Chaperone-mediated autophagy prevents apoptosis by degrading BBC3/PUMA

Wei Xie,Lei Zhang,Haifeng Jiao,Li Guan,Junmin Zha,Xiaotong Li,Mian Wu,Zhanxiang Wang,Jiahuai Han,Han You

doi:10.1080/15548627.2015.1075688

Abstract

Autophagy is a potentially inimical pathway and together with apoptosis, may be activated by similar stress stimuli that can lead to cell death. The molecular cues that dictate the cell fate choice between autophagy and apoptosis remain largely unknown. Here we report that the proapoptotic protein BBC3/PUMA (BCL2 binding component 3) is a bona fide substrate of chaperone-mediated autophagy (CMA). BBC3 associates with HSPA8/HSC70 (heat shock 70kDa protein 8), leading to its lysosome translocation and uptake. Inhibition of CMA results in stabilization of BBC3, which in turn sensitizes tumor cells to undergo apoptosis. We further demonstrate that upon TNF (tumor necrosis factor) treatment, IKBKB/IKKβ (inhibitor of kappa light polypeptide gene enhancer in B-cells, kinase β)-mediated BBC3 Ser10 phosphorylation is crucial for BBC3 stabilization via blocking its degradation by CMA. Mechanistically, Ser10 phosphorylation facilitates BBC3 translocation from the cytosol to mitochondria. BBC3 stabilization resulting from either Ser10 phosphorylation or CMA inhibition potentiates TNF-induced apoptotic cell death. Our findings thus reveal that the selective degradation of BBC3 underlies the prosurvival role of CMA and define a previously unappreciated proapoptotic role of IKBKB that acts through phosphorylation-mediated stabilization of BBC3, thereby promoting TNF-triggered apoptosis.

Highlights

Chaperone-mediated autophagy (CMA) plays an essential role in a diverse range of pathophysiological processes, including aerobic glycolysis, neurodegeneration, and killing of dormant cancer cells.[1,2,3,4,5,6,7,8] A variety of cellular stress stimuli, such as nutrient deprivation, exposure to toxic compounds, and mild oxidative stress can potentiate CMA activity.[9,10] In CMA, cytosolic substrates with a KFERQ-like motif are recognized by a chaperone-cochaperone complex that transports them to the lysosomal surface.[11]
Inhibition of CMA leads to BBC3 induction BBC3 is critical for apoptosis induced by a wide range of stress signals that act through a variety of transcriptional factors
HSPA8 depletion resulted in a significant increase of the cytosol-to-postnuclear supernatant (PNS) ratio of BBC3 in cells expressing IKBKB short hairpin RNA (shRNA) (Fig. 4E and F), compared to the ratio of BBC3 in control cells, suggesting that IKBKBmediated Ser[10] phosphorylation could stabilize BBC3 by promoting its mitochondrial localization, which in turn abrogates its degradation by CMA

Summary

Introduction

Chaperone-mediated autophagy (CMA) plays an essential role in a diverse range of pathophysiological processes, including aerobic glycolysis, neurodegeneration, and killing of dormant cancer cells.[1,2,3,4,5,6,7,8] A variety of cellular stress stimuli, such as nutrient deprivation, exposure to toxic compounds, and mild oxidative stress can potentiate CMA activity.[9,10] In CMA, cytosolic substrates with a KFERQ-like motif are recognized by a chaperone-cochaperone complex that transports them to the lysosomal surface.[11]. Genetic studies using bbc[3] knockout mice revealed a crucial role of BBC3 in the induction of apoptosis triggered by distinct apoptotic signals, including genotoxic damage, cytokine deprivation, dexamethasone, staurosporine, and PMA.[14] Distinct transcriptional programs have been reported to regulate BBC3. Genotoxic insults including g-irradiation and chemotherapeutic drugs induce BBC3 by TP53-dependent transactivation.[15,16] In addition to DNA damage signals, a variety of stress stimuli can induce BBC3 in a TP53-independent manner. SMAD4 proteins can induce BBC3 in response to growth factor deprivation, TNF or TGFB treatment, respectively.[17,18,19] In addition to transcriptional control, BBC3 has been found to undergo post-translational modification and is subject to proteasomemediated degradation,[20,21] or caspase-dependent degradation.[22]. Our results revealed that TNF-mediated BBC3 induction is fine-tuned via both post-translational and transcriptional mechanisms

Results

Discussion

Materials and Methods