High-throughput micro-patterning platform reveals Nodal-dependent dissection of peri-gastrulation-associated versus pre-neurulation associated fate patterning

TY - JOUR

T1 - High-throughput micro-patterning platform reveals Nodal-dependent dissection of peri-gastrulation-associated versus pre-neurulation associated fate patterning

AU - Tewary, Mukul

AU - Dziedzicka, Dominika

AU - Ostblom, Joel

AU - Prochazka, Laura

AU - Shakiba, Nika

AU - Woodford, Curtis

AU - Piccinini, Elia

AU - Vickers, Alice

AU - Louis, Blaise

AU - Rahman, Nafees

AU - Danovi, Davide

AU - Geens, Mieke

AU - Watt, Fiona Mary

AU - Zandstra, Peter W

PY - 2018/11/7

Y1 - 2018/11/7

N2 - In vitro models of post-implantation human development are valuable to the fields of regenerative medicine, and developmental biology. Here, we report characterization of a robust in vitro platform that enabled high-throughput screening of multiple human pluripotent stem cell (hPSC) lines for their ability to undergo peri-gastrulation-like fate patterning upon BMP4 treatment of geometrically-confined colonies and observed significant variability. Further, we observed that when hPSCs were permitted to differentiate in conditions unsupportive of pluripotency, upregulation of endogenous Nodal correlated with expression of a gastrulation-associated gene profile, whereas Nodal downregulation correlated with a neurulation-associated gene profile expression. Given these observations, we hypothesized that inhibiting Nodal during a peri-gastrulation-like induction would induce fate patterning associated with the early stages of neurulation and observed experimental results consistent with this hypothesis. Mechanistically, we demonstrate a reaction-diffusion mediated self-organization of phosphorylated-SMAD1 signaling, and a positional-information mediated patterning of pre-neurulation-associated fates. Our work identifies a Nodal signaling dependent switch in peri-gastrulation versus pre-neurulation-associated fate patterning in hPSC colonies and hints towards possible conserved mechanisms of self-organized fate specification in differentiating epiblast and ectodermal tissues.

AB - In vitro models of post-implantation human development are valuable to the fields of regenerative medicine, and developmental biology. Here, we report characterization of a robust in vitro platform that enabled high-throughput screening of multiple human pluripotent stem cell (hPSC) lines for their ability to undergo peri-gastrulation-like fate patterning upon BMP4 treatment of geometrically-confined colonies and observed significant variability. Further, we observed that when hPSCs were permitted to differentiate in conditions unsupportive of pluripotency, upregulation of endogenous Nodal correlated with expression of a gastrulation-associated gene profile, whereas Nodal downregulation correlated with a neurulation-associated gene profile expression. Given these observations, we hypothesized that inhibiting Nodal during a peri-gastrulation-like induction would induce fate patterning associated with the early stages of neurulation and observed experimental results consistent with this hypothesis. Mechanistically, we demonstrate a reaction-diffusion mediated self-organization of phosphorylated-SMAD1 signaling, and a positional-information mediated patterning of pre-neurulation-associated fates. Our work identifies a Nodal signaling dependent switch in peri-gastrulation versus pre-neurulation-associated fate patterning in hPSC colonies and hints towards possible conserved mechanisms of self-organized fate specification in differentiating epiblast and ectodermal tissues.

U2 - 10.1101/465039

DO - 10.1101/465039

M3 - Article

SP - 465039

JO - bioRxiv

JF - bioRxiv

ER -