Significance Statement High-resolution single-nucleus RNA-sequencing data indicate a clear separation between primary sites of calcium and magnesium handling within distal convoluted tubule (DCT). Both DCT1 and DCT2 express Slc12a3, but these subsegments serve distinctive functions, with more abundant magnesium-handling genes along DCT1 and more calcium-handling genes along DCT2. The data also provide insight into the plasticity of the distal nephron-collecting duct junction, formed from cells of separate embryonic origins. By focusing/changing gradients of gene expression, the DCT can morph into different physiological cell states on demand. Background The distal convoluted tubule (DCT) comprises two subsegments, DCT1 and DCT2, with different functional and molecular characteristics. The functional and molecular distinction between these segments, however, has been controversial. Methods To understand the heterogeneity within the DCT population with better clarity, we enriched for DCT nuclei by using a mouse line combining “Isolation of Nuclei Tagged in specific Cell Types” and sodium chloride cotransporter–driven inducible Cre recombinase. We sorted the fluorescently labeled DCT nuclei using Fluorescence-Activated Nucleus Sorting and performed single-nucleus transcriptomics. Results Among 25,183 DCT cells, 75% were from DCT1 and 25% were from DCT2. In addition, there was a small population (<1%) enriched in proliferation-related genes, such as Top2a, Cenpp, and Mki67. Although both DCT1 and DCT2 expressed sodium chloride cotransporter, magnesium transport genes were predominantly expressed along DCT1, whereas calcium, electrogenic sodium, and potassium transport genes were more abundant along DCT2. The transition between these two segments was gradual, with a transitional zone in which DCT1 and DCT2 cells were interspersed. The expression of the homeobox genes by DCT cells suggests that they develop along different trajectories. Conclusions Transcriptomic analysis of an enriched rare cell population using a genetically targeted approach clarifies the function and classification of distal cells. The DCT segment is short, can be separated into two subsegments that serve distinct functions, and is speculated to derive from different origins during development.

中文翻译：

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富集单核 RNA 测序揭示远端卷管细胞的独特属性

意义陈述 高分辨率单核 RNA 测序数据表明远端曲管 (DCT) 内钙和镁处理的主要位点之间存在明显分离。 DCT1和DCT2都表达Slc12a3，但这些亚片段具有独特的功能，DCT1上有更丰富的镁处理基因，DCT2上有更多的钙处理基因。这些数据还提供了对由不同胚胎起源的细胞形成的远端肾单位-集合管连接处的可塑性的深入了解。通过聚焦/改变基因表达梯度，DCT 可以根据需要转变为不同的生理细胞状态。背景 远曲小管 (DCT) 包括两个亚段，DCT1 和 DCT2，具有不同的功能和分子特征。然而，这些片段之间的功能和分子区别一直存在争议。方法 为了更清楚地了解 DCT 群体内的异质性，我们通过使用结合“特定细胞类型中标记的细胞核分离”和氯化钠协同转运蛋白驱动的诱导型 Cre 重组酶的小鼠品系来富集 DCT 细胞核。我们使用荧光激活核分选对荧光标记的 DCT 核进行分选，并进行单核转录组学。结果 25,183 个 DCT 细胞中，75% 来自 DCT1，25% 来自 DCT2。此外，还有一小部分群体（<1%）富含增殖相关基因，例如 Top2a、Cenpp 和 Mki67。尽管 DCT1 和 DCT2 均表达氯化钠协同转运蛋白，但镁转运基因主要沿 DCT1 表达，而钙、电钠和钾转运基因沿 DCT2 表达更为丰富。这两个片段之间的过渡是渐进的，有一个过渡区，其中散布有 DCT1 和 DCT2 细胞。 DCT细胞的同源框基因表达表明它们沿着不同的轨迹发育。结论 使用基因靶向方法对富集的稀有细胞群进行转录组分析，阐明了远端细胞的功能和分类。 DCT 段很短，可以分为两个具有不同功能的子段，并且推测在开发过程中源自不同的起源。