Kinase Riok2, A New Member Of The Apc / C Complexes

Control of cell cycle is an important research field of current life sience study,the cell cycle researches are not only an important way to understand life phenomenon,but also a vital method to cure cancer and tumor.APC/C(anaphase promoting complex/closome) is an ubiquitous E3 ubiquitin ligase of cycle control.APC/C is activated at anaphase of mitosis,and engages in cell cycle regulation.In mammales,APC/C is a 1.5MD molecular complex,consisting at least 11 subunits.Of them,Apc2 and Apcll together execute as catalyzing centre,and Apc10 can help substrates recognizing.We found a kinase interacting with Apc10 in yeast two hybrid system, and the kinase is highly homologous with Rio kinase2 in yeast,so we named it RIOK2.We' ve conducted coimmunoprecipitation and GST-pulldown experiment,and found that RIOK2 interact with Apc10 through its kinase domain at its N end,and it also interact with Apc11.We' ve expressed C end of RIOK2 in E.coli,and purified that by FPLC,then immunized New Zealand Rabbit,at last we got the antibody against RIOK2 with high specificity and sensitivity,which will benefit futher research a lot.Because APC/C is a key complex regulating cell cycle,we over-expressed RIOK2 in 293T cell and synchronized them by nocodazole. Using flow cytometry we tested and evaluated the effect RIOK2 exerting on cell cycle,and found that RIOK2 has delayed 293T cells at G₂/M period.