Cdc2/cyclin B1对中心体蛋白Nlp的调控在细胞生长中的作用

The Effect of Cdc2/cyclinB1 on Regulating Centrosome Protein Nlp in Cell Growth

目的研究中心体蛋白Nlp受Cdc2/cyclinB1磷酸化调控对细胞生长的影响。方法 Northern blot检测不同组织中Nlp的表达;EGFP-Nlp转染HeLa细胞,采用Double-Thymidine方法细胞同步化后释放,细胞免疫荧光方法观察Nlp在整个细胞周期中的亚细胞定位;构建Nlp磷酸化位点突变细胞系,采用人工计数和MTT法检测磷酸化位点突变对细胞生长的影响。结果 Nlp在不同组织中存在表达差异,Nlp在不同细胞周期呈现不同的亚细胞定位,磷酸化位点Ser185和Ser589的突变促进细胞的体外生长。结论 Cdc2/cyclin B1磷酸化位点突变后,Nlp获得了更强的促进细胞生长的能力。

Objective To study the effect of Cdc2/cyclinB1 on regulating centrosome protein Nlp in cell growth. Methods Nlp ex- pression in different tissues was detected by Northern blot. Transfection EGFP - Nip was transfected into HeLa cells, and then synchro- nized by Double -Thymidine. Nip subeellular localization throughout the ceil cycle was investigated by immunofluorescence. NIp phospho- rylation sites mutant cell lines were built. Phosphorylation site mutation on cell growth was detected by manually counting and MTT assay. Results Nip expression was distinct in different tissue. Nip showed different subcellular localization throughout the cell cycle. Phosphorylation sites Ser185 and Ser589 mutation promoted cell growth in vitro. Conclusion Nlp obtains stronger ability to promote cell growth after Cdc2/cvclin BI phosphorylation sites mutant.