Dissecting seipin function: the localized accumulation of phosphatidic acid at ER/LD junctions in the absence of seipin is suppressed by Sei1p(ΔNterm) only in combination with Ldb16p.
Dissecting seipin function: the localized accumulation of phosphatidic acid at ER/LD junctions in the absence of seipin is suppressed by Sei1p(ΔNterm) only in combination with Ldb16p.
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DOI：
10.1186/s12860-015-0075-3
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发表时间：
2015-12-04
期刊：
影响因子：
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通讯作者：
Goodman JM
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文献类型：
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作者：
Han S;Binns DD;Chang YF;Goodman JM
胞质脂滴的正确组装需要Seipin。在酵母菌素同源物Sei1p（原Fld1p）缺失的情况下，液滴从内质网出芽缓慢，表面缺乏正常的蛋白质成分，大小和形状高度不均匀，经常出芽进入细胞核，并促进内质网的局部增殖，并在其中缠结。但seipin催化脂滴形成的机制尚不清楚。Seipin防止磷脂酸（PA点）在er液滴连接处的局部积累。用Opi1p、Spo20p（51-91）和Pah1p三种探针检测PA点。液滴诱导系统表明，在液滴形成之前，PA点不存在；无论液滴是由三酰甘油还是steryl酯组成，斑点都出现了。缺失菌株被用来证明单一的磷脂酸产生酶不负责鱼点的产生，鱼点仍然抵抗代谢磷脂酸的酶的过度表达，这表明这种脂质被困在一个潜在的隔室中。PA点的抑制需要Sei1p （Nterm）的前14个氨基酸，该结构域对液滴组装的起始也很重要。与最近的证据一致，Ldb16p和Sei1p形成一个功能单元，ldb16Δ sei1Δ菌株的PA点表型被人类seipin拯救。此外，在表达Sei1pΔNterm的sei1Δ菌株中，点PA被Ldb16p的过表达所抑制，这表明Nterm与该蛋白存在功能相互作用。Sei1p和Ldb16p均过表达，但不单独表达Sei1p，足以引起液滴数量的大量增加。然而，在ldb16Δ sei1Δ背景下，Ldb16p单独增加了三酰甘油的积累。我们假设seipin可以防止内质网/液滴连接处形成具有极端曲率的膜，从而吸引磷脂酸。虽然Ldb16p单独可以影响三酰甘油的积累，但正确的液滴形成需要Sei1p和Ldb16的协同作用。本文的在线版本（doi:10.1186/s12860-015-0075-3）包含补充材料，可供授权用户使用。
Seipin is required for the correct assembly of cytoplasmic lipid droplets. In the absence of the yeast seipin homolog Sei1p (formerly Fld1p), droplets are slow to bud from the endoplasmic reticulum, lack the normal component of proteins on their surface, are highly heterogeneous in size and shape, often bud into the nucleus, and promote local proliferation of the endoplasmic reticulum in which they become tangled. But the mechanism by which seipin catalyzes lipid droplet formation is still uncertain. Seipin prevents a localized accumulation of phosphatidic acid (PA puncta) at ER-droplet junctions. PA puncta were detected with three different probes: Opi1p, Spo20p(51–91) and Pah1p. A system of droplet induction was used to show that PA puncta were not present until droplets were formed; the puncta appeared regardless of whether droplets consisted of triacylglycerol or steryl ester. Deletion strains were used to demonstrate that a single phosphatidic acid-producing enzyme is not responsible for the generation of the puncta, and the puncta remain resistant to overexpression of enzymes that metabolize phosphatidic acid, suggesting that this lipid is trapped in a latent compartment. Suppression of PA puncta requires the first 14 amino acids of Sei1p (Nterm), a domain that is also important for initiation of droplet assembly. Consistent with recent evidence that Ldb16p and Sei1p form a functional unit, the PA puncta phenotype in the ldb16Δ sei1Δ strain was rescued by human seipin. Moreover, PA puncta in the sei1Δ strain expressing Sei1pΔNterm was suppressed by overexpression of Ldb16p, suggesting a functional interaction of Nterm with this protein. Overexpression of both Sei1p and Ldb16p, but not Sei1p alone, is sufficient to cause a large increase in droplet number. However, Ldb16p alone increases triacylglycerol accumulation in the ldb16Δ sei1Δ background. We hypothesize that seipin prevents formation of membranes with extreme curvature at endoplasmic reticulum/droplet junctions that would attract phosphatidic acid. While Ldb16p alone can affect triacylglycerol accumulation, proper droplet formation requires the collaboration of Sei1p and Ldb16. The online version of this article (doi:10.1186/s12860-015-0075-3) contains supplementary material, which is available to authorized users.