University of Cambridge Metabolic Research Laboratories, Wellcome Trust-Medical Research Council Institute of Metabolic Science, University of Cambridge, Cambridge CB2 0QQ, UK.
Department of Medicine, Cambridge Institute for Medical Research, Cambridge Biomedical Campus, Hills Road, Cambridge, CB2 0XY, UK.
J Cell Sci. 2017 Oct 1;130(19):3234-3247. doi: 10.1242/jcs.206425. Epub 2017 Aug 3.
Galectins are a family of lectin binding proteins expressed both intracellularly and extracellularly. Galectin-3 (Gal-3, also known as LGALS3) is expressed at the cell surface; however, Gal-3 lacks a signal sequence, and the mechanism of Gal-3 transport to the cell surface remains poorly understood. Here, using a genome-wide CRISPR/Cas9 forward genetic screen for regulators of Gal-3 cell surface localization, we identified genes encoding glycoproteins, enzymes involved in N-linked glycosylation, regulators of ER-Golgi trafficking and proteins involved in immunity. The results of this screening approach led us to address the controversial role of N-linked glycosylation in the transport of Gal-3 to the cell surface. We find that N-linked glycoprotein maturation is not required for Gal-3 transport from the cytosol to the extracellular space, but is important for cell surface binding. Additionally, secreted Gal-3 is predominantly free and not packaged into extracellular vesicles. These data support a secretion pathway independent of N-linked glycoproteins and extracellular vesicles.
半乳糖凝集素是一类细胞内和细胞外表达的凝集素结合蛋白。半乳糖凝集素-3（Galectin-3，也称为 LGALS3）在细胞表面表达；然而，Gal-3 缺乏信号序列，其向细胞表面运输的机制仍知之甚少。在这里，我们使用全基因组 CRISPR/Cas9 正向遗传筛选来寻找调控 Gal-3 细胞表面定位的调节剂，鉴定出编码糖蛋白、参与 N 连接糖基化的酶、内质网-高尔基体运输调节剂以及参与免疫的蛋白质的基因。该筛选方法的结果促使我们解决 N 连接糖基化在 Gal-3 向细胞表面运输中的有争议作用。我们发现，N 连接糖蛋白成熟对于 Gal-3 从细胞质向细胞外空间的运输不是必需的，但对于细胞表面结合是重要的。此外，分泌的 Gal-3 主要是游离的，而不是包装到细胞外囊泡中。这些数据支持一种独立于 N 连接糖蛋白和细胞外囊泡的分泌途径。
