To characterize and compare the RAB39B protein expression and the endolysosomal trafficking function from two induced pluripotent stem cell (iPSC) lines derived from a Parkinson’s disease (PD) individual carrying the p.G192R RAB39B variant and his unaffected brother.
We previously reported a family with a classic PD phenotype in which a missense mutation in the RAB39B gene (c.574G>A; p.G192R) segregated with disease. Seven individuals (5 males and 2 females) were affected with a mean age at onset (AAO) of 46.1 years. The pattern of inheritance was X-linked dominant with reduced penetrance in females. The neuropathology of an affected male (AAO=31; deceased at age 54) showed neocortical Lewy bodies (LB), while another brother (AAO=53; deceased at age 65) presented no LB pathology.
We generated iPSCs from peripheral blood cells from both siblings. We differentiated the iPSCs into dopaminergic neurons (DN). We characterized endolysosomal trafficking utilizing antibodies for early (EEA1, Rab5), late endosomes (Rab11), lysosomes (LAMP1, Lysotracker), autophagic flux (p62), and lipid droplets (plin2). Additionally, we examined pathogenic protein aggregation by evaluating accumulation of insoluble ubiquitinated protein aggregates and alpha-synuclein oligomers.
There were no significant differences in RAB39B protein expression in the induced DNs. There were no significant differences in the early endosomal function, nor in the lysosome volume and size. We found significant differences in the p62-stained autophagolysosome volume and cellular compartment size between the affected subject's and the control's DNs and a dysfunction in lipid dynamics.
Our initial characterization supports the hypothesis that the p.G192R variant in RAB39B alters endolysosomal trafficking and lipid dynamics in RAB39B mutant DNs. We are currently evaluating protein aggregation and the implications of the observed alterations in lipid droplets on neurodegeneration. These studies will elucidate the role of RAB39B in PD pathogenesis and may reveal novel therapeutic targets for both genetic and idiopathic PD.