By Gerald Litwack
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Extra info for Biochemical Actions of Hormones. Volume 11
9a,b,c), Bt2cAMP (Fig. 9d,e,f), BrcAMP (Fig. 9g,h,i), MIX alone (Fig. 9j,k,l), or isoproterenol plus MIX (Fig. 9m,n,o) (Steinberg and Agard, 1981a). Although Bt2cAMP stabilized R as before (Fig. 7), both BrcAMP and isoproterenol plus MIX destabilized R to the extent that it was undetectable in labeled patterns after 8 or 15 hours of chase. In label-chase experiments using shorter time intervals, the halflife of R in wild-type cells stimulated with BrcAMP or isoproterenol was similar to that of R in kin ~ cells.
A. Steinberg and C. S. Murphy, unpublished results). With Bt2cAMP as a selective agent, up to 2 X 106 wild-type S49 cells can be plated in a 6-cm diameter culture dish without adversely affecting the isolation of rare mutants (Friedrich and Coffino, 1977). 5 mM, and, when feeder cells are used, a cAMP phosphodiesterase inhibitor is also necessary to ensure efficient killing of sensitive cells. This difference in conditions required for efficient killing with and without feeder cells suggests that feeder cells metabolize Bt2cAMP.
We had reasoned that reduced levels of R in kin ~ cells reflected either a primary effect of the putative kin~ regulatory mutation or a secondary effect resulting from absence of functional C subunit. If interaction with C affected R subunit metabolism, Bt 2 cAMP-mediated kinase activation was expected to mimic effects of kin~ mutations. Relative rates of R synthesis in wild-type and kin~ cells were compared by pulse-labeling experiments (Steinberg and Agard, 1981b). R synthesis in kin~ cells was about half that in untreated wild-type cells.
Biochemical Actions of Hormones. Volume 11 by Gerald Litwack