Mapping the phenotypic repertoire of the cytoplasmic 2-Cys peroxiredoxin – Thioredoxin system. 1. Understanding commonalities and differences among cell types

Abstract

The system (PTTRS) formed by typical 2-Cys peroxiredoxins (Prx), thioredoxin (Trx), Trx reductase (TrxR), and sulfiredoxin (Srx) is central in antioxidant protection and redox signaling in the cytoplasm of eukaryotic cells. Understanding how the PTTRS integrates these functions requires tracing phenotypes to molecular properties, which is non-trivial. Here we analyze this problem based on a model that captures the PTTRS’ conserved features. We have mapped the conditions that generate each distinct response to H2O2 supply rates (vsup), and estimated the parameters for thirteen human cell types and for Saccharomyces cerevisiae. The resulting composition-to-phenotype map yielded the following experimentally testable predictions. The PTTRS permits many distinct responses including ultra-sensitivity and hysteresis. However, nearly all tumor cell lines showed a similar response characterized by limited Trx-S- depletion and a substantial but self-limited gradual accumulation of hyperoxidized Prx at high vsup. This similarity ensues from strong correlations between the TrxR, Srx and Prx activities over cell lines, which contribute to maintain the Prx-SS reduction capacity in slight excess over the maximal steady state Prx-SS production. In turn, in erythrocytes, hepatocytes and HepG2 cells high vsup depletes Trx-S- and oxidizes Prx mainly to Prx-SS. In all nucleated human cells the Prx-SS reduction capacity defined a threshold separating two different regimes. At sub-threshold vsup the cytoplasmic H2O2 concentration is determined by Prx, nM-range and spatially localized, whereas at supra-threshold vsup it is determined by much less active alternative sinks and μM-range throughout the cytoplasm. The yeast shows a distinct response where the Prx Tsa1 accumulates in sulfenate form at high vsup. This is mainly due to an exceptional stability of Tsa1's sulfenate. The implications of these findings for thiol redox regulation and cell physiology are discussed. All estimates were thoroughly documented and provided, together with analytical approximations for system properties, as a resource for quantitative redox biology.

Repurposing drugs as pro‐oxidant redox modifiers to eliminate cancer stem cells and improve the treatment of advanced stage cancers

“…The intracellular levels of Trxs (approximately 17‐69 μM) can be two orders of magnitude lower than those of GSH (2‐4 mM) in some cancer and noncancer cell types. However, the cytosolic TrxR activity is similar or higher than those of GR and GPx, such that for the purpose of managing intracellular ROS levels, the TrxR/Trx system is essential, maintaining important antioxidant functions and regulating many key components. For instance, Trxs act on several key protein substrates providing reducing equivalents for these, including ribonucleotide reductase, Prxs(Trx peroxidases), glucocorticoid receptors, and transcription factors such as AP‐1, NF‐κB, and p53 (Figure ).…”

“…Trxs are small redox‐active proteins whose essential function inside normal cells is to facilitate DNA metabolism and repair, transcription, and cell‐cell communication . The intracellular levels of Trxs (approximately 17‐69 μM) can be two orders of magnitude lower than those of GSH (2‐4 mM) in some cancer and noncancer cell types. However, the cytosolic TrxR activity is similar or higher than those of GR and GPx, such that for the purpose of managing intracellular ROS levels, the TrxR/Trx system is essential, maintaining important antioxidant functions and regulating many key components.…”

“…However, the Prxs are of great importance for regulation of signaling cascades that are modulated by H 2 O 2 , lipid hydroperoxides, or peroxynitrite. Modeling of the activities of the Prxs‐Trxs‐TrxR1 (and Srx) system has suggested their synergetic coupling in cells and strongly correlated with their protein concentrations and activities in 10 different tumor cell lines . The net result is to maintain redox homeostasis with the Prx‐SS reduction capacity in slight excess over the maximal steady state level of Prx‐SS production.…”

“…Modeling analysis further indicated that at low oxidative loads (in the nanomolar range) the 2‐Cys Prxs consume approximately 99% of the H 2 O 2 in the cytosol with TrxR/Trx also adequately sufficient to maintain Prx function. In turn, GPx1 is much less active than TrxR, has a lower affinity for H 2 O 2 ( K m of 20 μM) and thus GPx is unable to sense nanomolar H 2 O 2 concentrations. Consequently, it has been predicted that GPx typically consumes less than 0.2% of the H 2 O 2 under basal, nonoxidative stress conditions .…”

“…In turn, GPx1 is much less active than TrxR, has a lower affinity for H 2 O 2 ( K m of 20 μM) and thus GPx is unable to sense nanomolar H 2 O 2 concentrations. Consequently, it has been predicted that GPx typically consumes less than 0.2% of the H 2 O 2 under basal, nonoxidative stress conditions . Despite their elevated protein content, GPx‐1 as well as GPx‐4 are markedly deacetylated in cancer cells, becoming inactive .…”