Marta Vaquero1, Sara Cuesta1, Carlos Anerillas1, Gisela Altés1, Joan Ribera1, M Albert Basson2, Jonathan D Licht3, Joaquim Egea4, Mario Encinas5. 1. Departments of Experimental Medicine and. 2. Centre for Craniofacial and Regenerative Biology, King's College London, Guy's Hospital, London, UK; and. 3. The University of Florida Health Cancer Center, The University of Florida Cancer/Genetics Research Complex, Gainesville, Florida. 4. Basic Medical Sciences, Universitat de Lleida/Institut de Recerca Biomèdica de Lleida, Lleida, Spain. 5. Departments of Experimental Medicine and mario.encinas@mex.udl.cat.
Abstract
BACKGROUND: Studies in mice suggest that perturbations of the GDNF-Ret signaling pathway are a major genetic cause of congenital anomalies of the kidney and urinary tract (CAKUT). Mutations in Sprouty1, an intracellular Ret inhibitor, results in supernumerary kidneys, megaureters, and hydronephrosis in mice. But the underlying molecular mechanisms involved and which structural domains are essential for Sprouty1 function are a matter of controversy, partly because studies have so far relied on ectopic overexpression of the gene in cell lines. A conserved N-terminal tyrosine has been frequently, but not always, identified as critical for the function of Sprouty1 in vitro. METHODS: We generated Sprouty1 knockin mice bearing a tyrosine-to-alanine substitution in position 53, corresponding to the conserved N-terminal tyrosine of Sprouty1. We characterized the development of the genitourinary systems in these mice via different methods, including the use of reporter mice expressing EGFP from the Ret locus, and whole-mount cytokeratin staining. RESULTS: Mice lacking this tyrosine grow ectopic ureteric buds that will ultimately form supernumerary kidneys, a phenotype indistinguishable to that of Sprouty1 knockout mice. Sprouty1 knockin mice also present megaureters and vesicoureteral reflux, caused by failure of ureters to separate from Wolffian ducts and migrate to their definitive position. CONCLUSIONS: Tyrosine 53 is absolutely necessary for Sprouty1 function during genitourinary development in mice.
BACKGROUND: Studies in mice suggest that perturbations of the GDNF-Ret signaling pathway are a major genetic cause of congenital anomalies of the kidney and urinary tract (CAKUT). Mutations in Sprouty1, an intracellular Ret inhibitor, results in supernumerary kidneys, megaureters, and hydronephrosis in mice. But the underlying molecular mechanisms involved and which structural domains are essential for Sprouty1 function are a matter of controversy, partly because studies have so far relied on ectopic overexpression of the gene in cell lines. A conserved N-terminal tyrosine has been frequently, but not always, identified as critical for the function of Sprouty1 in vitro. METHODS: We generated Sprouty1 knockin mice bearing a tyrosine-to-alanine substitution in position 53, corresponding to the conserved N-terminal tyrosine of Sprouty1. We characterized the development of the genitourinary systems in these mice via different methods, including the use of reporter mice expressing EGFP from the Ret locus, and whole-mount cytokeratin staining. RESULTS:Mice lacking this tyrosine grow ectopic ureteric buds that will ultimately form supernumerary kidneys, a phenotype indistinguishable to that of Sprouty1 knockout mice. Sprouty1 knockin mice also present megaureters and vesicoureteral reflux, caused by failure of ureters to separate from Wolffian ducts and migrate to their definitive position. CONCLUSIONS:Tyrosine 53 is absolutely necessary for Sprouty1 function during genitourinary development in mice.
Authors: Gisela Altés; Marta Vaquero; Sara Cuesta; Carlos Anerillas; Anna Macià; Carme Espinet; Joan Ribera; Saverio Bellusci; Ophir D Klein; Andree Yeramian; Xavi Dolcet; Joaquim Egea; Mario Encinas Journal: Cell Mol Life Sci Date: 2022-09-13 Impact factor: 9.207