Supplementary Figure 3: Effect of temperature and dilution on GLH-1, LAF-1 and DEPS-1.

a Time-lapse photomicrographs of a 2-cell embryos expressing GLH-1::GFP (n = 3), LAF-1::GFP (n = 4), and DEPS1::GFP (n = 5) cultured at 20 °C and then up-shifted to 34 °C for 1 minute. GLH-1 disperses whereas LAF-1 remains in granules throughout. DEPS-1 exists in two granule types: small, bright granules that are distributed throughout the cytoplasm and larger, less bright granules in the posterior cytoplasm. Both become reduced in intensity at 34 °C, consistent with partial dispersal. Images are representative of 3 independent experiments. Scale bar is 10 μm. b Time-lapse photomicrographs of 2-cell embryos expressing GLH-1::GFP, LAF-1::GFP, and DEPS1::GFP before and 10 s after laser puncture of the eggshell. Laser puncture causes the contents of the posterior blastomere to spill onto the slide mixing with the aqueous buffer. GLH-1 and LAF-1 immediately disperse. The majority of DEPS-1 also becomes dispersed, but a subset of smaller granules persists in the granule phase. Scale bar is 10 μm. c Graph showing the fraction of GLH-1::GFP (n = 6), LAF-1::GFP (n = 3), and DEPS1::GFP (n = 6) retained in the granular phase after extrusion. Total GFP fluorescence from granules was measured before laser puncture (I_B) and after laser puncture (I_A) and used to calculate a fluorescence ratio (I_A/I_B). Means are indicated along with error bars representing ± SD.