Abnormal Cell Properties and Down-Regulated FAK-Src Complex Signaling In B Lymphoblasts of Autistic Subjects

Background:  

Recent studies suggest that one of the major pathways to the pathogenesis of autism is reduced cell migration, since the linkage between RELN and autism is a frequently replicated genetic finding. The Reelin protein, encoded by the RELN gene, plays a pivotal role in neuronal migration.  Focal adhesion kinase (FAK) has an important functional role in neural migration, dendritic morphology, axonal branching and synapse formation. The FAK/Src complex activated by up-stream reelin and integrin β1 can initiate a cascade of phosphorylation events to trigger multiple intracellular pathways including MEK/ERK and PI3K/Akt signaling.

Objectives:  

The aims of this study is to 1) determine whether integrin β1 and FAK-Src signaling are abnormally regulated in autism by using B lymphoblasts  as a study model, and 2) whether abnormal FAK-Src signaling leads to defects in B lymphoblast adhesion, migration, proliferation, and IgG production.

Methods:  

Both autistic lymphoblasts and control lymphoblasts from the normal siblings were obtained from Autism Genetic Resource Exchange (AGRE).  In this study, Western Blot Analyses were used to detect the expression and activity levels of Integrin, FAK, Src and Paxillin.   Cell Adhesion Assay, Modified Boyden Chamber Migration Assay and Cell Proliferation Assay were used to examine the lymphoblasts’ adhesion, migration and proliferation.  A human IgG ELISA kit was used to measure the immunoglobulin production.  Fluorescence Staining and Computer-assisted morphometry were performed to determine the morphology of lymphoblasts.

Results:  

For the first time, we show that 1) the protein expression of both integrin β1 and FAK are significantly decreased in autistic lymphoblasts and that 2) Src protein expression and the phosphorylation of active site (Y416) are also significantly decreased. These findings suggest there is reduced FAK-Src signaling activity in autistic lymphoblasts. We also found that lymphoblasts from autistic subjects exhibit significantly decreased migration and increased adhesion properties, as well as an impaired capacity for IgG production. Overexpression of FAK in autistic lymphoblasts was found to counter the adhesion and migration defects. In addition, we demonstrate that FAK mediates its effect through the activation of Src, PI3K/Akt and MEK signaling cascades and that Paxillin is also likely involved in the regulation of adhesion and migration in autistic lymphoblasts.

Conclusions:  

Our findings suggest that compromised Intergin/FAK/Src signaling and reduced paxillin activites could be responsible for the observed abnormal adhesion, migration and IgG production of B lymphoblasts in autism.   FAK mediates its effect in lymphoblast migration through the activation of Src, PI3K/Akt and MEK signaling cascades. These various abnormalities could be used to develop an approach for early autism diagnosis.