补体因子I在斑马鱼松果体的表达调控及其功能研究

Complement factor I is an important component of complement pathway. Previous studies have shown CFI is regulated by different cytokines, and mailnly function as an inhibitory factor of complement pathway in adult animals. While CFI’s expression, function and its regulation in early development remain to be answered. .Our study has shown CFI is predominantly expressed in pineal gland in zebrafish early embryos, and preliminary experiment has proved it is rhythmical expressed and regulated by diffferent dark/light cycles. These results implies that CFI may function in circadian clockwork. The aims of this proposal were thus to examine how CFI are regulated by biological rhythms, dark/light cycles and circadian transcription factors, to investigate the role of CFI in circadian clock, and to explore the action mode. We plan to 1) study the expression pattern in early embryos by WISH and sectioned in situ hybridization; 2) clone and analyze CFI promoter; 3) examine CFI expression in different dark/light cycles and the regulation of CFI expression by circadian transcription factors; 4) analyze the role fo CFI in early development by knockdown or overexpression experiment; 5) construct CFI knockout model by Cas9/Talen; 6) observe the phenotype and locomotor activity of CFI knockout model and explore the possible action mode. The proposal will reveal a novel role of CFI in early development especially in circadian clockwork.

补体因子I (CFI) 是补体系统的重要因子，成体CFI受多种细胞因子的调控，主要起抑制补体途径的作用。而关于CFI在胚胎早期的表达调控以及功能均不明确。我们前期研究表明斑马鱼CFI在胚胎期在松果体有很强的特异性表达，提示CFI除调节补体途径外，可能在松果体发育中起重要作用。我们以斑马鱼为研究对象，拟1) 通过原位杂交、切片原位杂交等研究CFI的表达模式；2）克隆CFI的启动子，分析是否有松果体特异表达调控元件；3) 不同光周期处理及生物钟相关转录因子等抑制时，CFI的表达变化；4) 通过基因敲降、过表达等系统研究CFI在早期发育的功能，特别是是否与生物节律调节有关；5) Cas9/Talen方法建立敲除CFI的斑马鱼模型。6) 敲除CFI的斑马鱼的表型研究、运动活性改变以及作用机制探讨。基于以上研究，弄清CFI受生物节律、光周期等的调控，阐明CFI在松果体发育中的功能，探讨其作用机制。

补体因子I (CFI) 是补体系统的重要因子，成体CFI受多种细胞因子的调控，主要起抑制补体途径的作用。而关于CFI在胚胎早期的表达调控以及功能均不明确。我们的研究发现1）斑马鱼CFI在胚胎期有表达，主要在松果体和卵黄囊有较强表达2）CFI呈现节律性表达，其mRNA水平的表达在不同光周期表达不同，受光照的调控，在光照时其表达量明显高于黑暗时的表达。3) CFI在胚胎和成体中的表达均受LPS和LTA调控，参与免疫反应。4) Cas9/Talen方法建立了敲除CFI的斑马鱼模型，敲除CFI基因的斑马鱼早期发育并未出现明显异常。5) 敲降及敲除CFI的斑马鱼松果体发育未出现异常，这表明CFI可能并不参与松果体的发育。综上，我们的研究表明CFI参与免疫反应，且其早期的表达受生物节律、光周期等的调控。