Abstract
BACKGROUND: The renin-angiotensin system is highly conserved across vertebrates, including zebrafish, which possess orthologous genes coding for renin-angiotensin system proteins, and specialized mural cells of the kidney arterioles, capable of synthesising and secreting renin. METHODS: We generated zebrafish with CRISPR-Cas9-targeted knockout of renin (ren(-/-)) to investigate renin function in a low blood pressure environment. We used single-cell (10×) RNA sequencing analysis to compare the transcriptome profiles of renin lineage cells from mesonephric kidneys of ren(-/-) with ren(+/+) zebrafish and with the metanephric kidneys of Ren1(c-/-) and Ren1(c+/+) mice. RESULTS: The ren(-/-) larvae exhibited delays in larval growth, glomerular fusion and appearance of a swim bladder, but were viable and withstood low salinity during early larval stages. Optogenetic ablation of renin-expressing cells, located at the anterior mesenteric artery of 3-day-old larvae, caused a loss of tone, due to diminished contractility. The ren(-/-) mesonephric kidney exhibited vacuolated cells in the proximal tubule, which were also observed in Ren1(c-/-) mouse kidney. Fluorescent reporters for renin and smooth muscle actin (Tg(ren:LifeAct-RFP; acta2:EGFP)), revealed a dramatic recruitment of renin lineage cells along the renal vasculature of adult ren(-/-) fish, suggesting a continued requirement for renin, in the absence of detectable angiotensin metabolites, as seen in the Ren1YFP Ren1(c-/-) mouse. Both phenotypes were rescued by alleles lacking the potential for glycosylation at exon 2, suggesting that glycosylation is not essential for normal physiological function. CONCLUSIONS: Phenotypic similarities and transcriptional variations between mouse and zebrafish renin knockouts suggests evolution of renin cell function with terrestrial survival.