Tyrosine phosphorylation modulates binding preference to cyclin-dependent kinases and subcellular localization of p27Kip1 in the acute promyelocytic leukemia cell line NB4

Abstract

We have investigated the role of tyrosine phosphorylation of the cyclin-dependent kinase (cdk) inhibitor p27 Kip1 using the acute promyelocytic leukemia cell line NB4 together with granulocyte colonystimulating factor (G-CSF). Short-term G-CSF stimulation resulted in a rapid tyrosine dephosphorylation of p27 Kip1 accompanied by a change in its binding preferences to cdks. On G-CSF stimulation, p27 Kip1 dissociated from cdk4 and associated with cdk2. Binding assays with recombinant p27 Kip1 confirmed that tyrosine-phosphorylated p27 Kip1 preferentially bound to cdk4, whereas unphosphorylated protein preferentially associated with cdk2. In addition, studies with p27 Kip1 point mutations revealed a decisive role of Tyr88 and Tyr89 in binding to cdk4. Furthermore, phosphorylation of Tyr88 and Tyr89 was accompanied by strong nuclear translocation of p27 Kip1 .Taken together, this report provides the first evidence that tyrosine phosphorylation of p27 Kip1 plays a crucial role in binding to cdks and its subcellular localization. Moreover, both effects are mediated by application of G-CSF. (Blood.

Regulation of p27Kip1by mitogen-induced tyrosine phosphorylation

“…The discovery that p27 can be phosphorylated at tyrosine residues suggested a direct link between p27 and mitogen-induced signal transduction. [61][62][63] This link is supported by the observation that tyrosine phosphorylation of p27 is increased in G 1 phase and by mitogens. 63,64 Initially, the non-receptor kinases Lyn, Src, Yes, Abl and BCR-Abl were identified as tyrosine kinases that can phosphorylate p27.…”

Phosphorylation of p27Kip1 Regulates Assembly and Activation of Cyclin D1-Cdk4

“…28,2008 PKB PROMOTES p27-CYCLIN D1-Cdk4 ASSEMBLY 6463 appropriate antibodies, and the heat-stable cellular p27 was released from the immunoprecipitates by boiling in assembly buffer. Recovered p27 was titrated to ensure that equivalent p27 amounts were added to reaction mixtures, and their abilities to assemble recombinant cyclin D1-Cdk4 were then assayed as described above.…”

“…When differences in the expression levels of the p27 variants were VOL. 28,2008 PKB PROMOTES p27-CYCLIN D1-Cdk4 ASSEMBLY 6465…”

Exposure of neuroblastoma cell lines to imatinib results in the upregulation of the CDK inhibitor p27KIP1 as a consequence of c-Abl inhibition

“…Previous studies have demonstrated that overexpression of c-Abl in eukaryotic cells promotes p27 KIP1 phosphorylation on tyrosine residues [35] therefore, the potential involvement of c-Abl in p27 KIP1 tyrosine phosphorylation in neuroblastoma cell lines was investigated. To do this, neuroblastoma cells were transfected with siRNAs to c-Abl.…”