Genetic and Functional Mechanisms in Citrate Transporter Disorder associated with SLC13A5

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Scientific Abstract: SLC13A5 epilepsy is a newly recognized form of Developmental Epileptic Encephalopathy 25 (DEE25) with seizures beginning within the first days of life along with subsequent intellectual and motor symptoms. In these patients, mutations in the SLC13A5 gene, which encodes a plasma membrane citrate transporter, result in a severe, early onset multi-focal epilepsy and cognitive and behavioral symptoms. How disruption of SLC13A5 function results in dysfunction of neural circuitry is unknown. In patients and in rodent loss of function models of SLC13A5, plasma citrate concentrations are elevated and cytoplasmic citrate is decreased. Since citrate is a precursor to neurotransmitters, diminished glial and neuronal citrate may result in abnormal neuro-transmitter metabolism, contributing to functional defects. However, SLC13A5 loss of function may not account for the full severity of the disorder, and truncations of SLC13A5 are rarely observed in patients. Instead, human genetics show that certain mutations are over-represented as known causative mutations; SLC13A5 G219R (DNA G655A) and T227M (DNA C680T) are the most common recurrent mutations found in approximately two-thirds of all known patients. While the epilepsy is associated with bi-alleleic mutations, the presence of recurrent missense mutations suggests mechanisms more complex than simple autosomal recessive genetics. However, these have not been fully investigated, and no specific treatments for these patients exist. In order to better understand the genetics of SLC13A5 epilepsy, we have developed novel experimental systems. In fly, the entire Drosophila Slc13A5 gene was replaced with the human SLC13A5 coding region. This results in expression of only the human SLC13A5 expressed in the central nervous system. In the humanized line, the G219R mutation causes lethality in contrast to the null, suggesting gain of function mechanisms. In rodents, we show that the equivalent mutation to G219R in mouse SLC13A5 causes more severe epilepsy in direct comparison to the null, again suggesting the over-arching hypothesis of this proposal: that pathogenic mutations in SLC13A5 have gain of function effects, as well as, loss of function effects. However, the understanding of the mechanisms underlying these effects is incomplete, and the determination of both genetic and functional mechanisms are highly important for developing treatments for SLC13A5 epilepsy. We will determine in three Aims to determine:1) what is the normal function of SLC13A5 in brain physiology 2) how do pathogenic mutations in SLC13A5 result in neural dysfunction 3) if novel therapeutic strategies may ameliorate symptoms in SLC13A5 syndrome.