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Redefining the in vivo origin of nephron progenitors enables generation ofthree-dimensional kidney structures from pluripotent stem cells in vitro＊Authors’ name and affiliation: 11pt; Underline the lead presenter＊Title: Size=14pt; Style=BoldAtsuhiro Taguchi and Ryuichi Nishinakamura
Institute of Molecular Embryology and Genetics, Kumamoto University, Kumamoto, Japan＊Abstract: Not more than 300words; 11pt; Alignment=Justified; Do not insert fig., or any other graphics

Recapitulating three-dimensional structures of the kidney in vitro is a major challenge for developmental
biology and regenerative medicine. Adult kidney derives from embryonic metanephros, which develops by
the reciprocal interaction between the metanephric mesenchyme and the ureteric bud. Here, we define thedevelopmental origins of the nephron progenitors, which reside in the metanephric mesenchyme andgenerate most nephron components such as glomeruli and renal tubules. While nephron progenitors arebelieved to originate from the intermediate mesoderm that expresses a transcription factor Osr1, we
unexpectedly find that nephron progenitors are derived from posteriorly located T (Brachyury)-positive
population at the post-gastrulation stage, which is developmentally distinct from Osr1-positive ureteric bud
precursors. We also identify phasic Wnt stimulation and stage-specific growth factor addition as molecular
cues that promote the development of T-positive precursors into the nephron progenitors. We then use thisinformation to derive nephron progenitors, via the newly identified T-positive precursors, from mouseembryonic stem cells and human induced pluripotent stem cells. Upon Wnt4 stimulation, the induced
nephron progenitors readily reconstitute the three-dimensional structures of the kidney in vitro, including
glomeruli with podocytes and renal tubules with clear lumina. Furthermore, mouse glomeruli are efficiently
vascularized upon transplantation, because glomerular podocytes express vasculogenic factors including
VEGF. Thus, by redefining the developmental origin of nephron progenitors, we have revealed themolecular cascades of kidney specification in vivo and succeeded in generating the three-dimensionalnephrons in vitro from pluripotent stem cells both in mice and humans.

References＊References：11pt; Line-spacing=Minimum 0pt

1) Taguchi A, Kaku Y, Ohmori T, Sharmin S, Ogawa M, Sasaki H, and Nishinakamura R. Redefining the invivo origin of metanephric nephron progenitors enables generation of complex kidney structures frompluripotent stem cells. Cell Stem Cell 14: 53-67, 2014.
2) Fujimoto Y, Tanaka SS, Yamaguchi YL, Kobayashi H, Kuroki S, Tachibana M, Shinomura M, Kanai Y,Morohashi K, Kawakami K, and Nishinakamura R. Homeoprotein Six1 and Six4 regulate male sex
determination and mouse gonadal development. Dev. Cell 26: 416-430, 2013.
3) Sakaguchi M, Sharmin S, Taguchi A, Ohmori T, Fujimura S, Abe T, Kiyonari H, Komatsu Y, Mishina Y,Asashima M, Araki E, and Nishinakamura R. The phosphatase Dullard negatively regulates BMP signallingand is essential for nephron maintenance after birth. Nat. Commun. 4: 1398, 2013.