Inhibition of mitogenesis in Balb/c-3T3 cells by Trichostatin A. Multiple alterations in the induction and activation of cyclin-cyclin-dependent kinase complexes.
Trichostatin A (TSA), a global repressor of histone deacetylase activity, inhibits the proliferation of a number of cell types. However, the identification of the mechanisms underlying TSA-mediated growth arrests has remained elusive. In order to resolve in more detail the cellular process modulated during the growth inhibition induced by TSA, we studied the effect of the drug on G(0)/G(1) traverse in mitogen-stimulated quiescent Balb/c-3T3 cells. Cyclin D1 and retinoblastoma proteins were induced following the mitogenic stimulation of both control and TSA-treated cells, and cyclin D1 formed complexes with CDK4 under both conditions. However, cyclin D1-associated kinase was not increased in growth-arrested cells. The lack of cyclin D-associated kinase was paralleled by an accumulation of RB in a hypophosphorylated form, as would be expected. In contrast, p130 became partially phosphorylated, accompanied by a marked increase in p130-dependent E2F DNA binding activity and a partial release of free E2F-4. Despite the presence of E2F complexes not bound to pocket proteins, late G(1) E2F-dependent gene expression was not observed. The lack of cyclin D1-associated kinase in TSA-treated cultures was potentially due to high levels of the cyclin-dependent inhibitor p27(kip1). However, the modulation of p27(kip1) levels by the deacetylase inhibitor cannot be responsible for the induction of the cell cycle arrest, since the growth of murine embryo fibroblasts deficient in both p27(kip1) and p21(cip1) was also inhibited by TSA. These data support a model in which TSA inhibits very early cell cycle traverse, which, in turn, leads to a decrease in cyclin D1-associated kinase activation and a repression of late cell cycle-dependent events. Alterations in early G(0)/G(1) gene expression accompany the TSA-mediated growth arrest.
Wharton
W
W
Molecular Oncology Program, H. L. Moffitt Cancer Center and Research Institute and Departments of Pathology, University of South Florida College of Medicine, Tampa, Florida 33612, USA.
Savell
J
J
Cress
W D
WD
Seto
E
E
Pledger
W J
WJ
D016475:3T3 Cells; D000818:Animals; D002453:Cell Cycle; D018797:Cell Cycle Proteins; D002455:Cell Division / Q000187:drug effects*; D019938:Cyclin D1 / Q000378:metabolism*; D050759:Cyclin-Dependent Kinase Inhibitor p21; D050760:Cyclin-Dependent Kinase Inhibitor p27; D018844:Cyclin-Dependent Kinases / Q000378:metabolism*; D016213:Cyclins / Q000502:physiology; D004247:DNA / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D015870:Gene Expression / Q000187:drug effects; D056572:Histone Deacetylase Inhibitors; D006877:Hydroxamic Acids / Q000494:pharmacology*; D051379:Mice; D008869:Microtubule-Associated Proteins / Q000502:physiology; D010982:Platelet-Derived Growth Factor / Q000494:pharmacology; D025521:Tumor Suppressor Proteins
C493957:Cdkn1a protein, mouse; C493961:Cdkn1b protein, mouse; D018797:Cell Cycle Proteins; D050759:Cyclin-Dependent Kinase Inhibitor p21; D016213:Cyclins; D004791:Enzyme Inhibitors; D056572:Histone Deacetylase Inhibitors; D006877:Hydroxamic Acids; D008869:Microtubule-Associated Proteins; D010982:Platelet-Derived Growth Factor; D025521:Tumor Suppressor Proteins; D019938:Cyclin D1; D050760:Cyclin-Dependent Kinase Inhibitor p27; C012589:trichostatin A; D004247:DNA; D018844:Cyclin-Dependent Kinases