Elongation of Very Long chain fatty acids-4 (ELOVL4) is an elongase responsible for biosynthesis of very long chain (eC28) fatty acids (VLCFA) in brain, retina, skin, and testes. Heterozygous inheritance of mutant ELOVL4 causes juvenile macular degeneration in autosomal dominant Stargardt's disease (STGD3), while children with homozygous inheritance of the STGD3 mutation develop severe dry/itchy skin, seizures, intellectual disability, and spastic quadriplegia (similar to that associated with cerebral palsy). Mice with recessive STDG3 or global ELOVL4 knockout die shortly after birth due to loss of VLCFA-containing omega-O- acyl-ceramides, which form a critical water barrier in the skin. Using a skin-rescue approach, the Anderson lab generated transgenic mice that express wild-type ELOVL4 driven by human, skin-specific promoters. The skin- rescued mice recapitulated the recessive STGD3 condition seen in humans in that they develop a central nervous system (CNS) phenotype that includes hyperactivity and seizures. I determined that ELOVL4 is expressed within neurons (not glia) in the sub-granular layer of the dentate gyrus in the hippocampus, a known locus for medial temporal lobe seizures. Fatty acid analysis of wild-type hippocampus confirmed the presence of 28:0 and 30:0, the same fatty acids required for forming the skin permeability barrier. Positron emission tomography (PET) showed significant increase in the amount of fluoro-deoxyglucose (FDG) uptake in the brain of our double mutant mice compared to littermate controls, suggesting increased neural metabolism. Subsequent MRI studies with gadolinium ruled out any break down of the blood brain barrier, suggesting that there is an increased metabolic demand in the brain cells of these seizure-prone animals. My hypothesis is that the loss of VLC-FA within the CNS due to mutations in ELOVL4 leads to impaired neural development and that results in neuronal dysfunction, hyper-excitability, and seizures.
The specific aims of this training proposal are: 1. To characterize the expression pattern of ELOVL4 and its products within the CNS: This will allow me to establish the location of the ELOVL4 enzyme and its products within the central nervous system and to draw novel connections between ELOVL4 and specific brain functions. 2. To determine the mechanism of neural dysfunction mediated by mutations in ELOVL4 and/or loss of VLC-FA: By comparing neuronal function in Elovl4-experssing cells from brains of wild type and skin-rescued lines, I wil test my hypothesis both in vitro from primary culture with FM-Dye studies to assess synaptic function and ex vivo using extracellular electrophysiology to assess the integrity of the hippocampal circuitry.
The effects of bi-allelic mutations in ELOVL4 in human children are multi-factorial causing the following brain problems: delayed myelination, seizures, and intellectual disability. This proposed research plan will advance not only our understanding of the neuropathology suffered by children with bi-allelic, recessive STGD3 mutations, it will also potentially identify several novel mechanisms for ELOVL4 and its VLC-FA products within the brain, which could open possibilities for additional therapies at the clinical level.