Dynamics of a chemoattractant receptor in living neutrophils during chemotaxis
Dynamics of a chemoattractant receptor in living neutrophils during chemotaxis
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DOI：
10.1091/mbc.10.4.1163
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发表时间：
1999-04-01
影响因子：
3.3
通讯作者：
Bourne, HR
中科院分区：
文献类型：
--
作者：
Servant, G;Weiner, OD;Bourne, HR
中性粒细胞在浅趋化梯度中的持续定向运动增加了细胞相对于后部对前部趋化信号的敏感性的可能性。趋化剂受体重新分布到极化中性粒细胞的前极，可以通过增加细胞前缘检测到的信号的相对强度来施加不对称的敏感性。先前的实验对于运动中性粒细胞中受体的位置产生了矛盾的观察结果。为了在趋化过程中直接观察趋化剂受体，我们在白血病鳗鱼系 PLB-985 中表达了补体成分 C5a 的绿色荧光蛋白 (GFP) 标记受体。分化的 PLB-985 细胞，如中性粒细胞，响应均匀浓度的化学引诱剂而粘附、扩散和极化，并向含有化学引诱剂的微量移液器定向和爬行。在活细胞中记录的标记受体 C5aR-GFP 的荧光在极化和移动细胞的质膜上的任何地方都没有明显增加，即使在前缘也是如此。然而，在趋化过程中，一些细胞确实在前缘表现出大量高度折叠的质膜，如通过膜脂荧光探针检测到的那样。这伴随着 C5aR-GFP 荧光的明显增加，这与质膜的积累成正比。因此，中性粒细胞在趋化过程中不会主动将趋化剂受体集中在前缘，尽管膜的不对称分布可能会增加一些极化细胞前极的受体数量（相对于相邻的细胞质体积）。这种富集有助于维持化学引诱剂浅梯度下的持续迁移。
Persistent directional movement of neutrophils in shallow chemotactic gradients raises the possibility that cells can increase their sensitivity to the chemotactic signal at the front, relative to the back. Redistribution of chemoattractant receptors to the anterior pole of a polarized neutrophil could impose asymmetric sensitivity by increasing the relative strength of detected signals at the cell's leading edge. Previous experiments have produced contradictory observations with respect to receptor location in moving neutrophils. To visualize a chemoattractant receptor directly during chemotaxis, we expressed a green fluorescent protein (GFP)-tagged receptor for a complement component, C5a, in a leukemia eel line, PLB-985. Differentiated PLB-985 cells, like neutrophils, adhere, spread, and polarize in response to a uniform concentration of chemoattractant, and orient and crawl toward a micropipette containing chemoattractant. Recorded in living cells, fluorescence of the tagged receptor, C5aR-GFP, shows no apparent increase anywhere on the plasma membrane of polarized and moving cells, even at the leading edge. During chemotaxis, however, some cells do exhibit increased amounts of highly folded plasma membrane at the leading edge, as detected by a fluorescent probe for membrane lipids; this is accompanied by an apparent increase of C5aR-GFP fluorescence, which is directly proportional to the accumulation of plasma membrane. Thus neutrophils do not actively concentrate chemoattractant receptors at the leading edge during chemotaxis, although asymmetrical distribution of membrane may enrich receptor number, relative to adjacent cytoplasmic volume, at the anterior pole of some polarized cells. This enrichment could help to maintain persistent migration in a shallow gradient of chemoattractant.