ER滞留信号肽对外源CTL表位进入胞内MHC-Ⅰ类抗原呈递途径的促进作用

An endoplasmic reticulum retrieval signal sequence promotes MHC class Ⅰ-presentation of exogenous CTL epitope peptides

目的探讨哺乳动物内质网(endoplasmic reticulum,ER)滞留信号肽(retrieval signal sequence)能否促进外源CTL表位肽进入抗原呈递细胞(antigen-presenting cell,APC)内MHC-Ⅰ类抗原呈递途径。方法应用多肽固相合成技术将哺乳动物内质网滞留信号——赖氨酸-天冬氨酸-谷氨酸-亮氨酸(Lys-Asp-Glu-Leu,KDEL)基序融合在H-2Kb限制性CTL表位OVA257-264的羧基端,同时合成该表位和氨基端自然延伸四个氨基酸(TEWT)的对照肽OVA257-268。选择巨噬细胞系Ana-1作为本研究的APC,采用流式细胞仪(FACS)分析技术,检测各抗原肽在Ana-1内的MHC-Ⅰ类抗原呈递动力学。结果羧基端KDEL基序可明显增强与之偶联的CTL表位在APC内的MHC-Ⅰ类抗原呈递效率,并且显著延长APC表面MHC/肽复合物的呈递时间。结论羧基端KDEL基序修饰是将外源肽有效导入APC内MHC-Ⅰ类抗原呈递途径的一个简单有效的新策略,可为肿瘤治疗性肽疫苗的分子设计与研究提供新思路和实验依据。

Objective To detect the effect of an endoplasmic reticulum （ER） retrieval signal peptide on MHC class Ⅰ-associated antigen presentation of exogenous CTL epitope peptides. Methods Three peptides were synthesized, which were the minimal CTL epitope peptide OVA257-264, OVA257-264-KDEL （with an ER retrieval signal Lys-Asp-Glu-Leu sequence at the carboxyl-terminus of OVA257-264）, and OVA257-268 （with four naturally extended residues TEWT at the carboxyl terminus of OVA257-264）. Flow cytometric analysis was used to detect time-dependent kinetics of MHC class Ⅰ antigen presentation by APC fed with these peptides. Results The exogenous peptides which were artificially fused with a carboxyl-terminal KDEL sequence, could be efficiently presented by intracellular MHC class Ⅰ molecules, and retain the capacity of APCs to display surface MHC/peptide complexes for a prolonged period. Conclusion The ER retrieval signal modification can be regarded as a novel method for targeting exogenous peptides into the intracellular MHC class Ⅰ presentation pathway, and may improve the molecule design of CTL epitope based-peptide vaccines.