Rocha Susana, De Keersmaecker Herlinde, Hutchison James A, Vanhoorelbeke Karen, Martens Johan A, Hofkens Johan, Uji-i Hiroshi
Molecular Imaging and Photonics, Faculty of Science and ‡Centre for Surface Chemistry and Catalysis, Faculty of Bioscience Engineering, KU Leuven , Belgium.
Langmuir. 2014 Apr 29;30(16):4743-51. doi: 10.1021/la500121f. Epub 2014 Apr 15.
Important cellular events such as division require drastic changes in the shape of the membrane. These remodeling processes can be triggered by the binding of specific proteins or by changes in membrane composition and are linked to phospholipid metabolism for which dedicated enzymes, named phospholipases, are responsible. Here wide-field fluorescence microscopy is used to visualize shape changes induced by the action of phospholipase A1 on dye-labeled supported membranes of POPC (1-palmitoyl-2-oleoly-sn-glycero-3-phosphocholine). Time-lapse imaging demonstrates that layers either shrink and disappear or fold and collapse into vesicles. These vesicles can undergo further transformations such as budding, tubulation, and pearling within 5 min of formation. Using dye-labeled phospholipases, we can monitor the presence of the enzyme at specific positions on the membrane as the shape transformations occur. Furthermore, incorporating the products of hydrolysis into POPC membranes is shown to induce transformations similar to those observed for enzyme action. The results suggest that phospholipase-mediated hydrolysis plays an important role in membrane transformations by altering the membrane composition, and a model is proposed for membrane curvature based on the presence and shape of hydrolysis products.
诸如细胞分裂等重要的细胞事件需要细胞膜形状发生剧烈变化。这些重塑过程可由特定蛋白质的结合或膜成分的变化触发，并与磷脂代谢相关，磷脂代谢由名为磷脂酶的特定酶负责。在此，利用宽场荧光显微镜观察磷脂酶A1作用于染料标记的1-棕榈酰-2-油酰-sn-甘油-3-磷酸胆碱（POPC）支持膜时诱导的形状变化。延时成像表明，这些膜层要么收缩消失，要么折叠塌陷成囊泡。这些囊泡在形成后5分钟内可经历进一步的转变，如出芽、成管和珠状化。使用染料标记的磷脂酶，我们可以在形状转变发生时监测酶在膜上特定位置的存在情况。此外，将水解产物掺入POPC膜中可诱导出与酶作用所观察到的类似转变。结果表明，磷脂酶介导的水解通过改变膜成分在膜转变中起重要作用，并基于水解产物的存在和形状提出了一个膜曲率模型。
