Differential ROS-Mediated Phosphorylation of Drp1 in Mitochondrial Fragmentation Induced by Distinct Cell Death Conditions in Cerebellar Granule Neurons

Figure 7

K5 and ST induce mtROS and mitochondrial fragmentation not mediated by Drp1 p-Ser616 during neuronal death. The model proposed for both apoptotic conditions comprises an early stage of ROS produced by mitochondria, which was shown in this study, as well as a late stage of ROS generated by NOX [16, 45]. Based on the finding that MitoTEMPO inhibited Drp1 phosphorylation, we propose that the mtROS are involved in the Ser616 phosphorylation of Drp1. Further, we observed mitochondrial fragmentation and finally neuronal death. However, the inhibition of Drp1 activation by MitoTEMPO did not reduce mitochondrial fragmentation; nevertheless, MitoTEMPO reduced neuronal death. This suggests that mitochondrial fragmentation mediated by Drp1 Ser616 is not involved in CGN apoptotic death. On the other hand, ST treatment (right panel) also induced an increase in both mitochondrial and cytoplasmic ROS levels. Here, we found that ST markedly reduced total Drp1 and Ser616 Drp1 phosphorylation levels and, as with K5, MitoTEMPO did not inhibit the mitochondrial fragmentation, but it prevented the neuronal death. As for K5, we observed that Drp1 phosphorylation at Ser616 is not related to either mitochondrial fragmentation or neuronal death induced by ST. Although Drp1 activation is a determinant of the mitochondrial fission in other models of death, this does not seem to occur in our model, at least through its phosphorylation at Ser616 Drp1, which does not play a role in mitochondrial fragmentation and cell death of CGN. Based on these findings, we propose that there exists an alternative mechanism that regulates the mitochondrial fragmentation in CGN.