新发现Numb基因突变导致尿酸代谢异常引发高尿酸血症与痛风的分子机制研究

Primary hyperuricemia and gout is considered to be the result of interaction between environmental and genetic factors. However, the pathogenesis remains largely known. In the preliminary study, we discovered a gout-linked missense mutation in Numb gene, R630H, in a large pedigree of primary gout using whole genome sequencing technology. To our knowledge, no genetic variant in Numb gene was found to be related with hyperuricemia and gout up to now. Further studies showed that R630H mutation significantly reduces the expression of Numb and affects regulation of urate transporter protein MRP4 by Numb. At the same time, we found that Numb mutation causes the significant increase in the production of the IL-1β by monocyte treated with monododium urate crystal (MSU). These results suggest that Numb R630H mutation may cause gout by causing dysregulation in both uric acid metabolism and inflammation. In this study, Numb mutations screening will be carried out in more familial and sporadic gout patients. On the other hand, further in vitro and in vivo studies using R630H mutation knockin animal model will be carried out on the mechanism of how Numb mutation causes the defect in uric acid metabolism. This study is promising to define new pathogenic gene of hyperuricemia and gout, and identify new regulatory pathway of uric acid metabolism, leading to the identification of new molecular targets for the prevention and treatment of gout.

原发性高尿酸血症和痛风是遗传因素和环境因素相互作用的结果，其发病机制尚未阐明。课题组前期在原发性痛风大家系中发现位于Numb基因编码区的错义突变R630H，据了解尚没有 Numb基因变异与高尿酸血症和痛风相关的报道。进一步研究发现R630H突变显著减少Numb表达，并影响其对尿酸盐转运体蛋白MRP4的调节，同时发现突变明显增加尿酸盐晶体刺激下单核细胞分泌IL-1β。这提示Numb R630H突变可能通过引起尿酸代谢紊乱和炎症反应失调导致痛风病发生。本研究将一方面在更多痛风家系和散发患者中筛查Numb基因突变，另一方面从体外细胞水平和突变的基因敲入动物水平，着重探讨Numb R630H突变导致尿酸代谢紊乱的分子机制。本研究将有希望确定新的原发性高尿酸血症和痛风病致病基因，并阐明新的调控尿酸代谢的分子通路，从而为疾病防治提供新的分子靶点。

遗传因素在原发性痛风与高尿酸血症的发生中具有重要作用，致病基因或易感基因的发现可极大的推进疾病发病机制和尿酸代谢机制的研究。本项目前期通过全基因组测序技术在一遗传性痛风大家系中发现NUMB基因编码区错义突变R630H。在本项目的资助下，我们从以下四方面开展对NUMB R630H突变的致病性和NUMB参与尿酸代谢分子机制的探讨：其一，研究NUMB R630H突变是否影响NUMB的功能；其二，研究NUMB缺失与NUMB R630H突变是否影响肾小管上皮细胞尿酸转运；其三，研究NUMB促进肾小管上皮细胞尿酸排泄的机制；其四，利用基因敲入小鼠模型研究NUMB R630H突变是否导致尿酸排泄障碍从而导致血尿酸升高。研究发现NUMB R630H突变导致NUMB细胞内分布异常，并被自噬系统清除；NUMB促进肾小管上皮细胞的尿酸排泄，敲除NUMB后肾小管上皮细胞尿酸排泄显著减少；NUMB R630H突变导致NUMB促尿酸排泄的功能下降；NUMB与尿酸转运蛋白ABCG2相互作用，二者在内吞再利用区（endocytotic recycling compartment，ERC）共定位。NUMB介导ABCG2的内吞后分选，从而促进ABCG2分布到肾小管上皮细胞顶膜，敲除NUMB导致ABCG2在ERC中堆积，而在顶膜的分布显著减少。NUMB R630H突变基因敲入小鼠经肾尿酸排泄显著减少，血尿酸水平升高。本研究完成既定目标，从人类遗传学、体外以及模式生物水平证明NUMB R630H突变是痛风与高尿酸血症的致病突变，并阐明了该突变的致病机制，而更为重要的是揭示了NUMB通过介导关键尿酸转运蛋白ABCG2的极性分布而促进尿酸排泄这一崭新的尿酸代谢调控机制，NUMB可能作为细胞内外信号调节尿酸排泄的作用靶点。我们的研究对于阐明尿酸代谢机理，发现痛风与高尿酸血症的药物干预靶点具有重要价值。