Himmerkus Nina, Svendsen Samuel L, Quintanova Catarina, Bleich Markus, Von Schwerdtner Otto, Benzing Thomas, Welling Paul A, Leipziger Jens, Rinschen Markus M
Institute of Physiology, Christian Albrechts University Kiel, Kiel, Germany.
Department of Biomedicine, Aarhus University, Aarhus, Denmark.
Function (Oxf). 2020 Jul 2;1(1):zqaa007. doi: 10.1093/function/zqaa007. eCollection 2020.
The revolution of the omics technologies has enabled profiling of the molecules of any sample. However, the heterogeneity of the kidney with highly specialized nephron segments like the cortical collecting duct (CCD) poses a challenge regarding integration of omics data and functional analysis. We examined function and proteome from the same single CCDs of C57Bl6 mice by investigating them in a double-barreled perfusion system before targeted mass spectrometry. Transepithelial voltage (V), transepithelial resistance, as well as amiloride-sensitive voltage (ΔVamil) were recorded. CCDs were of 400-600 µm of length, showed lumen negative V between -8.5 and -32.5 mV and an equivalent short circuit current I' between 54 and 192 µA/cm. On a single-tubule proteome level, intercalated cell (IC) markers strongly correlated with other intercalated cell markers and negatively with principal cell markers. Integration of proteome data with phenotype data revealed that tubular length correlated with actin and Na-K-ATPase expression. ΔV(amil) reflected the expression level of the β-subunit of the epithelial sodium channel. Intriguingly, ΔV(amil) correlated inversely with the water channel AQP2 and the negative regulator protein NEDD4L (NEDD4-2). In pendrin knockout (KO) mice, the CCD proteome was accompanied by strong downregulation of other IC markers like CLCNKB, BSND (Barttin), and VAA (vH-ATPase), a configuration that may contribute to the salt-losing phenotype of Pendred syndrome. Proteins normally coexpressed with pendrin were decreased in pendrin KO CCDs. In conclusion, we show that functional proteomics on a single nephron segment scale allows function-proteome correlations, and may potentially help predicting function from omics data.
组学技术的革命使得对任何样本中的分子进行分析成为可能。然而，肾脏具有高度特化的肾单位节段，如皮质集合管（CCD），其异质性给组学数据整合和功能分析带来了挑战。我们通过在靶向质谱分析之前，在双管灌注系统中研究C57Bl6小鼠的单个CCD，来检测其功能和蛋白质组。记录跨上皮电压（V）、跨上皮电阻以及氨氯地平敏感电压（ΔVamil）。CCD长度为400 - 600 µm，管腔负电压V在-8.5至-32.5 mV之间，等效短路电流I'在54至192 µA/cm之间。在单肾小管蛋白质组水平上，闰细胞（IC）标志物与其他闰细胞标志物高度相关，与主细胞标志物呈负相关。蛋白质组数据与表型数据的整合显示，肾小管长度与肌动蛋白和钠钾ATP酶的表达相关。ΔV(amil)反映了上皮钠通道β亚基的表达水平。有趣的是，ΔV(amil)与水通道蛋白AQP2和负调节蛋白NEDD4L（NEDD4 - 2）呈负相关。在pendrin基因敲除（KO）小鼠中，CCD蛋白质组伴随着其他IC标志物如CLCNKB、BSND（Barttin）和VAA（vH - ATP酶）的强烈下调，这种情况可能导致了 Pendred综合征的失盐表型。在pendrin KO的CCD中，通常与pendrin共表达的蛋白质减少。总之，我们表明在单个肾单位节段水平上的功能蛋白质组学能够实现功能与蛋白质组的相关性分析，并且可能有助于从组学数据预测功能。
