Solid mechanobiology hypothesis with clean predictions. The YAP/TAZ-mesodermal bias story is well-grounded, but a few things worth pressure-testing: The ligand density confound is your biggest enemy here. Stiffer hydrogels almost invariably present more available RGD or fibronectin binding sites at equivalent coating concentrations, so you may be measuring integrin clustering effects rather than stiffness per se. Constant ligand density across your stiffness gradient (via PEG-based systems with defined peptide conjugation) is table stakes, but worth stating explicitly as a design constraint rather than just a limitation. On the predictions — the YAP/TAZ nuclear localization → mesoderm marker correlation is mechanistically plausible, but YAP/TAZ have context-dependent transcriptional partners. Are you expecting direct transcriptional activation of T/Brachyury downstream of TEAD, or an indirect effect through BMP/Wnt pathway amplification? Distinguishing these matters for interpreting the FAK inhibitor rescue experiment, because FAK sits upstream of multiple pathways beyond YAP. The scRNA-seq readout is the right call. One suggestion: include trajectory analysis (RNA velocity or pseudotime) rather than just endpoint marker quantification — early mechanosensitive bifurcation points might be transient and missed at fixed timepoints. Also worth considering: are your organoids spatially patterned or homogeneous? If patterned, stiffness gradients within the organoid itself may create competing signals that confound bulk interpretation. What stiffness range are you targeting? The mesoderm/neuroectoderm transition may have a relatively sharp mechanical threshold rather than a graded response.