Investigating the role of eukaryotic translation elongation factor eEF1A2 in autism, epilepsy and intellectual disability

Eukaryotic translation elongation factor eEF1A2 is responsible for delivering aminoacylated tRNAs to the ribosome during protein synthesis. Heterozygous de novo missense mutations in EEF1A2 have been identified in individuals with epilepsy, autism and intellectual disability. The primary aim of this thesis was to assess whether these mutations operated through a loss of function, gain of function or dominant negative mechanism. To investigate this, I analysed eEF1A2 protein interactions, how these interactions varied between mutations and the functional consequences of changes in the interactome. I used affinity purification mass spectrometry to identify the interactome of eEF1A2 and compared the changes resulting from mutations in eEF1A2. Co-immunoprecipitation experiments determined that mutations could be grouped by changes in their interaction with the cognate guanine exchange factor eEF1B. One group showed complete or partial loss of binding to all four subunits of the eEF1B complex, whilst another cluster showed no change in binding compared to wildtype eEF1A2. No specific patterns of clinical features or phenotypic severity could be attributed to these groupings. My results suggested that protein synthesis would be impaired by this disrupted interaction. In vivo and in vitro protein synthesis experiments failed to detect any differences that could be attributed to the presence or absence of the D252H mutation. As no apparent differences in protein synthesis could be detected between muscle tissue from Eef1a2⁻/⁻ and Eef1a2⁺/⁺ mice or between eEF1A2⁻/⁻ and wild-type cells, it is likely that in both cases the protein synthesis assay employed was not performing adequately. Comparative analysis of Eef1a2ᴰ²⁵²ᴴ/ᴰ²⁵²ᴴ and Eef1a2⁻/⁻ mouse phenotypes determined that there is a gain of function or dominant negative element to the eEF1A2ᴰ²⁵²ᴴ mutation, highlighting that this mutation does not operate simply through a loss of function mechanism. Interactome was unable to successfully identify what this might be. In conclusion eEF1A2 mutations may operate via both a loss and gain of function mechanism. Depending on the precise mutation, eEF1A2 may be compromised in its ability to operate in protein synthesis, but at least some mutations may also result in a degree of toxicity.