Cyclical Action of the WASH Complex: FAM21 and Capping Protein Drive WASH Recycling, Not Initial Recruitment

• Published in: Developmental cell 2013-01, Vol.24 (2), p.169-181
• Main Authors: Park, Laura, Thomason, Peter A., Zech, Tobias, King, Jason S., Veltman, Douwe M., Carnell, Michael, Ura, Seiji, Machesky, Laura M., Insall, Robert H.
• Format: Article
• Language: English
• Subjects: Actin Capping Proteins - metabolism; Actins - metabolism; Biological and medical sciences; Cell differentiation, maturation, development, hematopoiesis; Cell Line, Tumor; Cell physiology; Dictyostelium - genetics; Dictyostelium - metabolism; Exocytosis; Fundamental and applied biological sciences. Psychology; Humans; Lysosomes - metabolism; Microfilament Proteins - genetics; Microfilament Proteins - metabolism; Molecular and cellular biology; Mutation; Protozoan Proteins - genetics; Protozoan Proteins - metabolism; RNA Interference; RNA, Small Interfering; Vacuolar Proton-Translocating ATPases - metabolism; Vesicular Transport Proteins - genetics; Vesicular Transport Proteins - metabolism
• ISSN: 1534-5807; 1878-1551
• DOI: 10.1016/j.devcel.2012.12.014

WASH causes actin to polymerize on vesicles involved in retrograde traffic and exocytosis. It is found within a regulatory complex, but the physiological roles of the other four members are unknown. Here we present genetic analysis of the subunits’ individual functions in Dictyostelium. Mutants in each subunit are completely blocked in exocytosis. All subunits except FAM21 are required to drive actin assembly on lysosomes. Without actin, lysosomes never recycle vacuolar-type H+-adenosine triphosphatase (V-ATPase) or neutralize to form postlysosomes. However, in FAM21 knockout lysosomes, WASH generates excessive, dynamic streams of actin. These successfully remove V-ATPase, neutralize, and form huge postlysosomes. The distinction between WASH and FAM21 phenotypes is conserved in human cells. Thus, FAM21 and WASH act at different steps of a cyclical pathway in which FAM21 mediates recycling of the complex back to acidic lysosomes. Recycling is driven by FAM21’s interaction with capping protein, which couples the WASH complex to dynamic actin on vesicles. [Display omitted] ► Dictyostelium WASH complex neutralizes lysosomes, then is recycled in small vesicles ► FAM21 is required at a different step than the other four WASH complex subunits ► Mutants in FAM21 can make actin on vesicles but cannot recycle WASH complex after use ► WASH complex is recycled coupling between actin, capping protein, and FAM21 The WASH complex catalyzes actin polymerization on intracellular vesicles. Much of its biology remains unclear, including the different subunits’ roles. Park et al. show that the FAM21 subunit is—unexpectedly—not needed for actin polymerization. Instead, FAM21 drives recycling by connecting WASH complex to the inward flow of actin filaments.