Molecular Characterization, Protein-Protein Interaction Network, and Evolution of Four Glutathione Peroxidases from Tetrahymena thermophila.

Diana Ferro,Rigers Bakiu,Cristina Miceli,Sandra Pucciarelli,Adriana Vallesi,Gianfranco Santovito,Paola Irato

doi:10.3390/antiox9100949

Abstract

Glutathione peroxidases (GPxs) form a broad family of antioxidant proteins essential for maintaining redox homeostasis in eukaryotic cells. In this study, we used an integrative approach that combines bioinformatics, molecular biology, and biochemistry to investigate the role of GPxs in reactive oxygen species detoxification in the unicellular eukaryotic model organism Tetrahymena thermophila. Both phylogenetic and mechanistic empirical model analyses provided indications about the evolutionary relationships among the GPXs of Tetrahymena and the orthologous enzymes of phylogenetically related species. In-silico gene characterization and text mining were used to predict the functional relationships between GPxs and other physiologically-relevant processes. The GPx genes contain conserved transcriptional regulatory elements in the promoter region, which suggest that transcription is under tight control of specialized signaling pathways. The bioinformatic findings were next experimentally validated by studying the time course of gene transcription and enzymatic activity after copper (Cu) exposure. Results emphasize the role of GPxs in the detoxification pathways that, by complex regulation of GPx gene expression, enable Tethraymena to survive in high Cu concentrations and the associated redox environment.

Highlights

The maintenance of an equilibrium between production and elimination of reactive oxygen species (ROS) is a fundamental molecular process that has been playing a pivotal role in eukaryotic survival and aging since the appearance of the aerobic metabolism [1]
The results presented in this work confirm that the antioxidant system of T. thermophila, a single cell eukaryotic model species in genetic and molecular biology, presents a high degree of complexity comparable to that found in metazoan species
New information has been uncovered regarding the molecular evolution of Glutathione peroxidases (GPxs) of this species, the evolutionary events that led to the appearance of twelve functional isoforms in T. thermophila will have to be further investigated

Summary

Introduction

The maintenance of an equilibrium between production and elimination of reactive oxygen species (ROS) is a fundamental molecular process that has been playing a pivotal role in eukaryotic survival and aging since the appearance of the aerobic metabolism [1]. ROS are continuously generated in living organisms as a normal consequence of electrons moving across the inner mitochondrial membrane and free oxygen that is present inside the cell. In order to modulate ROS presence, antioxidant molecules are present in cells as a defense against the risk of Antioxidants 2020, 9, 949; doi:10.3390/antiox9100949 www.mdpi.com/journal/antioxidants. Antioxidants 2020, 9, 949 oxidative stress [6]. Antioxidant enzymes and metabolites should continuously and efficiently work together since small variations in their physiological concentrations can have dramatic effects on the resistance of cellular lipids, proteins, and DNA to oxidative damage. The antioxidant system is characterized by strong protein–protein interactions and associated signaling pathways intensely regulated by the intracellular oxidative state [1]

Methods

Discussion

Conclusion