ALS-causing SOD1 mutants regulate occludin phosphorylation/ ubiquitination and endocytic trafficking via the ITCH/Eps15/Rab5 axis

It is increasingly recognized that blood-spinal cord barrier (BSCB) breakdown is a hallmark of amyotrophic lateral sclerosis (ALS). BSCB integrity is disrupted prior to disease onset. Occludin, as the functional component of the endothelial barrier, is downregulated in mouse models expressing ALS-linked superoxide dismutase-1 (SOD1) mutants. However, the molecular mechanisms underlying the regulation of occludin expression remain elusive. Here, using SOD1G93A transgenic mice and endothelial cells expressing SOD1 mutants of different biochemical characteristics, we found that the SOD1 mutation disrupted endothelial barrier integrity and that the occludin expression level was downregulated with disease progression. Our mechanistic studies revealed that abnormal reactive oxygen species (ROS) in mutant SOD1-expressing cells induced occludin phosphorylation, which facilitated the subsequent occludin ubiquitination mediated by the E3 ligase ITCH. Moreover, ubiquitinated occludin interacted with Eps15 to initiate its internalization, then trafficked to Rab5-positive vesicles and be degraded by proteasomes, resulting in a reduction in cell surface localization and total abundance. Notably, either ITCH or Eps15 knockdown was sufficient to rescue occludin degradation and ameliorate endothelial barrier disruption. In conclusion, our study reveals a novel mechanism of occludin degradation mediated by ALScausing SOD1 mutants and demonstrates a role for occludin in regulating BSCB integrity.

Automated summary

The study reveals a novel mechanism of occludin degradation mediated by ALS-causing SOD1 mutants, showing that abnormal ROS induces occludin phosphorylation and ubiquitination leading to occludin degradation. Knockdown of either ITCH or Eps15 rescues occludin degradation, ameliorates endothelial barrier disruption.