# OCRL1 Deficiency Affects the Intracellular Traffic of ApoER2 and Impairs Reelin-Induced Responses
## 4 Discussion
OCRL1 deficient cells significantly increased colocalizations of TfR with EEA1 and ApoER2, suggesting that other receptors could be affected in neurons, including Megalin/LRP2, which is also present in CNS and accomplishes relevant roles in the development [97; 98; 99]. Further, many neuronal cargoes are recycled via the Retromer complex [83]. In OCRL1 deficient cells, we found that the Retromer subunit VPS26 [83; 84; 85] showed an increased colocalization with internalized ApoER2 and EEA1. Interestingly, SNX27, the endosomal protein in charge of cargo recognition for retrieval with the Retromer complex [100; 101], binds to PI3P via its PX domain [93] and to phosphoinosititides bi- and tri-phosphates via its FERM domain [102], a domain required for the cargo recycling to the cell surface [103]. In vitro, SNX27 has increased affinity for PI4,5P\({}_{2}\) and PI3,4,5P\({}_{3}\) rather than PI3P [104], and it is transiently recruited to the immune synapse by PI3,4,5P\({}_{3}\)[102]. Since the loss of function of OCRL1 leads to increased PI4,5P2, this could explain the increased presence of the Retromer complex to EEA1 in association with SNX27. Conversely, in other LS models, there is a well-documented alteration of the PIPs from different intracellular compartments [6], affecting the recycling of cargoes to the TGN and the late endosomes/lysosomes for degradation [21]. Regarding the signaling triggered by the binding of Reelin to ApoER2, the AKT-mediated pathway is known to be crucial for dendritic growth and branching [49]. Similarly, OCRL1-deficient kidney cells exposed to insulin have decreased AKT activation [22]. Also, zebrafish embryos deficient in OCRL1 have lower basal AKT activation and total AKT levels [105]. Although we did not measure AKT levels under basal conditions, our results in LS show alterations in this pathway, suggesting a deficiency in Reelin-mediated neuronal development as part of the LS phenotype. In contrast, the same study in zebrafish found no differences in ERK activation and total levels between the LS model and the control [105]. Therefore, unlike the changes observed in AKT, the reduction in ERK signaling could be specific to the response triggered by Reelin, as we recently described in neurons deficient in AP-4 adaptor complex [75], in which ERK and CREB activation, involved in the regulation of synaptic function [106; 107] were reduced. Our results show a decrease in ERK activation, so some of the cognitive problems presented by some LS patients may be due to this deficient response. Another process that we found affected in the OCRL1 deficient neurons is Golgi deployment, a fundamental process for CNS development, as it precedes neuronal polarization and migration [81; 108]. It is known that Reelin is capable of activating the Golgi deployment [109] through regulation of the small GTPases Cdc42 and Rac1 modulated by the GEF \(\alpha\)PIX [48]. In non-neuronal LS models, an imbalance in the signaling of Rho GTPases has been reported [27; 87]. In this line, and unlike the reduced activation of AKT and ERK, Reelin's ability to induce Golgi deployment was almost absent in OCRL1 deficient neurons; this could imply an evident developmental alteration in LS patients. In this regard, in the _OCRL_ KO neurons, we noticed an apparent dispersion or fragmentation of the Golgi complex in the soma. Previous studies in different LS models, but not neurons, have extensively demonstrated a Golgi fragmentation in LS [17; 18; 26; 80], with a strong influence of the 5-phosphatase domain of OCRL1 [17; 18]; however, the mechanism and significance for the disease are unclear. Additionally, Golgi mobilization is required for Golgi outpost generation. These compartments are believed to have local roles in microtubule nucleation, protein trafficking, and secretion [109], which would potentially be affected in LS.