pcDNA3．1(＋)-MER-Syk(L)载体构建及其对乳腺癌细胞增殖的影响

Construction of recombination vector of pcDNA3.1(+)-MER-Syk(L) and the effect of Syk(L) on the proliferation of breast cancer cells

目的:探讨乳腺癌细胞内4-OHT对MER-Syk(L)细胞定位的影响及对乳腺癌细胞增殖功能的影响。方法:采用不表达Syk的乳腺癌细胞系MDA-MB-231,建立稳定表达融合蛋白MER-Syk(L)的细胞株;蛋白质印迹和免疫荧光染色技术检测4-OHT处理前后融合蛋白MER-Syk(L)在细胞内的位置;二苯基溴化四氮唑蓝(MTT)法分析处理前后稳态细胞的增殖能力。结果:(1)无4-OHT的处理,融合蛋白MER-Syk(L)、MER-Syk(S)和MER均位于MDA-MB-231细胞浆内;4-OHT的作用下,融合蛋白MER-Syk(L)部分移位到细胞核内,而MER-Syk(S)和MER仍然位于细胞浆内;(2)MDA-MB-231细胞核内的MER-Syk(L)蛋白能够抑制细胞的增殖和生长,而细胞浆内的MER-Syk(L)、MER-Syk(S)和MER蛋白对细胞增殖和生长功能没有影响。结论:4-OHT可使融合蛋白MER-Syk(L)移位到MDA-MB-231细胞核内,从而抑制细胞的增殖。

AIM： To explore the effect of 4 - hydroxytomacifen （4-OHT） on MER-Syk（L） cellular localization and the function of Syk（L） on cell proliferation in breast cancer cells. METHODS ： pcDNA3.1 （ ＋ ） - MER - Syk （L） vector was constructed and the cell line MDA- MB -231, which expressed MER- Syk（L） stably, was established. Western blotting and immunofluorescence techniques were used to detect localization of MER - Syk （L） fusion protein in MDA - MB -231 cells cultured with or without 4 - OHT. MTT assay was used to explore the proliferation ability of MDA - MB -231 stable cells. RESULTS： （ 1 ） MER - Syk（L） fusion protein, not MER - Syk（S） and MER protein, translocated from cytoplasm to nucleus in the presence of 4 -OHT. （2） Nuclear not cytoplasmic MER- Syk （L） fusion protein inhibited MDA -MB -231 stable cell growth. （3） With or without the treatment of 4 -OHT, MER- Syk（S） and MER protein always located in cytoplasm and did not suppress cell growth. CONCLUSION： With 4 -OHT, MER -Syk（L） fusion protein translocates to nucleus and inhibits cell growth.