All restriction enzymes and T4 DNA ligase were from TaKaRa

their tasks in tissue surveillance and host defense. To control these activities before and after polarization, macrophages continuously form actin-rich membrane protrusions and extend filopodia from their cell surface [10,11]. Changes in the organization of the actin cytoskeleton thereby enable the cell to dynamically adapt its morphology to suit its particular function and differentiation state. For example, LPS induces polymerization of cytoskeletal actin filaments, cell spreading, and the formation of filopodia, lamellipodia, and membrane ruffles in monocytes and macrophages [12,13]. Likewise, IL-4, which is released during tissue injury, 1 May 2014 | Volume 9 | Issue 5 | e96786 PLOS ONE | www.plosone.org Glucose Controls Macrophage Morphodynamics 870281-82-6 custom synthesis 19653943″ title=View Abstract(s)”>PubMed ID:http://www.ncbi.nlm.nih.gov/pubmed/19653943 causes the rearrangement of actin-rich podosomes to form rosettes in M2 macrophages, enabling degradation of-and migration through-dense extracellular matrices [14]. The rearrangements of cytoskeletal actin filaments that steer this behavior comprise multiple steps, including the nucleation and elongation of new filaments from ATP-bound G-actin monomers, the addition of these monomers to the barbed ends of existing filaments, the hydrolysis of actin-bound ATP within the growing filament, and the dissociation of ADP-G-actin at the pointed end [15�18]. ADP on liberated G-actin is substituted with ATP, producing new ATP-G-actin monomers for incorporation. The ensemble of activities in this complex process is regulated by more than a hundred actin-associated proteins (ABPs), several of which are influenced by the availability of ATP. Metabolism, specifically the binding and hydrolysis of ATP and ATP-ADP exchange, thereby not only dictates the behavior of the actin filaments them