Does ROCK-dependant actomyosin activity kick-start nuclear descent during inter-kinetic nuclear migration in the mammalian neuroepithelium? (360G-Wellcome-211768_Z_18_Z)

In the posterior neuropore (PNP), actomyosin-driven apical constriction reduces the apical surface area of neuroepithelial cells, aiding neural fold apposition. This constriction is superimposed on changes in apical dimensions due to interkinetic nuclear migration (IKNM). Regional IKNM regulation is believed to alter PNP architecture, but the degree to which IKNM is linked to force generating apical constriction is unknown. The host lab recently showed that inhibiting the actomyosin-regulating Rho-associated protein kinase (ROCK) produces an atypical bimodal distribution of apical dimensions. This, alongside heterogeneous phospho-myosin light chain (pMLC)II staining, suggests only a subset of neuroepithelial cells are actively constricting at any one time. We hypothesise: ROCK-dependent active apical constriction at the end of mitosis initiates neuroepithelial nuclear descent in IKNM. To address this, I will use CD1 mouse embryos to: Compare apical dimensions and pMLCII immunofluorescent staining of neuroepithelial cells in M phase, G1, and intervening phases by confocal microscopy. Establish the time course of apical area changes and pMLCII staining following ROCK inhibitor treatment. Determine if apical area changes induced by ROCK inhibition is cell cycle phase dependent. Produce descriptive 3D in silico models of nuclear movements and apical dimensions for neuroepithelial cells with/without ROCK inhibition from confocal images.