto c in transfected cells. MCF-7 transfected with indicated GFP-Bax expression plasmids were stained with anti-Cyto c (red).
(C) Baxβ potently induces Cyto c release from isolated mitochondria. Mitochondria isolated from HCT116-Bax+/- cells were incubated with in vitro translated (IVT) proteins. The mitochondria were pelleted by centrifugation. The supernatants (Sup) and mitochondrial pellets were probed with Cyto c or HSP60 antibody.

Published in Molecular Cell. 2009. Jan 15, 33: 15-29

Their work has already been cited in a review by Kim and others (Kim et al., Deadly Splicing: Bax Becomes Almighty. Mol Cell 33, Jan 30, 2009):

“Activation of Bax and Bak by BH3-only molecules triggers mitochondrial apoptosis. In a recent issue of Molecular Cell, Fu et al. (2009) identify a constitutively active isoform of Bax, Baxb, whose activity is tightly controlled by the ubiquitin-proteasome system.”

 

For more information on Victor C. YU’s Lab, Please Click here.