Abnormal Crosstalk between Reelin-Dab1 and mTORC1 Signaling Pathways in Nonsyndromic Autism Spectrum Disorder

Poster Presentation
Friday, May 11, 2018: 5:30 PM-7:00 PM
Hall Grote Zaal (de Doelen ICC Rotterdam)
S. M. Sánchez-Sánchez1, J. Magdalon1, K. Griesi-Oliveira1, V. Abreu1, M. R. Passos-Bueno2 and A. L. Sertie1, (1)Centro de Pesquisa Experimental, Hospital Israelita Albert Einstein, Sao Paulo, Brazil, (2)Centro de Pesquisas Sobre o Genoma Humano e Células-Tronco, Instituto de Biociências, Universidade de São Paulo, Sao Paulo, Brazil
Background: Reelin-Dab1 and mTORC1 signaling pathways controls neuronal migration, layer formation, neurite outgrowth and plasticity of synapses in both the developing and the adult mammalian brain. Abnormal interplay between mTORC1- and Reelin/Dab1-signaling pathways has been implicated in the pathogenesis of syndromic Autism Spectrum Disorder (ASD). Recently, by using whole-exome sequencing in a subgroup of nonsyndromic ASD patients - in whom we found mTORC1 signaling hyperfunction, we identified rare and potentially damaging compound heterozygous variants in the gene encoding Reelin (RELN) in one patient (called as F2688). Although evidence from previous studies suggests that heterozygous loss-of-function variants in RELN can contribute to ASD, the molecular and cellular effects of RELN mutations identified in ASD patients are still poorly explored.

Objectives: To verify whether the variants we identified in RELN are functional and whether any crosstalk between Reelin-Dab1 and mTORC1 signaling pathways exists and potentiates neuropathological abnormalities.

Methods: We have generated iPSC-derived neural progenitor cells (NPCs) from patient F2688, from 5 idiopathic ASD patients (without RELN gene mutations) and from 5 control individuals. The gene expression levels of RELN were analyzed by RT-qPCR. The levels of Reelin protein in both whole NPC lysates and supernatants were analyzed by ELISA. Reelin and mTORC1 signal transduction was analyzed by RT-qPCR and western blot. Analysis of cell migration was performed using the Incucyte scratch wound cell invasion assay. F2688-derived NPCs were treated with rapamycin in order to verify whether an abnormal crosstalk exists between mTORC1 and Reelin-Dab1 pathways.

Results: We found that F2688-derived NPCs show impaired secretion of Reelin, impaired Reelin-Dab1 signaling (reduced levels of pDab1, pFyn and pSrc; and increased levels of total Dab1), hyperfunctional mTORC1 signaling (increased levels of RPS6K1, RPS6, pmTORC1 and pRPS6) and abnormal migration. Also, treatment of F2688-derived NPCs with rapamycin restored impaired Reelin-Dab1 signaling and migration.

Conclusions: Our results suggest that the variants we identified in the RELN gene are functionally relevant and suggest, for the first time, a dysfunctional interplay between Reelin-Dab1 and mTORC1 signaling pathways in nonsyndromic ASD.