Abstract
Lysosomal membrane permeabilization is an essential step in TRAIL-induced apoptosis of liver cancer cell lines. TRAIL-induced lysosomal membrane permeabilization is mediated by the multifunctional sorting protein PACS-2 and repressed by the E3 ligases cIAP-1 and cIAP-2. Despite the opposing roles for PACS-2 and cIAPs in TRAIL-induced apoptosis, an interaction between these proteins has yet to be examined. Herein, we report that cIAP-1 and cIAP-2 confer TRAIL resistance to hepatobiliary cancer cell lines by reducing PACS-2 levels. Under basal conditions, PACS-2 underwent K48-linked poly-ubiquitination, resulting in PACS-2 proteasomal degradation. Biochemical assays showed cIAP-1 and cIAP-2 interacted with PACS-2 in vitro and co-immunoprecipitation studies demonstrated that the two cIAPs bound PACS-2 in vivo. More importantly, both cIAP-1 and cIAP-2 directly mediated PACS-2 ubiquitination in a cell-free assay. Single c-Iap-1 or c-Iap-2 gene knock-outs in mouse hepatocytes did not lead to PACS-2 accumulation. However, deletion of both cIAP-1 and cIAP-2 reduced PACS-2 ubiquitination, which increased PACS-2 levels and sensitized HuH-7 cells to TRAIL-induced lysosomal membrane permeabilization and apoptosis. Correspondingly, deletion of cIAPs sensitized wild-type, but not PACS-2-deficient hepatocarcinoma cells or Pacs-2−/− mouse hepatocytes to TRAIL-induced apoptosis. Together, these data suggest cIAPs constitutively downregulate PACS-2 by polyubiquitination and proteasomal degradation, thereby restraining TRAIL-induced killing of liver cancer cells.
Highlights
The inhibitor of apoptosis proteins (IAPs) are evolutionarily conserved and implicated in a variety of cellular processes, including repressing apoptosis in response to both extrinsic and intrinsic signaling pathways [1,2]
The determination that JNK phosphorylation of Bim Ser69 is required for PIXosome assembly and lysosomal membrane permeabilization (LMP), together with the ability of Cellular Inhibitor of Apoptosis (cIAP) to regulate death receptor-induced JNK activation and apoptosis in liver cancer cells [15] [19] [34], led us to ask if cIAP inhibitors can block Tumor Necrosis Factor-Related Apoptosis-Inducing Ligand (TRAIL)-induced LMP by preventing Bim phosphorylation
The results of this study identify a novel mechanism by which cIAPs mediate resistance to TRAIL-induced apoptosis in liver cancer cells, namely, by regulating the formation of the PACS-2, Bim, Bax-containing signaling complex PIXosome on the lysosomes
Summary
The inhibitor of apoptosis proteins (IAPs) are evolutionarily conserved and implicated in a variety of cellular processes, including repressing apoptosis in response to both extrinsic (death receptors-mediated) and intrinsic (cell stress-mediated) signaling pathways [1,2]. TRAIL stimulates JNK-mediated phosphorylation of Bim, which promotes the formation of a lysosomal signaling complex (named PIXosome) with Bax and phosphofurin acidic cluster sorting protein 2 (PACS-2) [19], a member of the PACS family of multifunctional sorting proteins regulating membrane traffic within secretory pathways and interorganellar communication [20]. This complex triggers LMP and promotes apoptosis, in accordance with a previous observation that PACS-2 is essential for TRAIL-induced hepatocyte apoptosis in a model of viral hepatitis [21]. This study identifies PACS-2 as a key substrate for cIAPs and provides new insight into the mechanisms underlying the antiapoptotic effect of IAP proteins in TRAIL cytotoxic signaling
Sign-in/Register to view highlights & summary Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
