Monier-Gavelle F, Duband J L
Laboratoire de Biologie Cellulaire du Développement, Institut Jacques Monod, Université Paris 7-Denis Diderot, France.
J Cell Sci. 1995 Dec;108 ( Pt 12):3839-53. doi: 10.1242/jcs.108.12.3839.
Dispersion of neural crest cells and their ultimate regroupment into peripheral ganglia are associated with precisely coordinated regulations both in time and space of the expression and function of cell adhesion receptors. In particular, the disappearance of N-cadherin from the cell surface at the onset of migration and its reexpression during cell aggregation suggest that, during migration, N-cadherin expression is repressed in neural crest cells. In the present study, we have analyzed in vitro the mechanism of control of N-cadherin expression and function in migrating neural crest cells. Although these cells moved as a dense population, each individual did not establish extensive and permanent intercellular contacts with its neighbors. However, cells synthesized and expressed mature N-cadherin molecules at levels comparable to those found in cells that exhibit stable intercellular contacts, but in contrast to them, the bulk of N-cadherin molecules was not connected with the cytoskeleton. We next determined which intracellular events are responsible for the instability of the N-cadherin junctions in neural crest cells using various chemical agents known to affect signal transduction processes. Agents that block a broad spectrum of serine-threonine kinases (6-dimethylaminopurine, H7 and staurosporine) or that affect selectively protein kinases C (bisindolylmaleimide and sphingosine), inhibitors of protein tyrosine kinases (erbstatin, herbimycin A, and tyrphostins), and inhibitors of phosphatases (vanadate) all restored tight cell-cell associations among neural crest cells, accompanied by a slight increase in the overall cellular content of N-cadherin and its accumulation to the regions of intercellular contacts. The effect of the kinase and phosphatase blockers was inhibitable by agents known to affect protein synthesis (cycloheximide) and exportation (brefeldin A), indicating that the restored cell-cell contacts were mediated chiefly by an intracellular pool of N-cadherin molecules recruited to the membrane. Finally, N-cadherin molecules were constitutively phosphorylated in migrating neural crest cells, but their level and state of phosphorylation were apparently not modified in the presence of kinase and phosphatase inhibitors. These observations therefore suggest that N-cadherin-mediated cell-cell interactions are not stable in neural crest cells migrating in vitro, and that they are under the control of a complex cascade of intracellular signals involving kinases and phosphatases and probably elicited by surface receptors.
神经嵴细胞的分散及其最终重新聚集成外周神经节，与细胞黏附受体的表达和功能在时间和空间上精确协调的调控相关。特别是，在迁移开始时N-钙黏蛋白从细胞表面消失，以及在细胞聚集过程中其重新表达，这表明在迁移过程中，神经嵴细胞中N-钙黏蛋白的表达受到抑制。在本研究中，我们在体外分析了迁移的神经嵴细胞中N-钙黏蛋白表达和功能的调控机制。尽管这些细胞以密集群体形式移动，但每个细胞个体并未与其相邻细胞建立广泛且持久的细胞间接触。然而，细胞合成并表达成熟的N-钙黏蛋白分子，其水平与那些表现出稳定细胞间接触的细胞相当，但与它们不同的是，大部分N-钙黏蛋白分子并未与细胞骨架相连。接下来，我们使用各种已知会影响信号转导过程的化学试剂，确定了哪些细胞内事件导致神经嵴细胞中N-钙黏蛋白连接的不稳定性。阻断广泛的丝氨酸-苏氨酸激酶的试剂（6-二甲基氨基嘌呤、H7和星形孢菌素）或选择性影响蛋白激酶C的试剂（双吲哚马来酰胺和鞘氨醇）、蛋白酪氨酸激酶抑制剂（埃布他汀、除莠霉素A和酪氨酸磷酸化抑制剂）以及磷酸酶抑制剂（钒酸盐），均能恢复神经嵴细胞之间紧密的细胞-细胞关联，同时N-钙黏蛋白的总体细胞含量略有增加，并在细胞间接触区域积累。激酶和磷酸酶阻断剂的作用可被已知会影响蛋白质合成（放线菌酮）和输出（布雷菲德菌素A）的试剂抑制，这表明恢复的细胞-细胞接触主要由招募到膜上的N-钙黏蛋白分子的细胞内池介导。最后，N-钙黏蛋白分子在迁移的神经嵴细胞中持续磷酸化，但其磷酸化水平和状态在存在激酶和磷酸酶抑制剂的情况下显然未发生改变。因此，这些观察结果表明，在体外迁移的神经嵴细胞中，N-钙黏蛋白介导的细胞-细胞相互作用不稳定，且它们受涉及激酶和磷酸酶且可能由表面受体引发的复杂细胞内信号级联的控制。
