利用MHV-68感染小鼠模型分析Shiftless对免疫细胞浸润的调控机制

Shiftless (SFL) is a host factor that inhibits programmed -1 ribosomal frameshifting (-1PRF), a translation recoding mechanism. In addition to translation recoding, -1PRF can regulate mRNA stability when -1PRF causes a premature stop codon. CCR5 is the major chemokine receptor to induce leukocyte trafficking to the site of inflammation in viral infection. The -1PRF of CCR5 mRNA results in a premature stop codon and thereby mRNA degradation through the nonsense-mediated mRNA decay (NMD) pathway. In our previous studies, we observed that in SFL knockout mice, the CCR5 mRNA stability was decreased, due to the enhanced -1PRF efficiency. Based on these results, we propose that SFL can influence the immune cell infiltration through regulation of CCR5 expression. In the present study, we will use the MHV-68 infection mouse model to test this hypothesis. MHV-68 will be used to infect wildtype and SFL-KO mice, followed by analyses of the differences in the phenotypes, such as infiltration of immune cells to the infection site (lung). In addition, bioinformatics analyses predicted that the mRNAs of other chemokines and their receptors may also have -1PRF signals. We will use SFL as a tool to prove or disprove these predictions. We will use the mouse model to further elucidate the biological consequences of SFL regulation of the expressions of these genes. The aim of this project is to understand the molecular mechanisms underlying the roles of SFL in regulating inflammatory infiltration in viral infection.

宿主因子Shiftless(SFL)能够抑制RNA翻译过程中的-1位核糖体移码（-1PRF），从而影响mRNA 的稳定性。CCR5是重要的趋化因子受体，介导CCR5+免疫细胞向炎症部位迁移。CCR5的mRNA上含有-1PRF信号，-1PRF的发生会导致Premature终止密码子出现，引发NMD途径降解mRNA。我们的前期结果表明，敲除内源SFL促进CCR5的-1PRF活性， CCR5的mRNA的稳定性降低。鉴于CCR5在免疫细胞趋化中的作用，我们提出假说：SFL通过抑制-1PRF，调控CCR5表达水平，从而影响免疫细胞趋化。本项目将利用SFL敲除小鼠，从体内体外两方面验证该假说，重点研究SFL对MHV-68感染引起的细胞趋化的调节作用。此外，我们将基于生物信息学预测结果，验证SFL对其他含有-1PRF的趋化因子及受体的调控，并利用SFL敲除小鼠来研究这种调控的生物学意义。

细胞蛋白Shiftless(SFL)抑制RNA翻译过程中的-1位核糖体移码（-1PRF）。在之前工作中，多方面实验结果指向SFL可能参与了免疫应答调控。本项目中我们利用SFL基因敲除小鼠，建立了MHV68病毒感染、李斯特菌感染和皮下肿瘤三种免疫模型，发现敲除SFL显著抑制CCR2,CCR5等单核巨噬细胞向炎症部位的趋化。我们进一步揭示了其可能的分子机制：SFL调控趋化因子家族成员的表达水平，参与天然免疫通路的活化，进而影响炎症发生及免疫细胞趋化迁移。该工作揭示了一种由Shiftless介导的免疫调控机制。