Cellular Inhibitor of Apoptosis (cIAP)-mediated ubiquitination of Phosphofurin Acidic Cluster Sorting protein 2 (PACS-2) negatively regulates Tumor necrosis factor-Related Apoptosis-Inducing Ligand (TRAIL) cytotoxicity

Guicciardi, ME and Werneburg, NW and Bronk, SF and Franke, A and Yagita, H and Thomas, G and Gores, GJ (2014) Cellular Inhibitor of Apoptosis (cIAP)-mediated ubiquitination of Phosphofurin Acidic Cluster Sorting protein 2 (PACS-2) negatively regulates Tumor necrosis factor-Related Apoptosis-Inducing Ligand (TRAIL) cytotoxicity. PLoS ONE, 9 (3).

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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 polyubiquitination, 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. © 2014 Guicciardi et al.

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Details

Item Type:	Article
Status:	Published
Creators/Authors:	Creators Email Pitt Username ORCID Guicciardi, ME Werneburg, NW Bronk, SF Franke, A Yagita, H Thomas, G thomasg@pitt.edu THOMASG Gores, GJ
Contributors:	Contribution Contributors Name Email Pitt Username ORCID Editor Bratton, Shawn B. UNSPECIFIED UNSPECIFIED UNSPECIFIED
Date:	14 March 2014
Date Type:	Publication
Journal or Publication Title:	PLoS ONE
Volume:	9
Number:	3
DOI or Unique Handle:	10.1371/journal.pone.0092124
Schools and Programs:	School of Medicine > Microbiology and Molecular Genetics
Refereed:	Yes
Date Deposited:	23 Jun 2014 21:33
Last Modified:	23 Jun 2021 07:55
URI:	http://d-scholarship.pitt.edu/id/eprint/21941

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