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

TitleA llama-derived gelsolin single-domain antibody blocks gelsolin-G-actin interaction.
Publication TypeJournal Article
Year of Publication2010
AuthorsVan den Abbeele, A., S. De Clercq, A. De Ganck, V. De Corte, B. Van Loo, S. Hamdy Soror, V. Srinivasan, J. Steyaert, J. Vandekerckhove, and J. Gettemans
JournalCell Mol Life Sci
Volume67
Issue9
Pagination1519-35
Date Published2010 May
ISSN1420-9071
KeywordsActins, Animals, Calcium, Camelids, New World, Cell Line, Cell Movement, Crystallography, X-Ray, Epitopes, Gelsolin, Humans, Mitochondria, Models, Molecular, Molecular Sequence Data, Protein Structure, Tertiary, Recombinant Fusion Proteins, Single-Chain Antibodies
Abstract

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 Ca(2+)-activated gelsolin (K (d) approximately 10 nM) while GsnVHH 11 binds gelsolin irrespective of Ca(2+) (K (d) approximately 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.

DOI10.1007/s00018-010-0266-1
Alternate JournalCell. Mol. Life Sci.
PubMed ID20140750