AJP - Cell Physiology, Vol 255, Issue 2 C202-C213, Copyright © 1988 by American Physiological Society
Nonrandom turnover of actin and tubulin in cultured rabbit cardiac fibroblasts
W. R. Mostow, A. G. Ferguson, M. Lesch, R. S. Decker and A. M. Samarel
Department of Medicine, Northwestern University Medical School, Chicago, Illinois 60611.
Total protein fractional rates of growth, synthesis, and degradation were
assessed in primary cultures of rabbit cardiac fibroblasts. Differences in
fractional growth rates were produced by subculturing cells at low density
and growing them to confluence. Total protein fractional degradative rates
were then derived by subtracting fractional growth rates from measured
fractional synthetic rates (obtained in [3H]leucine pulse-labeling
experiments). Actin and tubulin degradation were studied in similar rapidly
and slowly growing cultures. [35S]methionine pulse-chase experiments,
followed by dodecyl sulfate-polyacrylamide gel electrophoresis,
fluorography, and densitometry were used to determine the amount of labeled
actin and tubulin remaining in cultures at various times during the chase
(0-96 h). The indirect study showed a substantially lower total protein
fractional degradative rate during rapid vs. slow growth (0.04 +/- 0.13 vs.
0.42 +/- 0.01 d-1 at 2 and 15 days after subculture, respectively; P less
than 0.01). At both growth rates, the disappearance of labeled actin and
tubulin was delayed, suggesting a more complex model for their degradation
than random decay. Serum deprivation of slowly growing fibroblasts
increased the rate of disappearance of both proteins by eliminating the
delay in their breakdown. Thus the suppression of protein degradation
during rapid growth appears to result from the presence of relatively
greater amounts of "new" actin and tubulin (and possible other long-lived
proteins) that are kinetically distinct from the total intracellular pools
of these proteins with respect to their susceptibility to proteolysis.
