Cell cycle regulation of the p34cdc2 inhibitory kinases.

Abstract

In cells of higher eukaryotic organisms the activity of the p34cdc2/cyclin B complex is inhibited by phosphorylation of p34cdc2 at two sites within its amino-terminus (threonine 14 and tyrosine 15). In this study, the cell cycle regulation of the kinases responsible for phosphorylating p34cdc2 on Thr14 and Tyr15 was examined in extracts prepared from both HeLa cells and Xenopus eggs. Both Thr14- and Tyr15- specific kinase activities were regulated in a cell cycle-dependent manner. The kinase activities were high throughout interphase and diminished coincident with entry of cells into mitosis. In HeLa cells delayed in G2 by the DNA-binding dye Hoechst 33342, Thr14- and Tyr15-specific kinase activities remained high, suggesting that a decrease in Thr14- and Tyr15- kinase activities may be required for entry of cells into mitosis. Similar cell cycle regulation was observed for the Thr14/Tyr15 kinase(s) in Xenopus egg extracts. These results indicate that activation of CDC2 and entry of cells into mitosis is not triggered solely by activation of the Cdc25 phosphatase but by the balance between Thr14/Tyr15 kinase and phosphatase activities. Finally, we have detected two activities capable of phosphorylating p34cdc2 on Thr14 and/or Tyr15 in interphase extracts prepared from Xenopus eggs. An activity capable of phosphorylating Tyr15 remained soluble after ultracentrifugation of interphase extracts whereas a second activity capable of phosphorylating both Thr14 and Tyr15 pelleted. The pelleted fraction contained activities that were detergent extractable and that phosphorylated p34cdc2 on both Thr14 and Tyr15. The Thr14- and Tyr15-specific kinase activities co-purified through three successive chromatographic steps indicating the presence of a dual-specificity protein kinase capable of acting on p34cdc2.

Andrographolide inhibits prostate cancer by targeting cell cycle regulators, CXCR3 and CXCR7 chemokine receptors

“…35 Our unpublished data shows AG affects Chk2 that could inhibit CDC25 family of protein, which in turn dephosphorylate (Wee1 phosphorylated) CDC2. [36][37][38] Activity of these phosphatases and kinases synchronize the firing of replication origin during S phase, which consequently controls the length of S phase. 39 Shortening of S phase could also be due to reduced p21 as low p21 levels are required for release from S phase arrest and cells with high p21 move slowly through S phase.…”

Hyperphosphorylation of the N-terminal Domain of Cdc25 Regulates Activity toward Cyclin B1/Cdc2 But Not Cyclin A/Cdk2

“…Single and double thymidine block/release synchronies of HeLa cultures were performed as described previously (19).…”

Localization of human Cdc25C is regulated both by nuclear export and 14-3-3 protein binding

“…Cell nuclei were observed by¯uorescence of FITC-dextran (left panels). LMB was added 60 min before injection HeLa cells were synchronized at the G1/S border using a double thymidine block and release protocol (Atherton-Fessler et al, 1994). Synchronized cells were then mock-infected or were infected with control adenoviruses or adenoviruses encoding mutant forms of Cdc25C (S216A, +NLS/S216A, or 7NES/S216A).…”