Université Pierre et Marie Curie, UMR S 872, Les Cordeliers, Paris, France.
PLoS One. 2013;8(1):e53017. doi: 10.1371/journal.pone.0053017. Epub 2013 Jan 2.
In enterocytes, the dynamic accumulation and depletion of triacylglycerol (TAG) in lipid droplets (LD) during fat absorption suggests that cytosolic LD-associated TAG contribute to TAG-rich lipoprotein (TRL) production. To get insight into the mechanisms controlling the storage/secretion balance of TAG, we used as a tool hepatitis C virus core protein, which localizes onto LDs, and thus may modify their protein coat and decrease TRL secretion. We compared the proteome of LD fractions isolated from Caco-2/TC7 enterocytes expressing or not hepatitis C virus core protein by a differential proteomic approach (isobaric tag for relative and absolute quantitation (iTRAQ) labeling coupled with liquid chromatography and tandem mass spectrometry). We identified 42 proteins, 21 being involved in lipid metabolism. Perilipin-2/ADRP, which is suggested to stabilize long term-stored TAG, was enriched in LD fractions isolated from Caco-2/TC7 expressing core protein while perilipin-3/TIP47, which is involved in LD synthesis from newly synthesized TAG, was decreased. Endoplasmic reticulum-associated proteins were strongly decreased, suggesting reduced interactions between LD and endoplasmic reticulum, where TRL assembly occurs. For the first time, we show that 17β-hydroxysteroid dehydrogenase 2 (DHB2), which catalyzes the conversion of 17-keto to 17 β-hydroxysteroids and which was the most highly enriched protein in core expressing cells, is localized to LD and interferes with TAG secretion, probably through its capacity to inactivate testosterone. Overall, we identified potential new players of lipid droplet dynamics, which may be involved in the balance between lipid storage and secretion, and may be altered in enterocytes in pathological conditions such as insulin resistance, type II diabetes and obesity.
在肠细胞中，脂肪吸收过程中脂滴（LD）中三酰基甘油（TAG）的动态积累和消耗表明细胞质 LD 相关的 TAG 有助于富含三酰基甘油的脂蛋白（TRL）的产生。为了深入了解控制 TAG 储存/分泌平衡的机制，我们使用丙型肝炎病毒核心蛋白作为工具，该蛋白定位于 LD 上，因此可能改变其蛋白外壳并减少 TRL 的分泌。我们通过差异蛋白质组学方法（相对和绝对定量同位素标记（iTRAQ）标记与液相色谱和串联质谱联用）比较了表达或不表达丙型肝炎病毒核心蛋白的 Caco-2/TC7 肠细胞中分离的 LD 级分的蛋白质组。我们鉴定了 42 种蛋白质，其中 21 种参与脂质代谢。脂滴包被蛋白 2/ADRP 被认为可以稳定长期储存的 TAG，在表达核心蛋白的 Caco-2/TC7 LD 级分中富集，而参与从新合成的 TAG 合成 LD 的脂滴包被蛋白 3/TIP47 则减少。内质网相关蛋白强烈减少，表明 LD 与内质网之间的相互作用减少，TRL 组装发生在内质网。这是首次表明，17β-羟甾脱氢酶 2（DHB2），它催化 17-酮向 17β-羟甾的转化，并且是表达核心蛋白的细胞中最丰富的蛋白质，定位于 LD 并干扰 TAG 的分泌，可能是通过其使睾酮失活的能力。总的来说，我们鉴定了潜在的新的脂滴动力学参与者，它们可能参与脂质储存和分泌之间的平衡，并且在胰岛素抵抗、2 型糖尿病和肥胖等病理条件下的肠细胞中可能会发生改变。
