Parkinson’s disease LRRK2 mutations dysregulate iron homeostasis and promote oxidative stress and ferroptosis in human neurons and astrocytes

Background: Iron accumulation is a hallmark of sporadic and familial Parkinson's disease (PD) pathology and correlates with clinical motor symptom severity. The biochemical mechanisms driving iron dyshomeostasis in PD brain and whether these are early or late event in the neurodegenerative process remain unknown. Elevated nigral iron levels have been reported in LRRK2 mutation carriers, both in PD patients compared to idiopathic PD and in asymptomatic carriers relative to controls, suggesting that iron accumulation precedes clinical onset in LRRK2-associated PD. However, the precise consequence of pathogenic LRRK2 mutations on cellular iron handling within neurons and glial cells remains unclear. Methods: Here, we investigated different readouts of iron homeostasis in iPSCs and iPSC-derived neurons and astrocytes from PD patients harboring G2019S or R1441C/G LRRK2 mutations or healthy controls, as well as an isogenic iPSC panel with the same variants. By using high-content and super-resolution microscopy of iron-specific probes, we assayed iron content and distribution in cells and examined the downstream effects of iron dyshomeostasis on ferroptosis signaling. Results: We show that heterozygous LRRK2 mutations dysregulate cellular iron across iPSCs, neurons and astrocytes, in a kinase-dependent manner. Lysosomal ferrous iron storage was consistently elevated across iPSCs, iPSC-derived neurons and astrocytes carrying LRRK2 mutations. LRRK2 regulates Rab GTPase function through their direct phosphorylation, and our prior work revealed significant but divergent lysosomal phenotypes between Rab8a and Rab10 knockout models. Here, we report that Rab8a knockout recapitulates key aspects of LRRK2 mutation phenotypes on intracellular iron and ferritin levels, although with differences in magnitude and specificity. By contrast, Rab10 deficiency showed opposing effects, suggesting distinct roles for these well-established LRRK2 substrates in iron homeostasis. Finally, we show that basal lipid peroxidation and ROS levels are elevated in isogenic LRRK2 mutant neurons, while iron chelation was sufficient to reduce LRRK2-dependent ROS. Conclusions: Together, our findings demonstrate that LRRK2 mutations disrupt iron homeostasis across disease-relevant cell types and establish a discrete biochemical pathway linking LRRK2 signaling and vulnerability to ferroptosis. Ongoing work aims to further dissect the roles of Rab substrates in these pathways.