FTY720诱导1型1-磷酸鞘氨醇受体内化与其保守基序的关系研究
Chinese Journal of Immunology（2009）
Abstract
目的:研究FTY720诱导1型1-磷酸鞘氨醇受体(S1P1)内化与后者保守基序之间的关系.方法:以HA-S1P1(WT)-Myc-EGFP-N1融合表达载体为模板,应用重叠PCR的方法,将S1P1保守基序ERY突变为ENY,构建HA-S1P1(R142N)-Myc-EGFP-N1融合表达载体.测序鉴定后,经Polyfect转染入HEK293细胞.G418筛选出稳定细胞株.100 nmol/L FTY720处理3、6、12小时后,荧光显微镜下观察S1P1在HEK293细胞上的表达情况.结果:HA-S1P1(R142N)-Myc-EGFP-N1载体被成功构建,S1P1(WT)和S1P1(R142N)蛋白表达在HEK293稳定细胞株的表面,FTY720能诱导S1P1(WT)内化,但不能诱导S1P1(R142N)内化.结论:FTY720诱导S1P1内化与ERY保守基序有关.
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Key words
internalization,sphingosine 1-phosphate receptor type 1,fty720,fluorescence microscopy,genes,genetic engineering,gene expression
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