A llama-derived gelsolin single-domain antibody blocks gelsolin-G-actin interaction

作者：Van den Abbeele Anske; De Clercq Sarah; De Ganck Ariane; De Corte Veerle; Van Loo Berlinda; Soror Sameh Hamdy; Srinivasan Vasundara; Steyaert Jan; Vandekerckhove Joel; Gettemans Jan^*

来源：Cellular and Molecular Life Sciences, 2010, 67(9): 1519-1535.

DOI：10.1007/s00018-010-0266-1

• VHH
• Single-domain antibody
• Gelsolin
• Calcium
• Immunomodulation
• Cytoskeleton
• Intrabody
• Cancer cell

RNA interference has tremendously advanced our understanding of gene function but recent reports have exposed undesirable side-effects. Recombinant Camelid single-domain antibodies (VHHs) provide an attractive means for studying protein function without affecting gene expression. We raised VHHs against gelsolin (GsnVHHs), a multifunctional actin-binding protein that controls cellular actin organization and migration. GsnVHH-induced delocalization of gelsolin to mitochondria or the nucleus in mammalian cells reveals distinct subpopulations including free gelsolin and actin-bound gelsolin complexes. GsnVHH 13 specifically recognizes Ca2+-activated gelsolin (K (d) similar to 10 nM) while GsnVHH 11 binds gelsolin irrespective of Ca2+ (K (d) similar to 5 nM) but completely blocks its interaction with G-actin. Both GsnVHHs trace gelsolin in membrane ruffles of EGF-stimulated MCF-7 cells and delay cell migration without affecting F-actin severing/capping or actin nucleation activities by gelsolin. We conclude that VHHs represent a potent way of blocking structural proteins and that actin nucleation by gelsolin is more complex than previously anticipated.

• 出版日期2010-5