LIM kinase 1 and cofilin regulate actin filament population required for dynamin-dependent apical carrier fission from the trans-Golgi network

• Published in: Molecular biology of the cell 2009-01, Vol.20 (1), p.438-451
• Main Authors: Salvarezza, Susana B, Deborde, Sylvie, Schreiner, Ryan, Campagne, Fabien, Kessels, Michael M, Qualmann, Britta, Caceres, Alfredo, Kreitzer, Geri, Rodriguez-Boulan, Enrique
• Format: Article
• Language: English
• Subjects: Actin Depolymerizing Factors - genetics; Actin Depolymerizing Factors - metabolism; Actins - metabolism; Animals; Biomarkers - metabolism; Cell Line; Cell Polarity; Cytoskeleton - metabolism; Dogs; Dynamins - genetics; Dynamins - metabolism; Golgi Apparatus - metabolism; Isoenzymes - genetics; Isoenzymes - metabolism; Lim Kinases - genetics; Lim Kinases - metabolism; Models, Biological; Neural Cell Adhesion Molecules - genetics; Neural Cell Adhesion Molecules - metabolism; Protein Transport - physiology; Receptor, Nerve Growth Factor - genetics; Receptor, Nerve Growth Factor - metabolism; Recombinant Fusion Proteins - genetics; Recombinant Fusion Proteins - metabolism; RNA, Small Interfering - genetics; RNA, Small Interfering - metabolism; trans-Golgi Network - metabolism
• ISSN: 1059-1524; 1939-4586
• DOI: 10.1091/mbc.E08-08-0891

The functions of the actin cytoskeleton in post-Golgi trafficking are still poorly understood. Here, we report the role of LIM Kinase 1 (LIMK1) and its substrate cofilin in the trafficking of apical and basolateral proteins in Madin-Darby canine kidney cells. Our data indicate that LIMK1 and cofilin organize a specialized population of actin filaments at the Golgi complex that is selectively required for the emergence of an apical cargo route to the plasma membrane (PM). Quantitative pulse-chase live imaging experiments showed that overexpression of kinase-dead LIMK1 (LIMK1-KD), or of LIMK1 small interfering RNA, or of an activated cofilin mutant (cofilin S3A), selectively slowed down the exit from the trans-Golgi network (TGN) of the apical PM marker p75-green fluorescent protein (GFP) but did not interfere with the apical PM marker glycosyl phosphatidylinositol-YFP or the basolateral PM marker neural cell adhesion molecule-GFP. High-resolution live imaging experiments of carrier formation and release by the TGN and analysis of peri-Golgi actin dynamics using photoactivatable GFP suggest a scenario in which TGN-localized LIMK1-cofilin regulate a population of actin filaments required for dynamin-syndapin-cortactin-dependent generation and/or fission of precursors to p75 transporters.