Lipid droplet (LD) formation from the endoplasmic reticulum (ER) is accompanied by the targeting and accumulation of specific hydrophobic, membrane-embedded proteins on LDs. The determinants of this process are unknown. Here, we study the hydrophobic membrane motifs of two Drosophila melanogaster proteins, GPAT4 and ALG14, that utilize this pathway, and we identify crucial sequence features that mediate LD accumulation. Molecular dynamics simulations and studies in cells reveal that LD targeting of these motifs requires deeply inserted tryptophans that have lower free energy in the LD oil phase and positively charged residues near predicted hairpin hinges that become less constrained in the LD environment. Analyzing hydrophobic motifs from similar LD-targeting proteins, it appears that the distribution of tryptophan and positively charged residues distinguishes them from non-LD-targeting membrane motifs. Our studies identify specific sequence features and principles of hydrophobic membrane motifs that mediate their accumulation on LDs.

从内质网 (ER) 形成的脂滴 (LD) 伴随着特定疏水性膜嵌入蛋白在 LD 上的靶向和积累。这个过程的决定因素是未知的。在这里，我们研究了两种黑腹果蝇的疏水膜基序利用该途径的蛋白质 GPAT4 和 ALG14，我们确定了介导 LD 积累的关键序列特征。细胞中的分子动力学模拟和研究表明，这些基序的 LD 靶向需要深度插入的色氨酸，这些色氨酸在 LD 油相中具有较低的自由能，并且在预测的发夹铰链附近带正电荷的残基在 LD 环境中受到的限制较少。分析来自类似 LD 靶向蛋白的疏水基序，似乎色氨酸和带正电荷的残基的分布将它们与非 LD 靶向膜基序区分开来。我们的研究确定了介导它们在 LD 上积累的疏水膜基序的特定序列特征和原理。