Kamiguchi H, Long K E, Pendergast M, Schaefer A W, Rapoport I, Kirchhausen T, Lemmon V
Department of Neurosciences, Case Western Reserve University, Cleveland, Ohio 44106, USA.
J Neurosci. 1998 Jul 15;18(14):5311-21. doi: 10.1523/JNEUROSCI.18-14-05311.1998.
Cell-cell interactions mediated via cell adhesion molecules (CAMs) are dynamically regulated during nervous system development. One mechanism to control the amount of cell surface CAMs is to regulate their recycling from the plasma membrane. The L1 subfamily of CAMs has a highly conserved cytoplasmic domain that contains a tyrosine, followed by the alternatively spliced RSLE (Arg-Ser-Leu-Glu) sequence. The resulting sequence of YRSL conforms to a tyrosine-based sorting signal that mediates clathrin-dependent endocytosis of signal-bearing proteins. The present study shows that L1 associates in rat brain with AP-2, a clathrin adaptor that captures plasma membrane proteins with tyrosine-based signals for endocytosis by coated pits. In vitro assays demonstrate that this interaction occurs via the YRSL sequence of L1 and the mu 2 chain of AP-2. In L1-transfected 3T3 cells, L1 endocytosis is blocked by dominant-negative dynamin that specifically disrupts clathrin-mediated internalization. Furthermore, endocytosed L1 colocalizes with the transferrin receptor (TfR), a marker for clathrin-mediated internalization. Mutant forms of L1 that lack the YRSL do not colocalize with TfR, indicating that the YRSL mediates endocytosis of L1. In neurons, L1 is endocytosed preferentially at the rear of axonal growth cones, colocalizing with Eps15, another marker for the clathrin endocytic pathway. These results establish a mechanism by which L1 can be internalized from the cell surface and suggest that an active region of L1 endocytosis at the rear of growth cones is important in L1-dependent axon growth.
在神经系统发育过程中，通过细胞粘附分子（CAMs）介导的细胞间相互作用受到动态调控。控制细胞表面CAMs数量的一种机制是调节它们从质膜的回收。CAMs的L1亚家族有一个高度保守的胞质结构域，其中包含一个酪氨酸，后面是可变剪接的RSLE（精氨酸-丝氨酸-亮氨酸-谷氨酸）序列。由此产生的YRSL序列符合基于酪氨酸的分选信号，该信号介导携带信号的蛋白质的网格蛋白依赖性内吞作用。本研究表明，L1在大鼠脑中与AP-2结合，AP-2是一种网格蛋白衔接蛋白，可捕获具有基于酪氨酸信号的质膜蛋白，以便被包被小窝内吞。体外实验表明，这种相互作用通过L1的YRSL序列和AP-2的μ2链发生。在L1转染的3T3细胞中，L1的内吞作用被显性负性发动蛋白阻断，该蛋白特异性破坏网格蛋白介导的内化作用。此外，内吞的L1与转铁蛋白受体（TfR）共定位，TfR是网格蛋白介导的内化作用的标志物。缺乏YRSL的L1突变体形式不与TfR共定位，表明YRSL介导L1的内吞作用。在神经元中，L1优先在轴突生长锥的后部被内吞，与Eps15共定位，Eps15是网格蛋白内吞途径的另一个标志物。这些结果建立了一种L1可以从细胞表面内化的机制，并表明生长锥后部L1内吞的活跃区域在L1依赖性轴突生长中很重要。
