B. Varnumfinney et Le. Reichardt, VINCULIN-DEFICIENT PC12 CELL-LINES EXTEND UNSTABLE LAMELLIPODIA AND FILOPODIA AND HAVE A REDUCED RATE OF NEURITE OUTGROWTH, The Journal of cell biology, 127(4), 1994, pp. 1071-1084
We have studied the role of vinculin in regulating integrin-dependent
neurite outgrowth in PC12 cells, a neuronal cell line. Vinculin is a c
ytoskeletal protein believed to mediate interactions between integrins
and the actin cytoskeleton. In differentiated PC12 cells, the cell bo
dy, the neurite, and the growth cone contain vinculin. Within the grow
th cone, both the proximal region of ''consolidation'' and the more di
stal region consisting of lamellipodia and filopodia contain vinculin.
To study the role of vinculin in neurite outgrowth, we generated vinc
ulin-deficient isolates of PC12 cell lines by transfection with vector
s expressing antisense vinculin RNA. In some of these cell lines, vinc
ulin levels were reduced to 18-23% of normal levels. In the vinculin-d
eficient cell lines, neurite outgrowth on laminin was significantly re
duced. In time-lapse analysis, growth cones advanced much more slowly
than normal. Further analysis indicated that this deficit could be exp
lained in large part by changes in the behaviors of filopodia and lame
llipodia. Filopodia were formed in normal numbers, extended at normal
rates, and extended to approximately normal lengths, but were much les
s stable in the vinculin deficient compared to control PC12 cells. Sim
ilarly, lamellipodia formed and grew nearly normally, but were dramati
cally less stable in the vinculin-deficient cells. This can account fo
r the reduction in rate of growth cone advance. These results indicate
that interactions between integrins and the actin-based cytoskeleton
are necessary for stability of both filopodia and lamellipodia.