Dynamic myosin phosphorylation is required for pulsed contractions during apical constriction. Claudia G Vásquez, Adam C. Martin. Biology, Massachusetts Institute of Technology, Cambridge, MA.

   The formation of tissue layers, such as germ layers, during gastrulation, is critical for embryonic development. A cell shape change that generates tissue layers is the apical constriction of epithelial cells, which promotes bending and invagination of cells in an epithelial sheet. During Drosophila gastrulation, a band of 18x70 prospective mesoderm cells on the ventral midline of the embryo apically constrict, forming a ventral furrow. While it is known that pulses of non-muscle myosin II (Myo-II) accumulations contract an actin meshwork to apically constrict these cells, how Myo-II is dynamically regulated to generate force is not understood. One regulatory candidate is phosphorylation of the Myo-II regulatory light chain (RLC) at two conserved sites, Thr20 and Ser21. Phosphorylation of these sites not only directs Myo-II mini-filament assembly, but also activates contractile motor activity. Using RLC phospho-mutants that lock Myo-II into different activity states, we find that mutants that disrupt modulation of RLC phosphorylation inhibit Myo-II contractile pulses. We observed that phospho-mimetic Myo-II mutants continuously constrict cells, causing the ventral tissue to tear. Thus, the pulsed Myo-II contractions are a possible mechanism to attenuate tissue tension while cells apically constrict. In contrast mutants that block Myo-II phosphorylation struggle to effectively generate cell contractions, and cells become round and appear to lose adhesion to each other. Analysis of Myo-II phospho-mutants will provide insight into the possible benefits of ratchet-like cell constriction versus continuous cell constriction.