ABSTRACT
Thioredoxin 1 (Trx1) and glutaredoxin 1 (Grx1) are two ubiquitous redox enzymes that are central for redox homeostasis but also are implicated in many other processes, including stress sensing, inflammation, and apoptosis. In addition to their enzymatic redox activity, a growing body of evidence shows that Trx1 and Grx1 play regulatory roles via protein-protein interactions with specific proteins, including Ask1. The currently available inhibitors of Trx1 and Grx1 are thiol-reactive electrophiles or disulfides that may suffer from low selectivity because of their thiol reactivity. In this report, we used a phage peptide library to identify a 7-mer peptide, 2GTP1, that binds to both Trx1 and Grx1. We further showed that a cell-permeable derivative of 2GTP1, TAT-2GTP1, disrupts the Trx1-Ask1 interaction, which induces Ask1 phosphorylation with subsequent activation of JNK, stabilization of p53, and reduced viability of cancer cells. Notably, as opposed to a disulfide-derived Trx1 inhibitor (PX-12), TAT-2GTP1 was selective for activating the Ask1 pathway without affecting other stress signaling pathways, such as endoplasmic reticulum stress and AMPK activation. Overall, 2GTP1 will serve as a useful probe for investigating protein interactions of Trx1.
Subject(s)
MAP Kinase Kinase Kinase 5/antagonists & inhibitors , MAP Kinase Signaling System/drug effects , Oligopeptides/pharmacology , Peptide Library , Stress, Physiological/physiology , Antineoplastic Agents/chemistry , Antineoplastic Agents/pharmacology , Cell Line, Tumor , Cell Membrane Permeability , Cell Survival/drug effects , Drug Screening Assays, Antitumor , Endoplasmic Reticulum Stress/drug effects , Enzyme Activation/drug effects , Enzymes, Immobilized , Glutaredoxins , HEK293 Cells , Humans , MAP Kinase Kinase Kinase 5/chemistry , MAP Kinase Kinase Kinase 5/physiology , NADP/analysis , Oligopeptides/isolation & purification , Oxidation-Reduction , Phosphorylation/drug effects , Protein Binding/drug effects , Protein Interaction Mapping , Protein Processing, Post-Translational/drug effects , Reactive Oxygen Species/metabolism , Recombinant Proteins/metabolismABSTRACT
Small molecules that block the altered metabolism in cancer or increase the production of reactive oxygen species (ROS) are emerging as potential anti-cancer agents. Considering that various carbohydrates can be used for cellular energetics or protein N-glycosylation of which interruption can lead to cellular stress, we have synthesized and evaluated a library of N-aryl glycosides for induction of ROS and cytotoxicity in H1299 cancer cell line. Two N-aryl glycosides (K8 and H8) were identified that induce about 2-fold induction of ROS and cytotoxicity in H1299 cells. We further showed that the acetylated form of K8 (K8A) activates AMPK, and stabilizes p53 in HEK293 cells, and induce a higher cytotoxicity than 2-deoxy-d-glucose in H1299 cell line.
