Limiting amounts of p27Kip1 correlates with constitutive activation of cyclin E-CDK2 complex in HTLV-I-transformed T-cells.
Human T-cells immortalized (interleukin-2 [IL-2] dependent) by the human T-cell lymphotropic/leukemia virus type I (HTLV-I), in time, become transformed (IL-2 independent). To understand the biochemical basis of this transition, we have used the sibling HTLV-I-infected T-cell lines, N1186 (IL-2 dependent) and N1186-94 (IL-2 independent), as models to assess the responses to antiproliferative signals. In N1186 cells arrested in G1 after serum/interleukin-2 (IL-2) deprivation, downregulation of the cyclin E-CDK2 kinase activity correlated with decreased phosphorylation of CDK2 and accumulation of p27Kip1 bound to the cyclin E-CDK2 complex, as seen in normal activated PBMCs (peripheral blood mononuclear cells). In contrast, N1186-94 cells failed to arrest in G1 upon serum starvation, displayed constitutive cyclin E-associated kinase activity, and, although CDK2 was partially dephosphorylated, the amount of p27Kip1 bound to the complex did not increase. This observation, extended to two other IL-2-dependent as well as to three IL-2-independent HTLV-I-infected T-cell lines, suggests that the lack of cyclin E-CDK2 kinase downregulation found in the late phase of HTLV-I transformation may correlate with insufficient amounts of p27Kip1 associated with the cyclin E-CDK2 complex. Reconstitution experiments demonstrated that the addition of p27Kip1 to lysates from N1186-94 starved cells resulted in the downregulation of cyclin E-associated kinase activity supporting the notion that the unresponsiveness of the cyclin E-CDK2 complex to growth inhibitory signals may be due to inadequate amounts of p27Kip1 assembled with the complex in HTLV-I-transformed T-cells. In fact, the amount of p27Kip1 protein was lower in most HTLV-I-transformed (IL-2-independent) than in the immortalized (IL-2-dependent) HTLV-I-infected T-cells. Furthermore, specific inhibitors of the phosphatidylinositol 3-kinase (P13K) induced an increase of p27Kip1 protein levels, which correlated with G1 arrest, in both IL-2-dependent and IL-2-independent HTLV-I-infected T-cells. Altogether, these results suggest that maintaining a low level of expression of p27Kip1 is a key event in HTLV-I transformation.
Cereseto
A
A
Basic Research Laboratory, Division of Basic Sciences, National Cancer Institute, Bethesda, Maryland 20892, USA.
Washington Parks
R
R
Rivadeneira
E
E
Franchini
G
G
D042846:CDC2-CDC28 Kinases; D002453:Cell Cycle; D018797:Cell Cycle Proteins; D002455:Cell Division; D002461:Cell Line, Transformed; D019927:Cyclin E / Q000378:metabolism*; D051357:Cyclin-Dependent Kinase 2; D050759:Cyclin-Dependent Kinase Inhibitor p21; D050760:Cyclin-Dependent Kinase Inhibitor p27; D018844:Cyclin-Dependent Kinases / Q000378:metabolism*; D016213:Cyclins / Q000235:genetics / Q000378:metabolism; D016147:Genes, Tumor Suppressor; D015490:HTLV-I Infections / Q000378:metabolism / Q000473:pathology; D015368:Human T-lymphotropic virus 1 / Q000472:pathogenicity*; D006801:Humans; D007376:Interleukin-2 / Q000378:metabolism; D008869:Microtubule-Associated Proteins / Q000235:genetics / Q000378:metabolism*; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D013601:T-Lymphocytes / Q000378:metabolism* / Q000473:pathology / Q000821:virology; D025521:Tumor Suppressor Proteins; D015854:Up-Regulation
D042846:CDC2-CDC28 Kinases; D018797:Cell Cycle Proteins; D019927:Cyclin E; D051357:Cyclin-Dependent Kinase 2; D050759:Cyclin-Dependent Kinase Inhibitor p21; D050760:Cyclin-Dependent Kinase Inhibitor p27; D018844:Cyclin-Dependent Kinases; D016213:Cyclins; D007376:Interleukin-2; D008869:Microtubule-Associated Proteins; D019869:Phosphatidylinositol 3-Kinases; D017346:Protein Serine-Threonine Kinases; D025521:Tumor Suppressor Proteins; C493956:CDKN1A protein, human; C495894:CDK2 protein, human