Dynamics of a chemoattractant receptor in living neutrophils during chemotaxis

Persistent directional movement of neutrophils in shallow chemotactic gradi ents raises the possibility that cells can increase their sensitivity to th e chemotactic signal at the front, relative to the back. Redistribution of chemoattractant receptors to the anterior pole of a polarized neutrophil co uld impose asymmetric sensitivity by increasing the relative strength of de tected signals at the cell's leading edge. Previous experiments have produc ed contradictory observations with respect to receptor location in moving n eutrophils. To visualize a chemoattractant receptor directly during chemota xis, we expressed a green fluorescent protein (GFP)-tagged receptor for a c omplement component, C5a, in a leukemia eel line, PLB-985. Differentiated P LB-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, fluoresc ence of the tagged receptor, C5aR-GFP, shows no apparent increase anywhere on the plasma membrane of polarized and moving cells, even at the leading e dge. During chemotaxis, however, some cells do exhibit increased amounts of highly folded plasma membrane at the leading edge, as detected by a fluore scent probe for membrane lipids; this is accompanied by an apparent increas e of C5aR-GFP fluorescence, which is directly proportional to the accumulat ion of plasma membrane. Thus neutrophils do not actively concentrate chemoa ttractant receptors at the leading edge during chemotaxis, although asymmet rical distribution of membrane may enrich receptor number, relative to adja cent cytoplasmic volume, at the anterior pole of some polarized cells. This enrichment could help to maintain persistent migration in a shallow gradie nt of chemoattractant.