Cold-inducible RNA-binding protein (CIRP) induces translation of the cell-cycle inhibitor p27Kip1.

The CDK inhibitor p27Kip1 plays a central role in controlling cell proliferation and cell-cycle exit. p27Kip1 protein levels oscillate during cell-cycle progression and are regulated by mitogen or anti-proliferative signaling. The abundance of the protein is frequently determined by post-transcriptional mechanisms including ubiquitin-mediated proteolysis and translational control. Here, we report that the cold-inducible RNA-binding protein (CIRP) selectively binds to the 5' untranslated region of the p27Kip1 mRNA. CIRP is induced, modified and relocalized in response to various stress stimuli and can regulate cell survival and cell proliferation particularly during stress. Binding of CIRP to the 5'UTR of the p27Kip1 mRNA significantly enhanced reporter translation. In cells exposed to mild hypothermia, the induction of CIRP correlated with increased translation of a p27Kip1 5'UTR reporter and with the accumulation of p27Kip1 protein. shRNA-mediated CIRP knockdown could prevent the induction of translation. We found that p27Kip1 is central for the decreased proliferation at lower temperature, since p27Kip1 KO mouse embryonic fibroblasts (MEFs) hardly increased their doubling time in hypothermic conditions, whereas wild-type MEFs significantly delayed proliferation in response to cold stress. This suggests that the CIRP-dependent p27Kip1 upregulation during mild hypothermia contributes to the cold shock-induced inhibition of cell proliferation.

The p27-Skp2 axis mediates glucocorticoid-induced cell cycle arrest in T-lymphoma cells.

Glucocorticoid therapy is an important treatment modality of hematological malignancies, especially T-cell acute lymphoblastic leukemia (T-ALL). Glucocorticoids are known to induce a cell cycle arrest and apoptosis in T-lymphoma cells. We could demonstrate that the cell cycle arrest induced by the synthetic glucocorticoid dexamethasone (Dex) clearly precedes apoptosis in human CEM T-ALL and murine S49.1 T-lymphoma cells. Cyclin D3 is strongly downregulated, whereas the CDK inhibitor p27 (Kip1) (p27) is strongly upregulated in response to dexamethasone in these cells. RNAi-mediated knockdown of p27 as well as overexpression of its negative regulator Skp2 revealed the critical function of p27 in the Dex-induced G 1 arrest of CEM cells. Our studies indicate that several mechanisms contribute to the increase of p27 protein in our T-lymphoma cell lines. We found a significant upregulation of p27 mRNA in S49.1 and CEM cells. In addition, Dex treatment activated the mouse p27 promotor in reporter gene experiments, indicating a transcriptional regulation. However, the relatively moderate induction of p27 mRNA levels by Dex did not explain the strong increase of p27 protein in CEM and S49.1 cells. We found clear evidence for a posttranslational mechanism responsible for the robust increase in p27 protein. Dex treatment of S49.1 and CEM cells increases the half-life of p27 protein, which indicates that decreased protein degradation is the primary mechanism of p27 induction by glucocorticoids. Interestingly, we found that Dex treatment decreased the protein and mRNA levels of the negative regulator of p27 protein and E3 ubiquitin ligase subunit Skp2. We conclude that the cell cycle inhibitor p27 and its negative regulator Skp2 are key players in the glucocorticoid-induced growth suppression of T-lymphoma cells and should be considered as potential drug targets to improve therapies of T-cell malignancies.