Zhakupova Assem, Debeuf Nincy, Krols Michiel, Toussaint Wendy, Vanhoutte Leen, Alecu Irina, Kutalik Zoltán, Vollenweider Peter, Ernst Daniela, von Eckardstein Arnold, Lambrecht Bart N, Janssens Sophie, Hornemann Thorsten
Institute of Clinical Chemistry, University Hospital Zurich, University of Zurich, Zurich, Switzerland.
Laboratory of Immunoregulation and Mucosal Immunology, Vlaams Instituut voor Biotechnologie (VIB) Inflammation Research Center, Ghent, Belgium.
FASEB J. 2016 Dec;30(12):4289-4300. doi: 10.1096/fj.201600639R. Epub 2016 Sep 19.
ORMDL proteins are believed to be negative regulators of serine palmitoyltransferase (SPT), which catalyzes the first and rate limiting step in sphingolipid (SL) de novo synthesis. Several single-nucleotide polymorphisms (SNPs) that are close to the ORMDL3 locus have been reported to increase ORMDL3 expression and to be associated with an elevated risk for early childhood asthma; however, the direct effect of ORMDL3 expression on SPT activity and its link to asthma remains elusive. In this study, we investigated whether ORMDL3 expression is associated with changes in SPT activity and total SL levels. Ormdl3-knockout (Ormdl3) and transgenic (Ormdl3) mice were generated to study the effect of ORMDL3 on total SL levels in plasma and tissues. Cellular SPT activity was measured in mouse embryonic fibroblasts from Ormdl3 mice, as well as in HEK293 cells in which ORMDL3 was overexpressed and silenced. Furthermore, we analyzed the association of the reported ORMDL3 asthma SNPs with plasma sphingoid bases in a population-based cohort of 971 individuals. Total C-long chain bases were not significantly altered in the plasma and tissues of Ormdl3 mice, whereas C-sphinganine showed a small and significant increase in plasma, lung, and liver tissues. Mouse embryonic fibroblast cells from Ormdl3 mice did not show an altered SPT activity compared with Ormdl3 and Ormdl3 mice. Overexpression or knockdown of ORMDL3 in HEK293 cells did not alter SPT activity; however, parallel knockdown of all 3 ORMDL isoforms increased enzyme activity significantly. A significant association of the annotated ORMDL3 asthma SNPs with plasma long-chain sphingoid base levels could not be confirmed. ORMDL3 expression levels seem not to be directly associated with changes in SPT activity. ORMDL3 might influence de novo sphingolipid metabolism downstream of SPT.-Zhakupova, A., Debeuf, N., Krols, M., Toussaint, W., Vanhoutte, L., Alecu, I., Kutalik, Z., Vollenweider, P., Ernst, D., von Eckardstein, A., Lambrecht, B. N., Janssens, S., Hornemann, T. ORMDL3 expression levels have no influence on the activity of serine palmitoyltransferase.
ORMDL蛋白被认为是丝氨酸棕榈酰转移酶（SPT）的负调节因子，SPT催化鞘脂（SL）从头合成的第一步且是限速步骤。据报道，几个靠近ORMDL3基因座的单核苷酸多态性（SNP）会增加ORMDL3的表达，并与儿童早期哮喘风险升高相关；然而，ORMDL3表达对SPT活性的直接影响及其与哮喘的联系仍不清楚。在本研究中，我们调查了ORMDL3表达是否与SPT活性和总SL水平的变化相关。构建了Ormdl3基因敲除（Ormdl3 -/-）和转基因（Ormdl3 -Tg）小鼠，以研究ORMDL3对血浆和组织中总SL水平的影响。在来自Ormdl3 -/-小鼠的小鼠胚胎成纤维细胞以及过表达和沉默ORMDL3的HEK293细胞中测量细胞SPT活性。此外，我们在一个由971名个体组成的基于人群的队列中分析了报道的ORMDL3哮喘SNP与血浆鞘氨醇碱的关联。在Ormdl3 -/-小鼠的血浆和组织中，总C -长链碱没有显著改变，而C -鞘氨醇在血浆、肺和肝组织中显示出小幅度但显著的增加。与Ormdl3 +/+和Ormdl3 -Tg小鼠相比，来自Ormdl3 -/-小鼠的小鼠胚胎成纤维细胞未显示出SPT活性改变。在HEK293细胞中过表达或敲低ORMDL3均未改变SPT活性；然而，同时敲低所有3种ORMDL同工型会显著增加酶活性。无法证实注释的ORMDL3哮喘SNP与血浆长链鞘氨醇碱水平之间存在显著关联。ORMDL3表达水平似乎与SPT活性的变化没有直接关联。ORMDL3可能在SPT下游影响鞘脂的从头代谢。 - 扎库波娃，A.，德伯夫，N.，克罗斯，M.，图森特，W.，万霍特，L.，阿莱库，I.，库塔利克，Z.，沃伦韦德，P.，恩斯特，D.，冯·埃卡德斯坦，A.，兰布雷希特，B. N.，扬森斯，S.，霍内曼，T. ORMDL3表达水平对丝氨酸棕榈酰转移酶的活性没有影响
