Novel CUL3 Variant Causing Familial Hyperkalemic Hypertension Impairs Regulation and Function of Ubiquitin Ligase Activity.
Novel CUL3 Variant Causing Familial Hyperkalemic Hypertension Impairs Regulation and Function of Ubiquitin Ligase Activity.
复制标题
DOI：
10.1161/hypertensionaha.121.17624
复制
发表时间：
2022-01
期刊：
影响因子：
--
通讯作者：
Sharma P
中科院分区：
文献类型：
--
作者：
Chatrathi HE;Collins JC;Wolfe LA;Markello TC;Adams DR;Gahl WA;Werner A;Sharma P
家族性高钾血症性高血压（FHHt）是由CUL 3-KLHL 3-WNK通路的致病性基因变异引起的，临床表现为高钾血症、代谢性酸中毒和高收缩压。泛素E3连接酶CUL 3-KLHL 3靶向WNK激酶进行降解，以限制噻嗪敏感性Na-Cl协同转运蛋白（NCC）的激活。CUL 3中所有已知的变体导致外显子9跳跃（CUL 3 Δ9），并且通常导致患者的严重FHHt和生长障碍。CUL 3中的其他变体是否导致FHHt尚不清楚。在此，我们在一名患有多种先天性异常的儿童FHHt患者中鉴定了一种新的新生杂合CUL 3变体（CUL 3 Δ474-477），并揭示了CUL 3 Δ474-477导致CUL 3-KLHL 3-WNK信号轴失调的分子机制。使用患者来源的尿细胞外囊泡（uEV）和真皮成纤维细胞、体外测定和培养的肾细胞，我们证明CUL 3 Δ474-477由于增加的自身泛素化而导致总CUL 3水平降低。逃脱自动降解的CUL 3 Δ474-477显示出用NEDD 8增强的修饰和在泛素化WNK 4中受损的CUL 3-KLHL 3复合物的形成增加。CUL 3复合物的蛋白质组学分析显示，除了增加的KLHL 3结合之外，CUL 3 Δ474-477变体还表现出与其他BTB底物衔接子的相互作用增加，为患者的多样性表型提供了理论基础。我们得出结论，CUL 3 Δ474-477的病理生理学效应是由CUL 3水平降低和催化受损的CUL 3连接酶复合物的形成引起的。
Familial hyperkalemic hypertension (FHHt) is caused by pathogenic variants in genes of the CUL3-KLHL3-WNK pathway, manifesting clinically as hyperkalemia, metabolic acidosis, and high systolic blood pressure. The ubiquitin E3 ligase CUL3-KLHL3 targets WNK kinases for degradation to limit activation of the thiazide-sensitive Na-Cl cotransporter (NCC). All known variants in CUL3 lead to exon 9 skipping (CUL3Δ9) and typically result in severe FHHt and growth disturbances in patients. Whether other variants in CUL3 cause FHHt is unknown. Here we identify a novel de novo heterozygous CUL3 variant (CUL3Δ474–477) in a pediatric FHHt patient with multiple congenital anomalies and reveal molecular mechanisms by which the CUL3Δ474–477 leads to dysregulation of CUL3-KLHL3-WNK signaling axis. Using patient-derived urinary extracellular vesicles (uEVs) and dermal fibroblasts, in vitro assays, and cultured kidney cells, we demonstrate that CUL3Δ474–477 causes reduced total CUL3 levels due to increased auto-ubiquitination. The CUL3Δ474–477 that escapes auto-degradation shows enhanced modification with NEDD8 and increased formation of CUL3-KLHL3 complexes that are impaired in ubiquitinating WNK4. Proteomic analysis of CUL3 complexes revealed that, in addition to increased KLHL3 binding, the CUL3Δ474–477 variant also exhibits increased interactions with other BTB substrate adaptors, providing a rationale for the patient’s diverse phenotypes. We conclude that the pathophysiological effects of CUL3Δ474–477 are caused by reduced CUL3 levels and formation of catalytically impaired CUL3 ligase complexes.