PERK Integrates Autophagy and Oxidative Stress Responses To Promote Survival during Extracellular Matrix Detachment

ABSTRACT Mammary epithelial cells (MECs) detached from the extracellular matrix (ECM) produce deleterious reactive oxygen species (ROS) and induce autophagy to survive. The coordination of such opposing responses likely dictates whether epithelial cells survive ECM detachment or undergo anoikis. Here, we demonstrate that the endoplasmic reticulum kinase PERK facilitates survival of ECM-detached cells by concomitantly promoting autophagy, ATP production, and an antioxidant response. Loss-of-function studies show that ECM detachment activates a canonical PERK-eukaryotic translation initiation factor 2α (eIF2α)-ATF4-CHOP pathway that coordinately induces the autophagy regulators ATG6 and ATG8, sustains ATP levels, and reduces ROS levels to delay anoikis. Inducible activation of an Fv2E-ΔNPERK chimera by persistent activation of autophagy and reduction of ROS results in lumen-filled mammary epithelial acini. Finally, luminal P-PERK and LC3 levels are reduced in PERK-deficient mammary glands, whereas they are increased in human breast ductal carcinoma in situ (DCIS) versus normal breast tissues. We propose that the normal proautophagic and antioxidant PERK functions may be hijacked to promote the survival of ECM-detached tumor cells in DCIS lesions.

[1]  A. Harris,et al.  Regulation of autophagy by ATF4 in response to severe hypoxia , 2010, Oncogene.

[2]  J. Debnath,et al.  Autophagy and Tumorigenesis , 2010, Seminars in Immunopathology.

[3]  E. Petricoin,et al.  Malignant Precursor Cells Pre-Exist in Human Breast DCIS and Require Autophagy for Survival , 2010, PloS one.

[4]  Jiangbin Ye,et al.  PERK promotes cancer cell proliferation and tumor growth by limiting oxidative DNA damage , 2010, Oncogene.

[5]  B. Hirst,et al.  Glycine transporter GLYT1 is essential for glycine‐mediated protection of human intestinal epithelial cells against oxidative damage , 2010, The Journal of physiology.

[6]  L. Moretti,et al.  Endoplasmic Reticulum Stress Mediates Radiation-Induced Autophagy via PERK-eIF2α in Caspase-3/7 Deficient Cells , 2010, Oncogene.

[7]  Philippe Lambin,et al.  The unfolded protein response protects human tumor cells during hypoxia through regulation of the autophagy genes MAP1LC3B and ATG5. , 2010, The Journal of clinical investigation.

[8]  Hanna Y. Irie,et al.  Antioxidant and oncogene rescue of metabolic defects caused by loss of matrix attachment , 2009, Nature.

[9]  A. Harris,et al.  The role of ATF4 stabilization and autophagy in resistance of breast cancer cells treated with Bortezomib. , 2009, Cancer research.

[10]  P. Maher,et al.  Basal Levels of eIF2α Phosphorylation Determine Cellular Antioxidant Status by Regulating ATF4 and xCT Expression* , 2009, Journal of Biological Chemistry.

[11]  C. Thompson,et al.  PERK-dependent regulation of lipogenesis during mouse mammary gland development and adipocyte differentiation , 2008, Proceedings of the National Academy of Sciences.

[12]  J. Debnath,et al.  Extracellular matrix regulation of autophagy. , 2008, Current opinion in cell biology.

[13]  P. Fisher,et al.  Regulation of autophagy by ceramide-CD95-PERK signaling , 2008, Autophagy.

[14]  Jayanta Debnath,et al.  Modeling morphogenesis and oncogenesis in three-dimensional breast epithelial cultures. , 2008, Annual review of pathology.

[15]  J. Debnath,et al.  Detachment-induced autophagy during anoikis and lumen formation in epithelial acini , 2008, Autophagy.

[16]  T. Hunter,et al.  Real-time imaging reveals that noninvasive mammary epithelial acini can contain motile cells , 2007, The Journal of cell biology.

[17]  J. Debnath,et al.  Induction of autophagy during extracellular matrix detachment promotes cell survival. , 2007, Molecular biology of the cell.

[18]  Masaaki Komatsu,et al.  Homeostatic Levels of p62 Control Cytoplasmic Inclusion Body Formation in Autophagy-Deficient Mice , 2007, Cell.

[19]  Sharon J. Sequeira,et al.  Inhibition of Proliferation by PERK Regulates Mammary Acinar Morphogenesis and Tumor Formation , 2007, PloS one.

[20]  Y Kouroku,et al.  ER stress (PERK/eIF2α phosphorylation) mediates the polyglutamine-induced LC3 conversion, an essential step for autophagy formation , 2007, Cell Death and Differentiation.

[21]  Per O. Seglen,et al.  Methods for Monitoring Autophagy from Yeast to Human , 2007, Autophagy.

[22]  Gordon B. Mills,et al.  The energy sensing LKB1–AMPK pathway regulates p27kip1 phosphorylation mediating the decision to enter autophagy or apoptosis , 2007, Nature Cell Biology.

[23]  F. Urano,et al.  Autophagy Is Activated for Cell Survival after Endoplasmic ReticulumStress , 2006, Molecular and Cellular Biology.

[24]  D. Ron,et al.  Perk-Dependent Translational Regulation Promotes Tumor Cell Adaptation and Angiogenesis in Response to Hypoxic Stress , 2006, Molecular and Cellular Biology.

[25]  S. Gruber,et al.  Anti-oncogenic role of the endoplasmic reticulum differentially activated by mutations in the MAPK pathway , 2006, Nature Cell Biology.

[26]  N. Bulleid,et al.  The role of glutathione in disulphide bond formation and endoplasmic‐reticulum‐generated oxidative stress , 2006, EMBO reports.

[27]  J. Aguirre-Ghiso,et al.  Functional coupling of p38-induced up-regulation of BiP and activation of RNA-dependent protein kinase-like endoplasmic reticulum kinase to drug resistance of dormant carcinoma cells. , 2006, Cancer research.

[28]  D. Scheuner,et al.  ER stress‐regulated translation increases tolerance to extreme hypoxia and promotes tumor growth , 2005, The EMBO journal.

[29]  Jayanta Debnath,et al.  Modelling glandular epithelial cancers in three-dimensional cultures , 2005, Nature Reviews Cancer.

[30]  Ying Huang,et al.  Cystine-glutamate transporter SLC7A11 in cancer chemosensitivity and chemoresistance. , 2005, Cancer research.

[31]  D. Hardie.  The AMP-activated protein kinase pathway – new players upstream and downstream , 2004, Journal of Cell Science.

[32]  D. Scheuner,et al.  Cytoprotection by pre‐emptive conditional phosphorylation of translation initiation factor 2 , 2004, The EMBO journal.

[33]  Randal J. Kaufman,et al.  Nrf2 Is a Direct PERK Substrate and Effector of PERK-Dependent Cell Survival , 2003, Molecular and Cellular Biology.

[34]  Jayanta Debnath,et al.  Morphogenesis and oncogenesis of MCF-10A mammary epithelial acini grown in three-dimensional basement membrane cultures. , 2003, Methods.

[35]  Jayanta Debnath,et al.  The Role of Apoptosis in Creating and Maintaining Luminal Space within Normal and Oncogene-Expressing Mammary Acini , 2002, Cell.

[36]  D. Ron,et al.  Perk is essential for translational regulation and cell survival during the unfolded protein response. , 2000, Molecular cell.

[37]  R. Mulcahy,et al.  Regulation of γ-glutamylcysteine synthetase subunit gene expression: Insights into transcriptional control of antioxidant defenses , 2000, Free radical research.

[38]  Wild Ac,et al.  Regulation of gamma-glutamylcysteine synthetase subunit gene expression by the transcription factor Nrf2. , 1999 .

[39]  D. Ron,et al.  Protein translation and folding are coupled by an endoplasmic-reticulum-resident kinase , 1999, Nature.

[40]  L. Philipson,et al.  CHOP (GADD153) and its oncogenic variant, TLS-CHOP, have opposing effects on the induction of G1/S arrest. , 1994, Genes & development.

[41]  P. Dennis,et al.  The GST-BHMT assay and related assays for autophagy. , 2009, Methods in enzymology.

[42]  R. Kaufman,et al.  A trip to the ER: coping with stress. , 2004, Trends in cell biology.

[43]  Arve-Olay Solumsmo,et al.  Methods of monitoring , 2002 .