Fig. 3.

Live cell imaging reveals that some genes are required for recruitment of RLC-GFP to the equatorial cortex and others only for its retention there. An S2 cell line stably expressing both RLC-GFP (green) and mRFP-α-tubulin (red) was treated with double-stranded RNAs for genes that showed an effect on myosin II localization in fixed cells, and then epifluorescence microscopy was used to determine RLC-GFP localization in cells as they transitioned out of metaphase. Cells depleted of rho1 (Top) did not contract and showed no RLC-GFP localization or central spindle formation. RNAi of rho kinase, pebble, and the myosin II heavy chain showed similar phenotypes (data not shown). Cells depleted of diaphanous (Middle) contract after myosin II becomes localized to the equatorial cortex, but localization is transient, and because myosin II disperses into patches about the cortex, those regions of the cortex contract. Chickadee-depleted cells (Bottom) show a similar phenotype. (Scale bar: 5 μm.).