# OCRL1 Deficiency Affects the Intracellular Traffic of ApoER2 and Impairs Reelin-Induced Responses
## 4 Discussion  
Lowe syndrome is a multisystemic disorder that affects the function of the kidneys, eyes, and brain of the patients [6]. Even though the neurological manifestations can vary in severity among individuals, impacting significantly cognitive abilities and daily functioning of the patients, the precise mechanisms behind these symptoms are still being studied. Here, we report in two human neuronal cellular models that the loss of OCRL1 impacts the intracellular trafficking and signaling function of ApoE2, providing the first documented evidence that links Reelin signaling, essential for CNS development and function, to the pathogenesis of LS. At the cellular level, the deficiency in OCRL1 causes enlargement of early endosomes [21; 22; 80; 86], persistent perinuclear localization of lysosomes [34], reduction in spreading area [18; 25; 28], and defective actin dynamics [31; 32], possibly reflecting an irregular balance of Rac1/ RhoA activation [27; 28; 87]. Our neuronal models recapitulate the cellular phenotype of LS, with some differences depending on the model. In OCRL1 deficient H4 cells, we detected a partial but significant alteration in the recycling of ApoE2, explaining its reduction at the cell surface. The impaired recycling is supported by the increased presence of ApoE2 in endosomal compartments labeled by EEA1, TfR, and the Retromer complex. Besides, we found a small but significant increase in ApoE2 in LAMP1-positive late endosomes, which correlates with faster degradation and lower total levels of ApoE2 in the absence of OCRL1. Different PIPs localized at specific membranes allow the recruitment of various proteins, giving identity to the compartment [88; 89]. Accordingly, we report a decrease in the number of EEA1-positive particles in OCRL KO i3 neurons, where we also found a  
Figure 8: The absence of OCRL1 interferes with ApoE2/Reelin-induced Golgi deployment. I3-neurons, wild-type, and _OCRL_ KO were differentiated for 12 days, starved for 2 h, and treated with 10 nM Reelin or mock for 30 min. (**A**) Fixed neurons were stained with anti-GM130 (green), anti-MAP2 (magenta), and DAPI (blue). The arrowhead highlights Golgi complex into the dendrite. Scale bar = 10 \(\upmu\)m. (**B**) The distance between the outermost Golgi and the nucleus was measured in Fiji using a straight-line tool. At least 50 cells were measured from three experiments. Two-way ANOVA with Sidak’s multiple comparisons test was calculated with GraphPad ns: \(p>0.05\), \(***p<0.001\), \(***p<0.0001\). decreased presence of the endosomal protein SNX17 in dendrites. SNX17 is the cargo recognition protein associated with the Retriever complex [90]. A significant part of this protein is present in EEA1 endosomes [91; 92] since SNX17 binds preferentially to PI3P [93]. Then, a reduction in SNX17 in dendrites with fewer EEA1 endosomes is expected. Furthermore, since SNX17 is needed to recycle ApoER2 and, thus, activate the Reelin-triggered signaling properly [41], the reduction of SNX17 endosomal structures in dendrites may contribute to the decrease in ApoER2 levels and Reelin signaling we detected in LS neurons. Besides, as the SNX17, probably as part of the Retriever, is in charge of recycling other neuronal proteins, including LRP1 [92; 94], VLDL-R [95], and APP [96], we speculate that the function of these receptors could be affected in LS due to this pathway. Nevertheless, more specific research is needed to comprehend the effect of the loss of function of OCRL1 in the molecular mechanisms that regulate the sorting of proteins. The evident alterations in ApoER2 trafficking and protein levels in our neuronal cellular models with loss of OCRL1 indicate a widespread disruption of intracellular traffic with a change in the identity of the endosomal compartments, as we found in kidney cells [22].