Figure 6.

Loss of SNX14 disrupts endoplasmic reticulum-associated neutral lipid metabolism. (A–D) Thin layer chromatography analysis of neutral lipids in SNX14^WT and SNX14^KO HEK293 cultured without (A, B) or with (C, D) exogenous cholesterol (Low-density lipoprotein; LDL). (A) Cholesterol esters (CE) decreased in SNX14^KO cells. (B) Triacylglycerides (TAG) increased in SNX14^KO cells on addition of U18666A. (C) CEs decreased when SNX14^KO cells were treated with U18666A and LDL. (D) TAG levels increased in SNX14^KO cells cultured with U18666A and LDL. N = 2, error bars = SD, **P ≤ 0.01, ns P ≥ 0.05, Student’s t-test. (E) Immunofluorescent images of U2OS cells that were either untreated, or treated with 600 μm oleic acid for 8 h. SNX14-FLAG (anti-FLAG) was observed to accumulate in peri-nuclear ring-like formations following oleic acid treatment (arrows). (F) Co-fluorescent imaging of SNX14-FLAG (Green), ER marker HSP90B1 (Red) and LD stain monodansylpentane (Blue). Scale bar = 5 μm.