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Figure 1


Fig. 1. PLK-1 is distributed asymmetrically in early C. elegans embryos. (A) Schematic representation of C. elegans PLK-1. The kinase (KD) and polo boxes (PB1 and PB2) are indicated, as is the region against which the C-terminal antibodies were raised (amino acids 589-648; underlined in red). (B) Western blot of wild-type (lane 1) and strong plk-1(RNAi) (lane 2) embryonic extracts probed with PLK-1 antibodies, which recognize a specific band at the expected size of ~70 kDa. The blot was reprobed with {alpha}-tubulin antibodies as a loading control. (C-H) Wild-type (C-F), strong plk-1(RNAi) (G) or GFP-PLK-1 (H) embryos stained for PLK-1 (C-G) or GFP (H) (shown alone in the left panels and in red in the merged panels), {alpha}-tubulin (green) and DNA (blue). In all figures, anterior is towards the left and scale bars correspond to 10 µm. Line scans of PLK-1 intensity of C-F (yellow rectangles) are shown on the right, with 1.0 corresponding to the average pixel intensity within the rectangle. (C) Shortly after the completion of meiosis, PLK-1 distribution in the cytoplasm is uniform. (D) In early prophase, cytoplasmic PLK-1 becomes asymmetric, with more protein present in the anterior. (E,F) PLK-1 asymmetry is maintained in anaphase (E) and in the early two-cell stage (F). PLK-1 is enriched at centrosomes (E, arrowheads) and the midbody (E,F, arrows). The signal detected by PLK-1 antibodies is specific, as it is essentially absent in strong plk-1(RNAi) embryos (G), which nevertheless allowed passage through the meiotic divisions in this particular embryo. Finally, asymmetric distribution in two-cell stage embryos is also apparent with GFP-PLK-1 (H). The fact that asymmetry with this particular transgenic line is less pronounced than that of endogenous PLK-1 may reflect the fact that excess GFP-PLK-1 cannot be efficiently retained at the anterior in the presence of endogenous PLK-1. Compatible with this view, anterior enrichment of GFP-PLK-1 to an extent comparable to that of endogenous PLK-1 was observed with a distinct transgenic line (a gift from Asako Sugimoto) that expresses lower protein levels (data not shown).