Autism spectrum disorders (ASD) are characterized by inheritance of heterogeneous genetic risk factors proposed to cause the specific neurologic behavioural phenotype. The cellular correlates dysfunctional in ASD are targeted by pathways of neurogenesis and synaptic plasticity. In addition, biological networks such as energy metabolism and oxidative stress seem to be disturbed at the cellular level.
Objectives:
To distinguish between general ASD-specific and individual patient-specific cellular network perturbations we employed proteomic studies on lymphoblastoid cell lines (LCL) established from patients with a known mutation in the ribosomal protein L10 gene (RPL10) and a set of patients without any RPL10 mutation. This approach would report on protein expression patterns and consequently on functional pathways involved in the pathophysiology of ASD patients with different genetic backgrounds.
Methods:
In this study we applied 2D-differential-in-gel-electrophoresis (2D-DIGE) coupled with tandem mass spectrometry (MS/MS) to analyze the overall protein expression pattern in individual LCLs. Validation of differentially regulated proteins was performed at mRNA and protein expression level using qRT-PCR and Western Blot methods. We studied LCLs from members of two independent families carrying a RPL10-H213Q mutation in comparison to a non-mutation carrier of one family and healthy controls. To investigate if similar differentially regulated protein patterns could be observed in the general ASD population we analyzed a distinct set of 10 ASD patients not harbouring any RPL10 mutation in comparison with 10 random controls. A yeast model system was employed to validate and compare in wild-type and RPL10-deficient cells, respectively, a possible oxidative stress effect on protein expression.
Results:
In the RPL10 mutation analysis we discovered alterations in the expression level of different protein isoforms operative in glycolysis/energy metabolism (TPI1 and GAPDH), oxidative stress (ECH1) and mRNA regulation (HNRNPK). Then, in the additional set of LCLs from ASD patients who do not carry a RPL10 mutation we identified 19 differentially expressed candidates. These 19 proteins map to energy metabolism, mRNA and protein metabolism, cytoskeleton, and redox regulation. Therefore, the proteome profile of both experimental ASD groups overlap in energy metabolism and oxidative stress response, but with different protein representatives. Interestingly, most of the candidates have been previously described as redox-sensitive. Furthermore, the RPL10-deficient yeast cells under standard conditions show a subset of differentially expressed proteins that are observed in wild-type cells only under oxidative stress, and these again mapped to energy metabolism and oxidative stress response.
Conclusions:
We conclude that there is a redox-sensitive cellular response under conditions of RPL10 deficiency, which is part of an altered protein expression profile observed in the general ASD patient set. Of note, dysfunctions of energy metabolism and oxidative stress regulation as implicated by our results may be tolerated in LCLs, but be more deleterious in neuronal cells by causing a detrimental redox imbalance under the high oxygen pressure of neural cells. Thus, imbalance in energy/redox metabolism or a redox-sensitive dysfunction may lead to altered processes of neuronal development or function causing the ASD phenotype.
See more of: Cell Biology
See more of: Biological Mechanisms