The kidney releases in the urine and circulation a non-polymerizing form of Uromodulin that retains the external hydrophobic patch domain

Uromodulin (Tamm-Horsfall protein, THP) is a glycoprotein uniquely produced in the kidney. It is released by cells of the thick ascending limbs (TAL) apically in the urine, and basolaterally in the renal interstitium and systemic circulation. Processing of mature urinary THP, which polymerizes into supra-molecular filaments, requires cleavage of an external hydrophobic patch (EHP) at the C terminus. However, THP in the circulation is not polymerized, and it remains unclear if non-aggregated forms of THP exist natively in the urine. We propose that an alternative processing path, which retains the EHP domain, can lead to a non-polymerizing form of THP. We generated an antibody that specifically recognizes THP with retained EHP (THP+EHP) and established its presence in the urine in a non-polymerized native state. Proteomic characterization of urinary THP+EHP revealed its C-terminus to end at F617. In the human kidney, THP+EHP was not only detected in TAL cells, but also diffusely in the renal parenchyma. Using C-terminus proteomic sequencing and immunoblotting, we then demonstrated that serum THP has also retained EHP. In a small cohort of patients at risk for acute kidney injury (AKI), admission urinary THP+EHP was significantly lower in patients who subsequently developed AKI during hospitalization. Our findings uncover novel insights into uromodulin biology by establishing the presence of an alternative path for cellular processing, which could explain the release of non-polymerizing THP in the circulation. Larger studies as needed to establish the utility of urinary THP+EHP as a sensitive biomarker of kidney health and susceptibility to injury.

尿调蛋白（Uromodulin，又称Tamm-Horsfall蛋白，THP）是一种仅由肾脏合成的糖蛋白。它由髓袢升支粗段（thick ascending limbs, TAL）细胞分泌：经顶端途径释放入尿液，经基底侧途径释放入肾间质与体循环。成熟尿液源性THP可聚合为超分子丝，其加工过程需切除C端的外部疏水结构域（external hydrophobic patch, EHP）。然而，循环中的THP并未发生聚合，目前仍不清楚尿液中是否天然存在非聚集形式的THP。本研究提出一条可保留EHP结构域的替代性加工通路，可生成非聚合型THP。我们制备了可特异性识别携带完整EHP结构域的THP（THP+EHP）的抗体，并证实尿液中存在非聚合状态的天然THP+EHP。对尿液源性THP+EHP的蛋白质组学表征显示，其C端终止于F617位点。在人体肾脏中，THP+EHP不仅可在髓袢升支粗段细胞中被检测到，还可弥散分布于肾实质内。通过C端蛋白质组测序与免疫印迹（immunoblotting）实验，我们进一步证实血清THP同样保留了EHP结构域。在一小队列存在急性肾损伤（acute kidney injury, AKI）风险的患者中，住院期间后续发生AKI的患者，其入院时尿液中的THP+EHP水平显著降低。本研究通过证实存在一条替代性细胞加工通路，揭示了尿调蛋白生物学的全新机制，该通路可解释循环中非聚合型THP的释放来源。未来需开展更大规模的研究，以确立尿液THP+EHP作为评估肾脏健康状态与损伤易感性的敏感生物标志物的应用价值。