Phosphatidylinositol Cycle Disruption is Central to Atypical Hemolytic-Uremic Syndrome Caused by Diacylglycerol Kinase Epsilon Deficiency
| Autor: | Richard M. Epand, Vincent So, Christopher H. Choy, Alexis Traynor-Kaplan, Jing Wu, Roberto J. Botelho, Mathieu Lemaire |
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| Rok vydání: | 2019 |
| Předmět: |
0303 health sciences
Phospholipase C Kinase Phosphatidic acid Cell biology 03 medical and health sciences chemistry.chemical_compound 0302 clinical medicine chemistry 030220 oncology & carcinogenesis Lipidomics lipids (amino acids peptides and proteins) Human umbilical vein endothelial cell Arachidonic acid Phosphatidylinositol 030304 developmental biology Diacylglycerol kinase |
| Popis: | BackgroundLoss-of-function mutations in diacylglycerol kinase epsilon (DGKE) cause a rare form of atypical hemolytic-uremic syndrome (aHUS) for which there is no treatment besides kidney transplantation. Highly expressed in kidney endothelial cells, DGKE is a lipid kinase that phosphorylates diacylglycerol (DAG) to phosphatic acid (PA). Specifically, DGKE’s preferred substrate is 38:4-DAG, that is DAG containing stearic acid (18:0) and arachidonic acid (20:4). DAG is produced when phosphatidylinositol 4,5-bisphosphate (PtdIns(4,5)P2) is cleaved by phospholipase C (PLC). A better understanding of how DGKE deficiency impacts the endothelial lipid landscape is critical to developing a treatment for this condition.MethodsWe used orthogonal methods to compare the lipid levels in two novel models of DGKE deficiency to their respective controls: an immortalized human umbilical vein endothelial cell (iHUVEC) engineered with CRISPR/Cas9 and a blood outgrowth endothelial cell (BOEC) from an affected patient. Methods included mass spectrometry lipidomics, radiolabeling of phosphoinositides with [3H]myo-inositol, and live-tracking of a transfected fluorescent PtdIns(4,5)P2 biosensor.ResultsUnexpectedly, mass spectrometry lipidomics data revealed that high 38:4-DAG was not observed in the two DGKE-deficient models. Instead, a reduction in 38:4-PtdIns(4,5)P2 was the major abnormality.These results were confirmed with the other two methods in DGKE-deficient iHUVEC.ConclusionReduced 38:4-PtdIns(4,5)P2—but not increased 38:4-DAG—is likely to be key to the pro-thrombotic phenotype exhibited by patients with DGKE aHUS.TRANSLATIONAL STATEMENTMutations in DGKE cause a severe renal thrombotic microangiopathy that affects young children and leads to end-stage renal disease before adulthood. DGKE preferentially phosphorylates diacylglycerol to its corresponding phosphatidic acid (PA), which is then used to synthesize PtdIns(4,5)P2 via the phosphatidylinositol cycle. Understanding the disease pathophysiology is necessary to develop a treatment to prevent this outcome. This paper describes how we applied mass spectrometry lipidomics to two novel models of DGKE deficiency to investigate how this defect impacts the levels of diacylglycerol, PA and related phosphoinositides in endothelia. Unexpectedly, our data show that the critical abnormality caused by DGKE deficiency is not high diacylglycerol, but rather low PtdIns(4,5)P2. Restoring endothelial PtdIns(4,5)P2 homeostasis may be the cornerstone to treat these patients. |
| Databáze: | OpenAIRE |
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