8. Actin nucleation and elongation in the lamellipodium
Protein assemblies are recruited to the tips of lamellipodia and filopodia that regulate and drive the polymerisation of actin for protrusion.
For reviews see:
Stradal et al., 2004; Chesarone and B.L. Goode, 2009; Takenawa and Suetsugu, 2007; Derivery and Gautreau, 2010.
Live cell microscopy with GFP-tagged proteins has shown that members of this complex are specifically targeted to lamellipodia at the onset of protrusion (Fig. 8-1). (Links to pdf files of tip protein papers: VASP; Abi; Scar; IRS/WAVE)
| Figures 8-1 A-E: Some members of the tip complex in lamellipodia and filopodia. Video sequences show the lamellipodia region of mouse melanoma cells that were transfected with the proteins indicated, tagged with GFP. Click on pictures to watch movies A: VASP (Movie 2.67Mb) B: Abi-1 (Movie 794Kb) C: Scar WAVE1 (Movie 4Mb) D: Scar WAVE2 (Movie 1.64Mb) and E: IRSp53. (Movie 816 Kb) |
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The WAVE complex comprises in mammals Scar/WAVE and Abi, Sra1/PIR121, Nap1 (or haematopoietic Hem1) and Brick1/HSPC300. The role of WAVE is to activate the Arp2/3 complex to initiate actin branching for the generation of actin networks in lamellipodia. Actin branching can be reproduced in vitro by mixing actin with an active domain of the WAVE or WASP protein and the Arp2/3 complex (Amann and Pollard, 2002) (Fig. 8-2).
Figure 8-2. Branching of actin in vitro by WAVE and the Arp2/3 complex observed using TIRF microscopy. Courtesy of Kai Schlueter and Theresia Stradal (unpublished).
As shown by the Welch group, the Arp2/3 complex is localized over the whole lamellipodium network. Photobleach experiments show that the Aprp2/3 complex treadmills with actin in the lamellipodium (Fig. 8-3)
Figure 8-3. Fluorescence recovery after photobleaching (FRAP) in the lamellipodium of a B16 melanoma cell expressing Actin-RFP and ArpC5-GFP. Note that the recovery of the Arp label parallels the recovery of actin (Lai et al., 2008). As shown by the Svitkina group, the Arp complex is absent from the core of actin bundles (microspikes) that transverse the lamellipodium.
Video courtesy of Malgozata Szcozodrak and Klemens Rottner (unpublished).
VASP is a member of the Ena/VASP family of proteins that act as processive elongators of actin filaments (Breitsprecher et al., 2008, 2010), as shown in Fig 8-4 and 8-5.
Figure 8-4. Plastic beads coated with VASP promote the growth of actin filaments at the bead surface by tethering actin filaments and feeding monomers into the plus ends. From Breitsprecher et al., 2008.
(Movie 2.3Mb) Click on picture to watch movie
Figure 8-5. Shows again the localization of VASP at protruding lamellipodia and filopodia tips in a fish fibroblast expressing VASP-GFP and Actin-RFP.
(Movie 4.8Mb) Click on picture to watch movie
