Molecular mechanisms underlying interferon-alpha-induced G0/G1 arrest: CKI-mediated regulation of G1 Cdk-complexes and activation of pocket proteins.
One prominent effect of IFNs is their cell growth-inhibitory activity. The mechanism behind this inhibition of proliferation is still not fully understood. In this study, the effect of IFN-alpha treatment on cell cycle progression has been analysed in three lymphoid cell lines, Daudi, U-266 and H9. Examination of the growth-arrested cell populations shows that Daudi cells accumulate in a G0-like state, whereas U-266 cells arrest later in G1. H9 cells are completely resistant to IFN-alpha's cell growth-inhibitory effects. The G0/G1-phase arrest is preceded by a rapid induction of the cyclin-dependent kinase inhibitors (CKIs), p21 and p15. In parallel, the activities of the G1 Cdks are significantly reduced. In addition to p21/p15 induction, IFN-alpha regulates the expression of another CKI, p27, presumably by a post-transcriptional mechanism. In the G1 Cdk-complexes, there is first an increased binding of p21 and p15 to their respective kinases. At longer exposure times, when Cdk-bound p15 and p21 decline, p27 starts to accumulate. Furthermore, we found that IFN-alpha not only suppresses the phosphorylation of pRb, but also alters the phosphorylation and expression of the other pocket proteins p130 and p107. These data suggest that induction of p21/p15 is involved in the primary IFN-alpha response inhibiting G1 Cdk activity, whereas increased p27 expression is part of a second set of events which keep these Cdks in their inactive form. Moreover, elevated levels of p27 correlated with a dissociation of cyclin E/Cdk2-p130 or p107 complexes to yield cyclin E/Cdk2-p27 complexes. In resistant H9 cells, which possess a homozygous deletion of the p15/p16 genes and lack p21 protein expression, IFN-alpha causes no detectable changes in p27 expression and, furthermore, no effects are observed on either pocket proteins in this cell line. Taken together, these data suggest that the early decline in G1 Cdk activity, subsequent changes in phosphorylation of pocket proteins, and G1/G0 arrest following IFN-alpha treatment, is not primarily due to loss of the G1 kinase components, but result from the inhibitory action of CKIs on these complexes.
Sangfelt
O
O
Department of Oncology/Pathology, Karolinska Hospital and Institute, Stockholm, Sweden.
Erickson
S
S
Castro
J
J
Heiden
T
T
Gustafsson
A
A
Einhorn
S
S
Grandér
D
D
D002051:Burkitt Lymphoma / Q000188:drug therapy / Q000378:metabolism / Q000473:pathology; D042846:CDC2-CDC28 Kinases; D002352:Carrier Proteins / Q000187:drug effects / Q000378:metabolism; D018797:Cell Cycle Proteins; D051357:Cyclin-Dependent Kinase 2; D051358:Cyclin-Dependent Kinase 4; D050762:Cyclin-Dependent Kinase Inhibitor p15; D019941:Cyclin-Dependent Kinase Inhibitor p16; D050759:Cyclin-Dependent Kinase Inhibitor p21; D050760:Cyclin-Dependent Kinase Inhibitor p27; D018844:Cyclin-Dependent Kinases / Q000037:antagonists & inhibitors* / Q000187:drug effects / Q000378:metabolism*; D016213:Cyclins / Q000187:drug effects / Q000378:metabolism; D016193:G1 Phase / Q000187:drug effects*; D006801:Humans; D016898:Interferon-alpha / Q000378:metabolism / Q000494:pharmacology*; D008869:Microtubule-Associated Proteins / Q000187:drug effects / Q000378:metabolism; D009101:Multiple Myeloma / Q000188:drug therapy / Q000378:metabolism / Q000473:pathology; D009687:Nuclear Proteins / Q000187:drug effects / Q000378:metabolism; D010750:Phosphoproteins / Q000187:drug effects / Q000378:metabolism; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000187:drug effects / Q000378:metabolism; D017027:Protein Tyrosine Phosphatases / Q000187:drug effects / Q000378:metabolism; D011506:Proteins; D011518:Proto-Oncogene Proteins; D016192:Resting Phase, Cell Cycle / Q000187:drug effects*; D016160:Retinoblastoma Protein / Q000187:drug effects / Q000378:metabolism; D050720:Retinoblastoma-Like Protein p107; D050717:Retinoblastoma-Like Protein p130; D014407:Tumor Cells, Cultured; D025521:Tumor Suppressor Proteins; D020687:cdc25 Phosphatases
C493956:CDKN1A protein, human; C493966:CDKN2B protein, human; D002352:Carrier Proteins; D018797:Cell Cycle Proteins; D050762:Cyclin-Dependent Kinase Inhibitor p15; D019941:Cyclin-Dependent Kinase Inhibitor p16; D050759:Cyclin-Dependent Kinase Inhibitor p21; D016213:Cyclins; D016898:Interferon-alpha; D008869:Microtubule-Associated Proteins; D009687:Nuclear Proteins; D010750:Phosphoproteins; D011506:Proteins; D011518:Proto-Oncogene Proteins; C493844:RBL1 protein, human; D016160:Retinoblastoma Protein; D050720:Retinoblastoma-Like Protein p107; D050717:Retinoblastoma-Like Protein p130; D025521:Tumor Suppressor Proteins; D050760:Cyclin-Dependent Kinase Inhibitor p27; D017346:Protein Serine-Threonine Kinases; D042846:CDC2-CDC28 Kinases; C495894:CDK2 protein, human; C495900:CDK4 protein, human; D051357:Cyclin-Dependent Kinase 2; D051358:Cyclin-Dependent Kinase 4; D018844:Cyclin-Dependent Kinases; C517004:CDC25A protein, human; D017027:Protein Tyrosine Phosphatases; D020687:cdc25 Phosphatases