Local accumulations of B-50/GAP-43 evoke excessive bleb formation in PC12 cells.
B-50 (GAP-43) is an axonal, plasma membrane-associated protein involved in growth cone morphology and function. We have conducted immunocytochemical, electron microscopic, and time-lapse experiments to visualize morphological consequences of local accumulations of B-50 at the plasma membrane of B-50-transfected PC-B2 cells, a clonal PC12 cell line with very low expression of endogenous B-50. The distribution of the transfected B-50 within these cells was inhomogeneous. At sites where the B-50 concentration was locally increased up to twofold, numerous filopodia were present in growth cone-like, substrate-attached regions. When local B-50 concentrations were even higher (up to 6.2-fold), blebs were formed, often containing vesicular structures, heavily decorated with B-50 immunoreactivity. Double labeling with f-actin binding phalloidin revealed that local B-50 accumulations were accompanied by increased actin filament concentrations. Colocalization of B-50 with actin filaments was prominent in filopodia, but was virtually absent in blebs, suggesting a disconnection of the bleb plasma membrane from the actin cytoskeleton. We conclude that B-50 evokes distinct effects on cell-surface activity in PC12 cells depending on its local concentration.
Aarts
L H
LH
Department of Physiological Chemistry, Rudolf Magnus Institute for Neurosciences, Utrecht University, The Netherlands.
Verkade
P
P
Schrama
L H
LH
Oestreicher
A B
AB
Gispen
W H
WH
Schotman
P
P
D000818:Animals; D002462:Cell Membrane / Q000502:physiology* / Q000648:ultrastructure; D019922:GAP-43 Protein / Q000235:genetics / Q000378:metabolism*; D018613:Microscopy, Confocal; D016253:Microscopy, Immunoelectron; D018715:Microscopy, Video; D009068:Movement; D016716:PC12 Cells; D051381:Rats; D011994:Recombinant Proteins / Q000378:metabolism; D014162:Transfection
D019922:GAP-43 Protein; D011994:Recombinant Proteins