Transforming growth factor beta(1) selectively inhibits the cyclic AMP-dependent proliferation of primary thyroid epithelial cells by preventing the association of cyclin D3-cdk4 with nuclear p27(kip1)

Dog thyroid epithelial cells in primary culture constitute a physiologicall y relevant model of positive control of DNA synthesis initiation and G0-S p rereplicative phase progression by cAMP as a second messenger for thyrotrop in (thyroid-stimulating hormone [TSH]). As previously shown in this system, the cAMP-dependent mitogenic pathway differs from growth factor cascades a s it stimulates the accumulation of p27(kip1) but not cyclins D. Neverthele ss, TSH induces the nuclear translocations and assembly of cyclin D3 and cd k4, which are essential in cAMP-dependent mitogenesis. Here we demonstrate that transforming growth factor beta(1) (TGF beta(1)) selectively inhibits the cAMP-dependent cell cycle in mid-G1 and various cell cycle regulatory e vents, but it weakly affects the stimulation of DNA synthesis by epidermal growth factor (EGF), hepatocyte growth factor, serum, and phorbol esters. E GF+serum and TSH did not interfere importantly with TGF beta receptor signa ling, because they did not affect the TGF beta-induced nuclear translocatio n of Smad 2 and 3. TGF beta inhibited the phosphorylation of Rb, p107, and p130 induced by TSH, but it weakly affected the phosphorylation state of Rb -related proteins in EGF+serum+treated cells. TGF beta did not inhibit c-my c expression. In TSH-stimulated cells, TGF beta did not affect the expressi on of cyclin D3, cdk4, and p27(kip)1, nor the induced formation of cyclin D 3-cdk4 complexes, but it prevented the TSH-induced relocalization of p27(ki p1) from cdk2 to cyclin D3-cdk4. It prevented the nuclear translocations of cdk4 and cyclin D3 without altering the assembly of cyclin D3-cdk4 complex es probably formed in the cytoplasm, where they were prevented from sequest ering nuclear p27(kip1) away from cdk2. This study dissociates the assembly of cyclin D3-cdk4 complexes from their nuclear localization and associatio n with p27(kip1). It provides anew mechanism of regulation of proliferation by TGF beta, which points out the subcellular location of cyclin D-cdk4 co mplexes as a crucial factor integrating mitogenic and antimitogenic regulat ions in an epithelial cell in primary culture.