Defining the Mechanism of Altered Very Long Chain Fatty Acid Metabolism in Neuro-Ichthyotic Disorders.

超长链脂肪酸延伸-4 （ELOVL4）基因突变导致以干鳞状皮肤（鱼鳞病）、认知缺陷、癫痫发作和年龄相关性小脑变性为特征的神经鱼鳞病。ELOVL4是介导超长链（≥C28）饱和脂肪酸（VLC-SFA）和多不饱和脂肪酸（VLC-PUFA）生物合成的必需酶，这些脂肪酸统称为超长链脂肪酸（VLC-FA）。ELOVL4在特定组织中表达，对VLC- SFA和/或VLC- pufa生物合成具有组织特异性选择性。在皮肤、大脑和睑板腺中，ELOVL4主要制造VLC- SFA，而在视网膜和睾丸中，ELOVL4主要制造VLC- pufa。然而，一种组织产生VLC-SFA和另一种VLC-PUFA的机制，以及这些VLC-FA如何在每种组织中发挥重要的保护作用，仍然未知。考虑到ELOVL4的突变影响皮肤和神经元组织，答案一定是不同突变蛋白的先天酶活性，或者突变蛋白对WT酶合成的VLC-FA的质量和数量有显性的负面影响。本提案的目的是研究哪些因素决定了VLC-SFA生物合成相对于VLC-PUFA的组织特异性选择性，以及突变体ELOVL4如何改变这些因素从而导致不同的组织特异性疾病。为了实现他的目标，我们建立了第一个ELOVL4突变（c.736T>G, p.W246G）的Long Evans大鼠模型，该突变导致脊髓小脑性共济失调34 （SCA34）伴变异性红细胞角化病（EKV）。与野生型对照大鼠相比，杂合子和纯合子SCA34大鼠从出生起就出现皮肤病变，后来表现出人类疾病典型的小脑功能障碍症状19。对皮肤VLC-SFA的分析表明，与WT对照组相比，纯合子KI组织仅占总VLC-SFA的33%左右。这些发现表明，该突变型SCA34模型的神经鱼鳞病变可能是由ELOVL4突变导致的皮肤和神经元VLC-FA水平下降引起的。我们假设组织特异性因素决定了哪种类型的VLC-FA（饱和或多不饱和）被制造，并且在SCA34患者中发现的皮肤和小脑病理缺陷是由于突变型ELOVL4酶对野生型ELOVL4合成VLC-FA的能力的影响。我们提出两个具体目标来检验这一假设。在成功完成所提出的实验后，我们将回答不同突变体ELOVL4蛋白如何影响VLC-FA生物合成从而导致神经鱼鳞疾病的未解决问题。我们预计这将使我们能够在未来的研究中寻求实验性治疗方案，这些研究可能会迅速进展到治疗一些人类突变ELOVL4疾病的人类临床试验。

Mutations in the Elongation of Very Long Chain Fatty Acids-4 (ELOVL4) gene cause neuro-ichthyotic disorder characterized by dry scaly skin (ichthyosis) and cognitive deficits, seizures and age-related cerebellar degeneration. ELOVL4 is an essential enzyme that mediates biosynthesis of very long chain (≥C28) saturated (VLC-SFA) and polyunsaturated fatty acids (VLC-PUFA) that are collectively called very long chain fatty acids (VLC-FA). ELOVL4 is expressed in specific tissues in which it exhibits tissue-specific selectivity towards VLC- SFA and/or VLC-PUFA biosynthesis. In the skin, brain and Meibomian glands, ELOVL4 makes mainly VLC- SFA, while in the retina and testes it makes VLC-PUFA. However, the mechanisms by which one tissue makes VLC-SFA and the other VLC-PUFA, and how these VLC-FA exert their important protective roles in each tissue, remain unknown. Considering the fact that mutations in ELOVL4 affect skin and neuronal tissues, the answer must be in either the innate enzyme activity of the different mutant proteins or in a dominant negative effect of the mutant proteins on the quality and quantity of VLC-FA synthesized by the WT enzyme. The objective of this proposal is to examine what factors determine the tissue-specific selectivity of VLC-SFA biosynthesis relative to VLC-PUFA and how mutant ELOVL4 alters such factors to cause the different tissue- specific disorders. To achieve his goal, we generated the first Long Evans rat model of ELOVL4 mutation (c.736T>G, p.W246G) that causes Spinocerebellar ataxia 34 (SCA34) with Erythrokeratodermia variabilis (EKV). Compared to wild type control rats, heterozygous and homozygous SCA34 rats developed skin lesions from birth and later showed signs of cerebellar dysfunction typical of the human disease19. Analysis of skin VLC-SFA showed that the homozygous KI tissue had only about 33% of total VLC-SFA compared to WT controls. These findings indicate that the neuro-ichthyotic pathology in this mutant SCA34 model is likely caused by decreased skin and neuronal VLC-FA levels, an effect of the ELOVL4 mutation. We hypothesize that tissue-specific factors determine which type of VLC-FA (saturated versus polyunsaturated) is made and that the skin and cerebellar pathological defects found in SCA34 patients result from effects of the mutant ELOVL4 enzyme on the wild type ELOVL4’s ability to synthesize VLC-FA. We propose two specific aims to test this hypothesis. Upon successful completion of the proposed experiments, we will answer unresolved questions of how the different mutant ELOVL4 proteins affect VLC-FA biosynthesis to cause neuro-ichthyotic disorders. We anticipate this will allow us to pursue experimental treatment options in future studies which could progress rapidly into human clinical trials for treating some of the human mutant ELOVL4 disorders.