Cdk2-dependent phosphorylation of the NF-Y transcription factor is essential for the expression of the cell cycle-regulatory genes and cell cycle G1/S and G2/M transitions

Abstract

We previously reported that cdk2 phosphorylates two serine residues near the DNA-binding domain of the YA subunit of NF-Y transcription factor and this phosphorylation is essential for DNA binding of NF-Y. In this study, we examined the effects of a phosphorylationdeficient mutant form of YA, YA-aa, in which the two serine residues are replaced with alanine, on the cell cycle and expression of the NF-Y target genes. Transient transfection assays show that YA-aa inhibits transcription from the NF-Y target promoters, such as cdc2, cyclin A, and cdc25C. Moreover, this inhibitory function of YA-aa can be suppressed by the expression of wild-type YA, implying that YA-aa inhibits transcription of those NF-Y target genes by inactivating wild-type YA. Since NF-Y target genes include the cell cycle-regulatory genes that ensure orderly progression of the cell cycle, we examined the effects of YA-aa in cell cycle progression. We constructed a recombinant adenovirus encoding YA-aa and found that YA-aa expression leads to repression of cell cycle-regulatory genes, such as cyclin A, RNR R2, DNA polymerase a, cdc2, cyclin B, and cdc25C. Consistently, YA-aa expression results in the inactivation of both cdc2 and cdk2. Furthermore, cell cycle analysis reveals that YA-aa induces cell cycle arrest at both G1 and G2/M. These results suggest that cdk2-dependent phosphorylation of NF-Y is essential for the expression of the cell cycle-regulatory genes and therefore for cell cycle progression at both G1/S and G2/M.

Deregulation of Cdk2 causes Bim-mediated apoptosis in p53-deficient tumors following actin damage

“…immunoprecipitation (IP)-kinase assay showed that while both Cdk2 and Cdc2 were not detected in HCT116 cells, both kinases remain unchanged in p53-and p21-knockout cells following PTX-2 treatment (Figure 1c). Therefore, inactivation of Cdk2 and Cdc2 kinases in p53-positive cells should be due to p21-mediated inhibition, supporting our previous report that p21-mediated Cdk2 inhibition lead to Cdc2 inactivation (Yun et al, 2003;Chae et al, 2004).…”

“…To select stable cell lines expressing Bcl-x L , we transfected p53-knockout cells with pPuro-Bcl-x L . A recombinant adenovirus encoding Cdk2dn was constructed as previously described (Chae et al, 2004).…”

A small molecule based on the pRb2/p130 spacer domain leads to inhibition of cdk2 activity, cell cycle arrest and tumor growth reduction in vivo

“…The crucial role of cdk2 activity is known to induce the expression of cell cycle-regulatory genes, thus leading to cell cycle progression (Chae et al, 2004;Yu et al, 2004). Therefore, inhibiting the phosphorylation ability of this kinase with the Spa310 small molecule is likely to induce cell cycle arrest.…”

Oocyte-based screening of cytokinesis inhibitors and identification of pectenotoxin-2 that induces Bim/Bax-mediated apoptosis in p53-deficient tumors

“…p21 induction causes subsequent arrest in the G1 phase by binding to cyclin E-and A-associated cdk2 complexes (Gu et al, 1993;Harper et al, 1993;Xiong et al, 1993). Recently, we reported that p53 and p21 also inactivate cyclin B-associated cdc2 kinase by repressing the transcription of cyclin B, cdc2, and cdc25C genes, and that this repression is mediated by the cdk2-dependent phosphorylation of NF-Y (Park et al, 2000;Yun et al, 2003;Chae et al, 2004). Other recent studies have described additional p53 target genes that are likely to activate cell cycle checkpoints and induce apoptosis (Vousden and Lu, 2002).…”