TRANSLATIONAL REPRESSION OF ENDOGENOUS THYMIDINE KINASE MESSENGER-RNAIN DIFFERENTIATING AND ARRESTING MOUSE CELLS

We observed that decline of thymidine kinase (TK) enzyme activity was severalfold faster than the decay of full length TK mRNA during growth arrest of 3T6 mouse fibroblasts or during differentiation of myoblast s (C2Cl12) or F9 embryonal carcinoma cells. In order to study the mole cular mechanism of this disparate behavior, a polyclonal antiserum aga inst mouse TK was raised in rabbit. High level expression of mouse TK polypeptide in Escherichia coli was achieved with a T7 RNA polymerase- directed expression system. Using the antiserum in immunoblotting, no indication for a pool of inactive enzyme was found during differentiat ion of F9 or growth arrest of 3T6 cells. Pulse labeling of these cells in vivo with [S-35]methionine showed a more than 6-fold decrease in t he rate of TK-protein synthesis of in F9 cells after 3 days of treatme nt with retinoic acid as well as in 3T6 cells after 16 h under low ser um. This was not due to increased turnover of the protein as measured in pulse chase experiments. In addition, full length TK mRNA stayed as sociated with polysomes under these conditions in F9 as well as 3T6 ce lls. Taken together the results suggest that endogenous TK mRNA become s translationally repressed under a variety of conditions when mouse c ells cease to grow.