A lay summary of this work is now available: hdbi.org/human-epibla...

Special thanks once again to Kathy @niakanlab.bsky.social and Richard Acton at @babrahaminst.bsky.social

Special shout-out to Michele Marass, our outstanding steward through the turbulent waters that is the review process.

I’m grateful for the continued support from
the Niakan Lab, and our collaborators
in the labs of L. Vallier, P. Rugg-Gunn, and C. Hill, who made this work possible. 9/10

These data suggest differences in the signalling requirements regulating pluripotency in the pre-implantation human epiblast compared with existing human embryonic stem cell culture. 8/10

Transcriptional analysis of A8301-treated human blastocysts indicated that NANOG transcription did not decrease under NODAL signalling inhibition, in contrast to naïve human embryonic stem cells in t2iLifGo conditions pubmed.ncbi.nlm.nih.gov/34463252/ 7/10

A8301 treatment throughout human blastocyst development from 5 to 7 days post fertilisation did not impact the number of cells expressing epiblast markers NANOG, SOX2, KLF17 or primitive endoderm marker GATA4. 6/10

We next characterised three NODAL signalling inhibitors, yet only A8301 reliably inhibited SMAD2/3 phosphorylation and nuclear accumulation. The required high concentration of A8301 also inhibited BMP signalling, found in the primitive endoderm of human blastocysts. 5/10

We found that NANOG expression emerges in the inner cell mass of human blastocysts at 5 days post fertilisation, in the absence of NODAL signalling activity. NODAL signalling then becomes active throughout the pluripotent epiblast and extra-embryonic primitive endoderm. 4/10

To characterise NODAL signalling activity, we optimised an immunofluorescence protocol to detect the intracellular effectors SMAD2/3, which become phosphorylated and accumulate in the nucleus upon ligand binding. See also Kruger et al doi.org/10.1242/dev.... 3/10

In vitro, NODAL regulates pluripotency, by promoting NANOG expression, as shown by Ludovic Vallier, AnnaOsnato and others. We thus wondered whether NODAL signalling activity was active in pre-implantation human development and involved in human pluripotency in vivo. 2/10

In my PhD work, published last Monday, we show that both NODAL and BMP signalling are active in the human blastocyst, but NODAL signalling is not required to initiate or maintain the human pluripotent epiblast. 1/10

Thank you so much @cellysally.bsky.social - great slogan!
I'll cross-post my Twitter thread below.