LAUSR

The distal convoluted tubule (DCT) undergoes hypertrophy and hyperplasia to increase salt absorption in response to physiologic and pathological stresses, and has been implicated in the development of loop-diuretic resistance. However, the molecular mechanisms underlying DCT remodeling are unknown. We investigated progressive changes in the transcriptome of the early DCT (DCT1) in a time series (12, 24, 48, and 96 hours) after the administration of furosemide to identify initiating gene regulatory pathways. For high-resolution coverage, RNAseq was performed on isolated DCT1 segments, identified using a DCT1-specific Cre-driven fluorescent protein, tdTomato. An average of 17 million sequences were read per sample, providing genome-wide coverage of the DCT transcriptome (15499 genes). Sequence reads aligned on the mouse transcripts were quantified using Salmon, and differential gene expression analysis was performed using edgeR. All samples were enriched with known DCT-specific genes and lacked other nephron segment specific markers, indicating uniform purification of DCT tubules. Pathway and pattern clustering analysis of the time-resolved DCT transcriptome profiles revealed the early-induced genes were highly enriched in gene sets associated with cell cycle progression and cell proliferation. Meta-analysis of transcription factor binding enrichment at promoters of the early-induced genes revealed enrichment of genomic binding sites of Hypoxia-inducible factor 1 (HIF-1) signaling components (HIF1A, STAT3, RELA) across the early-induced gene promoters. Immunoblot analysis in kidney cortical tissues confirmed rapid activation of the HIF-1 signaling components following furosemide treatment. In summary, a core transcriptional regulatory axis in the DCT is activated rapidly after loop-diuretic treatment to drive a program of cell proliferation in the DCT. The results identify HIF1 signaling as a proximal pathway in the initiation of proliferative DCT remodeling. NIDDK, LeDucq Foundation This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.