Differential correlations between changes to glutathione redox state, protein ubiquitination, and stress-inducible HSPA chaperone expression after different types of oxidative stress

Abstract

In primary bovine fibroblasts with an hspa1b/luciferase transgene, we examined the intensity of heat-shock response (HSR) following four types of oxidative stress or heat stress (HS), and its putative relationship with changes to different cell parameters, including reactive oxygen species (ROS), the redox status of the key molecules glutathione (GSH), NADP(H) NAD(H), and the post-translational protein modifications carbonylation, S-glutathionylation, and ubiquitination. We determined the sub-lethal condition generating the maximal luciferase activity and inducible HSPA protein level for treatments with hydrogen peroxide (HO), UVA-induced oxygen photo-activation, the superoxide-generating agent menadione (MN), and diamide (DA), an electrophilic and sulfhydryl reagent. The level of HSR induced by oxidative stress was the highest after DA and MN, followed by UVA and HO treatments, and was not correlated to the level of ROS production nor to the extent of protein S-glutathionylation or carbonylation observed immediately after stress. We found a correlation following oxidative treatments between HSR and the level of GSH/GSSG immediately after stress, and the increase in protein ubiquitination during the recovery period. Conversely, HS treatment, which led to the highest HSR level, did not generate ROS nor modified or depended on GSH redox state. Furthermore, the level of protein ubiquitination was maximum immediately after HS and lower than after MN and DA treatments thereafter. In these cells, heat-induced HSR was therefore clearly different from oxidative stress-induced HSR, in which conversely early redox changes of the major cellular thiol predicted the level of HSR and polyubiquinated proteins.

Hsp70 in Redox Homeostasis

“…Oxidative stress can activate the HSR and Kelch-like ECH-associated protein 1 (Keap1)/Nrf2 pathway to upregulate expression of HSP70 , and the increased protein level of Hsp70 protects cells against oxidative-stress-related damage [ 15 , 98 , 99 ]. However, different types of oxidative stress differ in their capacity to induce the HSR and elevate expression of HSP70 , and the same oxidative stimulus may lead to distinct stress responses in different cells [ 100 , 101 , 102 ]. Oxidative-stress induced by 15-deoxy-Δ12,14-prostaglandin J2 (15d-PGJ2) enhances HSP70 expression to exert anti-inflammatory effects [ 103 ].…”

Imbalanced GSH/ROS and sequential cell death

“…The redox state of cellular GSH is a key regulatory element of the protein ubiquitination system. [59] The protein ubiquitination system is an important nonlysosomal corridor through which proteins involved in cell proliferation, differentiation, and apoptosis in eukaryotic cells are degraded. [35] The imbalance of ROS and GSH would affect the protein ubiquitination system in T A B L E 2 Factors that consume GSH MeHg (44,45,46) MeHg can be chelated with GSH to form the Hg-GSH complex.…”

Early Weaning Affects Liver Antioxidant Function in Piglets

“…In this study, weaning down-regulated gene expression of GSH in the liver of piglets on the 3rd day after weaning. However, the enzyme activity of GSH did not change significantly after weaning, the reason for which might be that the modification and activation of antioxidant proteins are affected when piglets are under weaning stress [41,42].…”