patterning the primitive streak – Patterning

We aim to understand how the primitive streak (PS) is patterned in the mouse embryo. This transient structure plays a crucial role in the organization of the body plan. Classical embryology has established that cells emerging from it at different levels and times adopt different cell identities, but how this is achieved is still not understood. The BMP, WNT and NODAL signaling pathways are known to be involved, but altering precisely the level and duration of these signals to examine how it impacts cell-fate decisions is challenging in the mouse embryo. However, the development of an embryonic stem-cell-based in vitro model system recapitulating key aspects of gastrulation, and resulting in the formation of 2D gastruloids (2Dgas), has allowed us to investigate its underlying mechanisms. The results we have obtained so far are consistent with those previously obtained in the embryo, but are at odds with current theoretical models of PS patterning. In particular, we found that in addition to promoting proximal PS cell fates in m2Dgas, BMP actively suppresses distal ones.
We propose to address two fundamental questions: First, what are the BMP, WNT and NODAL signaling requirements for the correct allocation of cell-fates within the PS? Second, what are the molecular mechanisms underlying the patterning of the PS, and in particular, how is BMP signaling controlling the expression of NODAL and WNT signaling targets?
To carry out these investigations we recently designed a microfluidic chip in which pluripotent cells can be exposed to defined gradients of molecules for defined length of time. This allows us to explore in vitro the signaling parameters that ensure the patterning of a complete PS and the formation of all its derivatives. In combination with imaging, genome editing, single cell transcriptomics and epigenomics approaches, our in vitro models will give us the means to dissect the respective roles played by the BMP and NODAL signaling pathways in PS patterning. The results of our experiments will we processed and analyzed using a machine learning-based program, designed by one of us, to infer causality. These analyses will infer possible regulatory interactions playing critical roles in the PS cell identity decision process. The validity of these inferences will tested both in vitro and in vivo.
We expect that the completion of our project will lead to advances in our understanding of the signaling landscape that shapes the formation and patterning of the PS. We should also finally understand how, within the PS, BMP signaling redirects the action of NODAL and WNT signaling, and affects the process that underlies cell-fate decisions. Finally, we should make some significant advance in our understanding of how the grammar of TF binding sites at an enhancer translates into a specific pattern of activity. Completing PATTERNING will be an opportunity to identify general principles that will apply to other biological contexts, and may have applications in healthcare or bioengineering.