Figure 1. Using a 405 nm laser to wound the worm epidermis reveals rapid membrane reorganisation.

(A) Schematic view of a section through an adult C. elegans worm near the mid-body. The internal organs are omitted for the sake of clarity. The main syncytial epidermis, hyp7 (buff), with its adjacent cells (seam cells in pink, nerve cord in red and muscles in green) can be divided into two main regions, the lateral epidermis (delimited by the dashed black line) and the thin dorso-ventral epidermis where muscles are anchored to the cuticle. Microtubules (thin taupe lines) are disordered on the apical surface of the lateral epidermis. The typical position of a wound is indicated by the yellow circle. Figure adapted from one kindly provided by Christopher Crocker, WormAtlas (Altun and Hall, 2014). (B) Wounding the lateral epidermis with a 405 nm laser causes a rapid increase in intracellular Ca²⁺, measured using GCamp3. Representative spinning disk images from a worm carrying a col-19p::GCamp3 reporter transgene. The dashed circle is centred on the wound site; bw, before wound; pw, post-wound; sc, seam cells. Scale bar 10 µm. (C–D) Upon laser wounding, membrane and lipids (red arrows) are rapidly recruited to the wound site (asterisk). Representative spinning disk images of worms carrying dpy-7p::GFP::CAAX (C) and wrt-2p::GFP::PH-PLC1δ transgenes (D). bw, before wound; time post-wound [min:s]; sc, seam cells. Scale bar 5 µm. The last panel on the right in both C and D is a focal plane two microns deeper than the previous images, showing recruitment of cytoplasmic vesicles to the wound.

Figure 1—figure supplement 1. Reproduction of known wound hallmarks following injury using a 405 nm laser.

Wounding C. elegans lateral epidermis with a 405 nm laser causes immediate formation of an autofluorescent scar (A) and later an actin ring, revealed with the actin-binding protein GFP::moesin (B) at the wound site. Representative images of wild-type (A) and col-19p::GFP::moesin (B) worms. The dashed circle is centred on the wound site; bw, before wound; pw, post wound; sc, seam cells. Scale bar 10 µm except in the inset 5 µm. (C) 3 hr after wounding, young adult worms exhibit an increased expression of a nlp-29p::GFP reporter, a read out of the immune response (right), compared to unwounded worms (left). The worms express DsRed constitutively in the epidermis. Red and green fluorescence are visualized here simultaneously.

Figure 1—video 1. Plasma membrane rapidly reorganizes at the wound site.

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Imaging Ex dpy-7p::GFP::CAAX; * indicates the wound site; [min:sec]. Scale bar 5 µm.

Figure 1—video 2. Rapid PIP₂ domain reorganization at the wound site.

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Imaging Is wrt-2p::GFP::PH-PLC1δ; * indicates the wound site; [min:sec]. At resting conditions, PIP₂ membrane domains are large and static. Upon wounding, the GFP signal first disappears in a large area around the wound (15 s), it then re-appears and the domains close to the wound reorganise. Scale bar 5 µm.