Laser ablation

Mechanical disruption of the actomyosin ring within the YSL was performed as previously described by conducting laser ablation on a UV laser ablation setup (Behrndt et al., 2012) equipped with a Zeiss 63 × 1.2 NA water immersion lens using Tg(actb1::myl12.1-eGFP) embryos. Embryos were mounted at 50% epiboly (5.3 hpf) and the YSL actomyosin cortex close to the EVL margin was repeatedly ablated by applying 10 UV pulses at 1000 Hz on a rectangular ROI. Suboptimal ablation intensity was applied to just disrupt the cortical actomyosin flux and marginal ring, and avoid the activation of a wound response within the yolk cell. The kinetic of EVL cells, DFCs, and YSL marginal actomyosin network adjacent to the disrupted cortex was compared with the kinetic of close neighbouring tissues showing intact regions of the actomyosin network as an internal control. Cortical laser ablation of the EVL was performed parallel to the EVL margin and perpendicular to the EVL actomyosin cortex by applying 25 UV pulses at 1000  Hz along a 10 μm line. Retraction of apical ties and DFCs were quantified from maximum z-projections images.