ABSTRACT
Interleukin 24 (IL-24) is a tumor-suppressing protein, which inhibits angiogenesis and induces cancer cell-specific apoptosis. We have shown that IL-24 regulates apoptosis through phosphorylated eukaryotic initiation factor 2 alpha (eIF2α) during endoplasmic reticulum (ER) stress in cancer. Although multiple stresses converge on eIF2α phosphorylation, the cellular outcome is not always the same. In particular, ER stress-induced apoptosis is primarily regulated through the extent of eIF2α phosphorylation and activating transcription factor 4 (ATF4) action. Our studies show for the first time that cyclic adenosine monophosphate (cAMP)-dependent protein kinase A (PKA) activation is required for IL-24-induced cell death in a variety of breast cancer cell lines and this event increases ATF4 activity. We demonstrate an undocumented role for PKA in regulating IL-24-induced cell death, whereby PKA stimulates phosphorylation of p38 mitogen-activated protein kinase and upregulates extrinsic apoptotic factors of the Fas/FasL signaling pathway and death receptor 4 expression. We also demonstrate that phosphorylation and nuclear import of tumor suppressor TP53 occurs downstream of IL-24-mediated PKA activation. These discoveries provide the first mechanistic insights into the function of PKA as a key regulator of the extrinsic pathway, ER stress, and TP53 activation triggered by IL-24.
Subject(s)
Apoptosis , Breast Neoplasms/metabolism , Breast Neoplasms/pathology , Cyclic AMP-Dependent Protein Kinases/metabolism , Interleukins/metabolism , Activating Transcription Factor 4/metabolism , Cell Line, Tumor , Cell Nucleus/drug effects , Cell Nucleus/metabolism , Enzyme Activation/drug effects , Female , Humans , Models, Biological , Phosphorylation/drug effects , Protein Kinase Inhibitors/pharmacology , Protein Transport/drug effects , Tumor Suppressor Protein p53/metabolismABSTRACT
Interleukin-24 (IL-24), a member of the IL-10 cytokine family, is an immunomodulatory cytokine that also displays broad cancer-specific suppressor effects. The tumor suppressor activities of IL-24 include inhibition of angiogenesis, sensitization to chemotherapy, and cancer-specific apoptosis. We show that Sigma 1 Receptor (S1R), a ligand-regulated protein chaperone contributes to IL-24 induction of apoptosis. IL-24 generated from an adenovirus expressing IL-24 (Ad.IL-24) induces cancer-specific apoptosis by inducing an endoplasmic reticulum (ER) stress, reactive oxygen species production, and calcium mobilization. The present studies reveals that S1R is required for Ad.IL-24-induced cell death. We provide several lines of evidence to confirm a physical and functional interaction between IL-24 and S1R including: (a) S1R and IL-24 co-localize, as judged by immunocytochemical analysis studies; (b) S1R and IL-24 co-immunoprecipitate using either S1R or IL-24 antibody; (c) S1R agonist (+)-SKF10047 inhibits apoptosis by Ad.IL-24; (d) (+)-SKF10047-mediated inhibition of Ad.IL-24 results in: diminished ER stress protein expression; (e) Calcium mobilization; and (f) ROS production. Collectively, these data demonstrate that S1R interacts with IL-24 and suggest that IL-24:S1R interaction determines apoptosis induction by Ad.IL-24. These studies define Sigma 1 Receptor as a key initial mediator of IL-24 induction of cancer-specific killing. These findings have important implications for our understanding of IL-24 as a tumor suppressor protein as well as an immune modulating cytokine.
Subject(s)
Apoptosis , Interleukins/immunology , Neoplasms/immunology , Receptors, sigma/immunology , Calcium Signaling , Caspase 3/immunology , Cell Line, Tumor/immunology , Endoplasmic Reticulum Stress , Humans , Neoplasms/pathology , Reactive Oxygen Species/immunology , Receptors, sigma/agonists , Sigma-1 ReceptorABSTRACT
The inability of the adult mammalian retina to regenerate can be partly attributed to the expression of injury-induced inhibitory extracellular matrix (ECM) and cell adhesion molecules. In particular, photoreceptor degeneration stimulates deposition of the inhibitory ECM proteins neurocan and CD44 at the outer limits of the dystrophic retina, where they act as a barrier against cellular migration and axonal extension. We have previously shown that degradation of these molecules, via induction of MMP2, promotes host-donor integration and retinal repopulation following transplantation. Here we present a biodegradable/biocompatible polymer scaffold that has the ability to deliver MMP2, in conjunction with retinal progenitor cells, directly to the site of retinal injury in an attempt to enhance cellular integration and promote retinal repopulation. Pre-activated MMP2, loaded into a PLGA polymer, maintained its activity throughout polymer fabrication and hydrolysis. Following delivery, significant degradation of CD44 and neurocan from the outer limits of the dystrophic retina, without further disruption of retinal architecture, was observed. As a result, the number of retinal progenitor cells that migrated beyond the glial barrier into the degenerating host increased significantly. These cells took up residence in the retinal outer nuclear layer, adopted appropriate photoreceptor morphology and expressed the mature photoreceptor markers recoverin and rhodopsin. Thus, we have created a cell delivery platform that upon transplantation provides controlled release of active-MMP2 directly to the site of retinal injury, stimulating inhibitory ECM barrier removal and enhancement of stem cell integration and retinal repopulation.
