Cellular and Chemical Biology Unit, Institut Curie, Paris Sciences & Lettres Research University, U1143 INSERM, UMR3666 CNRS, 75248 Paris, France.
CurieCoreTech-Metabolomics and Lipidomics Platform, Institute Curie, 75248 Paris, France.
Biomolecules. 2024 Sep 18;14(9):1169. doi: 10.3390/biom14091169.
Essentially all plasma membrane proteins are glycosylated, and their activity is regulated by tuning their cell surface dynamics. This is achieved by glycan-binding proteins of the galectin family that either retain glycoproteins within lattices or drive their endocytic uptake via the clathrin-independent glycolipid-lectin (GL-Lect) mechanism. Here, we have used immunofluorescence-based assays to analyze how lattice and GL-Lect mechanisms affect the internalization of the cell adhesion and migration glycoprotein αβ integrin. In retinal pigment epithelial (RPE-1) cells, internalized αβ integrin is found in small peripheral endosomes under unperturbed conditions. Pharmacological compounds were used to competitively inhibit one of the galectin family members, galectin-3 (Gal3), or to inhibit the expression of glycosphingolipids, both of which are the fabric of the GL-Lect mechanism. We found that under acute inhibition conditions, endocytic uptake of αβ integrin was strongly reduced, in agreement with previous studies on the GL-Lect driven internalization of the protein. In contrast, upon prolonged inhibitor treatment, the uptake of αβ integrin was increased, and the protein was now internalized by alternative pathways into large perinuclear endosomes. Our findings suggest that under these prolonged inhibitor treatment conditions, αβ integrin containing galectin lattices are dissociated, leading to an altered endocytic compartmentalization.
基本上所有的质膜蛋白都被糖基化，它们的活性通过调节其细胞表面动力学来调节。这是通过半乳糖凝集素家族的糖结合蛋白来实现的，这些蛋白要么将糖蛋白保留在晶格中，要么通过网格蛋白非依赖性糖脂-凝集素（GL-Lect）机制驱动它们的内吞作用。在这里，我们使用基于免疫荧光的测定法来分析晶格和 GL-Lect 机制如何影响细胞黏附和迁移糖蛋白 αβ 整联蛋白的内化。在视网膜色素上皮（RPE-1）细胞中，在未受干扰的条件下，内化的 αβ 整联蛋白存在于小的外周内体中。我们使用药理学化合物竞争性抑制半乳糖凝集素家族成员之一半乳糖凝集素-3（Gal3），或抑制糖脂的表达，这两者都是 GL-Lect 机制的组成部分。我们发现，在急性抑制条件下，αβ 整联蛋白的内吞作用明显减少，这与以前关于该蛋白通过 GL-Lect 驱动的内化的研究结果一致。相比之下，在延长抑制剂处理后，αβ 整联蛋白的摄取增加，并且该蛋白现在通过替代途径被内化到大型核周内体中。我们的研究结果表明，在这些延长的抑制剂处理条件下，含有半乳糖凝集素晶格的 αβ 整联蛋白解离，导致内吞作用的区室化发生改变。
