白血病抑制因子 LIF 第 29 位氨基酸 突变导致其功能完全丧失

白血病抑制因子 (LIF) 为一种多功能细胞因子，在生物发育及维持正常生理功能中发挥着重要的作用. LIF 在所有的哺乳动物中都具有高度的保守性，某些氨基酸在不同物种中保持不变. LIF 中 29 位氨基酸在所有的哺乳动物中都是Q. 为了研究 Q 对 LIF 功能的重要性，利用随机 PCR 的方法将 29 位氨基酸突变为 R，并将突变后的 LIF 克隆到真核表达载体 pcDNA6 ，在哺乳动物细胞中成功表达. 同时以克隆到的野生型 LIF 作为对照. 将分泌到细胞培养基中的野生型和突变 LIF 因子收集，通过 EMSA 实验以及荧光素酶报告系统检测，发现野生型 LIF 因子具有激活 STAT3 信号通路的生物学活性. 同时， ³H-TdR 掺入实验结果表明，野生型 LIF 因子能显著抑制鼠骨髓白血病细胞系 M1 的增殖. 而 29 位氨基酸突变的 LIF 因子则完全丧失了以上功能，但所发现的突变没有显性负的作用. 结果充分说明， LIF 中高度保守的 29 位氨基酸对 LIF 功能的维持具有重要的作用.

A LIF Mutation at The 29th Amino Acid Totally Abolished The Biological Functions

Leukemia inhibitory factor (LIF) plays important roles in varieties of biological processes. This factor is highly conserved in mammalian animals and only one heterozygous LIF mutation was reported to cause the infertility of women. A LIF mutation was generated and the evidences were provided that the mutation of mature LIF at the 29th amino acid totally abolished its functions, including stimulation of STAT activation assayed by Luciferase reporter gene expression and EMSA experiments. In addition, the mutated LIF failed to inhibit the proliferation of M1 cells. The data indicated that the mutation of LIF did not have a dominant negative effect but lost the biological functions, suggesting that the 29th amino acid is critical for maintaining the activities of LIF.