ER stress was monitored by
ER stress was monitored by both an increase of the ER chaperone (GRP78, GRP94) and the activation of the ER resident caspase-12. ER stress response (also known as the unfolded protein response) is a defense mechanism against various cellular stresses which causes accumulation of unfolded proteins in the ER. Three proximal sensors (PERK, IRE1, and ATF6) act in concert to regulate the UPR through their respective signaling cascades (Roussel et al., 2013). The protein chaperone GRP78/94 is the master regulator of these pathways. With the accumulation of unfolded proteins, GRP78/94 release enables PERK dimerization and activation to phosphorylate eIF2a, and eIF2a phosphorylation induces the translation of ATF4 mRNA. Similarly, GRP78/94 is released from IRE1 and permitted to dimerize, which activates its kinase and RNase activities to initiate Xbp1 mRNA splicing, thereby producing a potent transcriptional activator. Finally, GRP78/94 release from ATF6 (ATF6 p90) permits ATF6 transport to the Golgi compartment, where it is cleaved to yield a cytosolic fragment (ATF6 p50) that migrates to the nucleus to further activate the transcription of UPR-responsive genes (Wang and Kaufman, 2012). Our results corroborated with previous findings (Yoon et al., 2011a), which demonstrated that Mn was found to upregulate GRP78/94 expression. Furthermore, PERK phosphorylation, PERK-mediated phosphorylation of eIF2a and ATF4 mRNA po1 also increased. IRE1 activation and Xbp1 mRNA splicing also increased. However, ATF6 p90 levels did not change. Thus, the ATF6 pathway did not appear to be involved in Mn-induced ER stress. These data suggested that Mn could activate the PERK and IRE1 signal pathways in cultured slices.
Although the initial role might be neuroprotective, failure to restore proteostasis and prolonged UPR activation could play a critical role in the neurodegenerative process. CHOP is a non-ER-localized transcription factor that is induced by a variety of adverse physiological and pharmacological conditions, including ER stress. ER stress regulates CHOP transcription through two UPR pathways (PERK and ATF6) (Ohri et al., 2012). Caspase-12, the most recently characterized member of the ICE (interleukin-1β-converting enzyme) subfamily of caspases, mediates ER specific apoptosis. In addition, the activation of caspase-12 in response to ER stress has been suggested as a putative molecular marker for apoptosis (Moserova and Kralova, 2012). Our study provided evidence that the treatment with Mn resulted in a time-dependent increase in the expression of CHOP and cleaved caspase-12 in cultured slices. Moreover, growing evidence suggested that ER stress was involved in Mn-induced nerve cell apoptosis (Yeh et al., 2011, Yoon et al., 2011b).
Conflict of interest statement
Acknowledgment This work was supported by grants from the National Natural Science Foundation of China (Grant No. 81102098).