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Fig. 4. Dynamics of GFP–LIS-1 localization. (A,B) Spinning-disc confocal time-lapse fluorescence microscopy of one-cell-stage (A) and four-cell-stage (B) embryos derived from homozygous lis-1(1550) adults expressing GFP–LIS-1 (see also corresponding Movies 9-10, in supplementary material, http://jcs.biologists.org/cgi/content/full/117/19/4571/DC1). Time elapsed since the beginning of each movie is shown in minutes and seconds; live embryos in this and all other figures are 50 µm long. Note enrichment at nuclear periphery during prophase (A, time 01:55 and 03:50; B, time 00:00 and 02:15), entry in pronuclei/nuclei during late prophase (A, time 03:50; B, time 02:15), as well as further enrichment in the vicinity of chromosomes during prometaphase (A, time 05:40; not visible in B due to orientation of spindle in ABa and ABp). (C) Quantification of average pixel intensities in the nucleus and a comparable area in the cytoplasm of the four-cell-stage embryo displayed in panel B; results are shown for the time interval between the moment GFP–LIS-1 enrichment becomes apparent at the nuclear periphery and NEBD. Similar plots were obtained in three other embryos. (D,E) FRAP experiments. Images from representative confocal time-lapse sequences of four-cell-stage embryos derived from homozygous lis-1(1550) adults expressing GFP–LIS-1 (D) or wild-type animals injected with FITC-labelled Dextran 70 kDa (D) (see also corresponding Movies 11 and 12, in supplementary material). Time elapsed is shown in minutes and seconds. Photobleaching of a 4.5 µm² square (indicated in white) started at time –00:02 and ended at time 00:00. Note very rapid recovery (t_1/2 10 seconds) of fluorescence for both embryos.

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