An organized cytoskeleton is required for the cAMP-induced reverse transformation reaction in CHO-K1 cells. In the course of the reaction a considerable fraction of the genome changes its nuclease sensitivity. The current paper presents the following evidence that cAMP-induced phosphorylation of vimentin is an early step in this reaction complex. (i) Vimentin is only slightly phosphorylated in transformed CHO-K1 cells but is heavily phosphorylated in normal fibroblasts. (ii) cAMP addition almost triples the vimentin phosphorylation of CHO-K1 cells but does not change that of normal cells. (iii) Vimentin phosphorylation is one of the earliest phenomena to occur after addition of cAMP to CHO-K1 cells, preceding the cell-stretching reaction and other manifestations of reverse transformation. (iv) Indirect immunofluorescence experiments demonstrate that vimentin appears as a condensed mass in transformed CHO-K1 cells but cAMP addition restores the filamentous structure characteristic of the normal fibroblast. (v) Other transformed cells unresponsive to reverse transformation by cAMP failed to demonstrate increased phosphorylation of vimentin on treatment with cAMP. These results support the proposed scheme that phosphorylation of cytoskeletal elements initiates a large-scale genetic regulatory action in which a substantial change in the spectrum of genome exposure and sequestration occurs. A function for intermediate filaments in reverse transformation is implied.
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