Mechanisms of TLR4-Mediated Autophagy and Nitroxidative Stress

Pathogenic infections have badly affected public health and the development of the breeding industry. Billions of dollars are spent every year fighting against these pathogens. The immune cells of a host produce reactive oxygen species and reactive nitrogen species which promote the clearance of these microbes. In addition, autophagy, which is considered an effective method to promote the destruction of pathogens, is involved in pathological processes. As research continues, the interplay between autophagy and nitroxidative stress has become apparent. Autophagy is always intertwined with nitroxidative stress. Autophagy regulates nitroxidative stress to maintain homeostasis within an appropriate range. Intracellular oxidation, in turn, is a strong inducer of autophagy. Toll-like receptor 4 (TLR4) is a pattern recognition receptor mainly involved in the regulation of inflammation during infectious diseases. Several studies have suggested that TLR4 is also a key regulator of autophagy and nitroxidative stress. In this review, we describe the role of TLR4 in autophagy and oxidation, and focus on its function in influencing autophagy-nitroxidative stress interactions.

Dynamic posttranslational modifications of cytoskeletal proteins unveil hot spots under nitroxidative stress

The cytoskeleton is a supramolecular structure consisting of interacting protein networks that support cell dynamics in essential processes such as migration and division, as well as in responses to stress. Fast cytoskeletal remodeling is achieved with the participation of regulatory proteins and posttranslational modifications (PTMs). Redox-related PTMs are emerging as critical players in cytoskeletal regulation. Here we used a cellular model of mild nitroxidative stress in which a peroxynitrite donor induced transient changes in the organization of three key cytoskeletal proteins, i.e., vimentin, actin and tubulin. Nitroxidative stress-induced reconfiguration of intermediate filaments, microtubules and actin structures were further correlated with their PTM profiles and dynamics of the PTM landscape. Using high-resolution mass spectrometry, 62 different PTMs were identified and relatively quantified in vimentin, actin and tubulin proteins, including 12 enzymatic, 13 oxidative and 2 nitric oxide-derived modifications as well as 35 modifications by carbonylated lipid peroxidation products, thus evidencing the occurrence of a chain reaction with formation of reactive species and the activation of multiple signaling pathways. Our results unveil the presence of certain modifications under basal conditions and their modulation in response to stress in a target-, residue- and reactive species-dependent manner. Moreover, we identified protein PTM “hot spots”, such as the single cysteine residue of vimentin, supporting its role in PTM cross-talk and redox sensing. Finally, identification of novel PTMs in these proteins may pave the way for unveiling novel cytoskeleton regulatory mechanisms.