NADP+/NADPH是细胞内保持还原性环境的关键，是参与细胞合成代谢和抗氧化应激的重要辅因子。NADK催化NAD+磷酸化为NADP+，控制细胞内NADP+/NADPH的含量，但是NADK的活性调节机制仍然不清楚。申请人通过质谱、Co-IP等实验发现ERK1/2结合并磷酸化NADK的第48和50位丝氨酸；体外酶活实验表明ERK1/2可以激活NADK的活性。在此基础上，申请人将利用多种体外和细胞实验进一步证明ERK1/2通过磷酸化NADK促进NADP+/NADPH合成，并探究其在肿瘤细胞生长中的功能。结合前人的研究，进一步证明ERK1/2和AKT协同性的磷酸化激活NADK，并且在KRas激活突变的肿瘤细胞中探讨它们对肿瘤细胞生长的作用。本项目研究揭示了新的NADK活性调节机制，丰富了细胞内NADP+/NADPH代谢的调控方式，进一步证明了NADK在肿瘤细胞中的重要功能。

NADP+/NADPH is the key to maintain the reducing environment in cell, and is an important co-factor involved in cell anabolism and anti-oxidant stress. NADK catalyzes the phosphorylation of NAD+ to NADP+ and controls the content of NADP+/NADPH in cells, but the mechanism of NADK activity regulation remains unclear. Through experiments such as mass spectrometry and Co-IP assay, the applicant found that ERK1/2 binds to NADK and phosphorylates its serine at positions 48 and 50; in vitro enzyme activity experiments showed that ERK1/2 can activate NADK activity. On this basis, the applicant will use a variety of in vitro and cellular experiments to further demonstrate that ERK1/2 promotes NADP+/NADPH synthesis through phosphorylating NADK and explore its function in tumor cell growth. Combined with previous research, the applicant will further prove that ERK1/2 and AKT synergistically phosphorylating and activating NADK, and explore their effect on cell growth in tumor cells with KRas activation mutations. The research in this project will reveal a new mechanism for regulating NADK activity, enrich the regulation of NADP+/NADPH metabolism in cells, and further highlight the important function of NADK in tumor cells.