Scott Hoffmann1, Linda Mullins1, Sebastien Rider1,2, Cara Brown1, Charlotte B Buckley1,3, Adrienne Assmus1, Ziwen Li1, Mariana Sierra Beltran4, Neil Henderson4,5, Jorge Del Pozo6, Alexandre De Goes Martini7, Maria Luisa S Sequeira-Lopez7, R Ariel Gomez7, John Mullins1. 1. Centre for Cardiovascular Science (S.H., L.M., S.R., C.B., C.B.B., A.A., Z.L., J.M.), The Queen's Medical Research Institute, The University of Edinburgh, United Kingdom. 2. Now with DSM Nutritional Products Ltd, Switzerland (S.R.). 3. Now with Strathclyde Institute of Pharmacy and Biomedical Sciences, University of Strathclyde, Glasgow, United Kingdom (C.B.B.). 4. Centre for Inflammation Research (M.S.B., N.H.), The Queen's Medical Research Institute, The University of Edinburgh, United Kingdom. 5. MRC Human Genetics Unit, Institute of Genetics and Molecular Medicine, University of Edinburgh, United Kingdom (N.H.). 6. Veterinary Pathology, Royal (Dick)School of Veterinary Studies and The Roslin Institute, The University of Edinburgh, Easter Bush Campus, United Kingdom (J.d.P.). 7. Department of Pediatrics, School of Medicine, University of Virginia, Charlottesville (A.D.G.M., M.L.S.S.-L., R.A.G.).
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 Ren1c-/- and Ren1c+/+ 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 Ren1c-/- 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 Ren1c-/- 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.
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 Ren1c-/- and Ren1c+/+ 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 Ren1c-/- 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 Ren1c-/- 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.
Authors: Cuong Q Diep; Dongdong Ma; Rahul C Deo; Teresa M Holm; Richard W Naylor; Natasha Arora; Rebecca A Wingert; Frank Bollig; Gordana Djordjevic; Benjamin Lichman; Hao Zhu; Takanori Ikenaga; Fumihito Ono; Christoph Englert; Chad A Cowan; Neil A Hukriede; Robert I Handin; Alan J Davidson Journal: Nature Date: 2011-01-26 Impact factor: 49.962
Authors: Johannes Schindelin; Ignacio Arganda-Carreras; Erwin Frise; Verena Kaynig; Mark Longair; Tobias Pietzsch; Stephan Preibisch; Curtis Rueden; Stephan Saalfeld; Benjamin Schmid; Jean-Yves Tinevez; Daniel James White; Volker Hartenstein; Kevin Eliceiri; Pavel Tomancak; Albert Cardona Journal: Nat Methods Date: 2012-06-28 Impact factor: 28.547
Authors: R Ariel Gomez; Brian Belyea; Silvia Medrano; Ellen S Pentz; Maria Luisa S Sequeira-Lopez Journal: Pediatr Nephrol Date: 2013-12-15 Impact factor: 3.714
Authors: Ellen Steward Pentz; Maria Alejandra Moyano; Barbara A Thornhill; Maria Luisa S Sequeira Lopez; R Ariel Gomez Journal: Am J Physiol Regul Integr Comp Physiol Date: 2003-10-16 Impact factor: 3.619