Recruitment of a myosin heavy chain kinase to actin-rich protrusions in Dictyostelium

Abstract

Nonmuscle myosin II plays fundamental roles in cell body translocation during migration and is typically depleted or absent from actin-based cell protrusions such as lamellipodia, but the mechanisms preventing myosin II assembly in such structures have not been identified [1-3]. In Dictyostelium discoideum, myosin II filament assembly is controlled primarily through myosin heavy chain (MHC) phosphorylation. The phosphorylation of sites in the myosin tail domain by myosin heavy chain kinase A (MHCK A) drives the disassembly of myosin II filaments in vitro and in vivo [4]. To better understand the cellular regulation of MHCK A activity, and thus the regulation of myosin II filament assembly, we studied the in vivo localization of native and green fluorescent protein (GFP)-tagged MHCK A. MHCK A redistributes from the cytosol to the cell cortex in response to stimulation of Dictyostelium cells with chemoattractant in an F-actin-dependent manner. During chemotaxis, random migration, and phagocytic/endocytic events, MHCK A is recruited preferentially to actin-rich leading-edge extensions. Given the ability of MHCK A to disassemble myosin II filaments, this localization may represent a fundamental mechanism for disassembling myosin II filaments and preventing localized filament assembly at sites of actin-based protrusion.

Myosin heavy chain kinases play essential roles in Ca2+, but not cAMP, chemotaxis and the natural aggregation of Dictyostelium discoideum

“…MhckA had been shown to localize to the pseudopod (Steimle et al, 2001a;Steimle et al, 2001b;Steimle et al, 2002;Liang et al, 2002), which, when extending, is relatively devoid of myosin II (Moores et al, 1996). The accumulation of myosin II in the pseudopods of mhckA 2 cells, as we have shown here, supports the suggestion that MhckA phosphorylates Mhc in the pseudopod, thus blocking myosin II polymerization in the dynamic actin gel that drives pseudopodial expansion (Steimle et al, 2001a;Steimle et al, 2001b;Steimle et al, 2002).…”

“…6I,J) also stained intensely for myosin II. These latter results suggest that in the absence of MhckA, which localizes in pseudopods (Steimle et al, 2001a;Steimle et al, 2001b;Steimle et al, 2002;Liang et al, 2002), myosin II abnormally accumulates in that structure, presumably in an unphosphorylated and thus polymerized state.…”

Identification and Characterization of a Novel α-Kinase with a von Willebrand Factor A-like Motif Localized to the Contractile Vacuole and Golgi Complex inDictyostelium discoideum

“…The thoroughly analyzed members to date include MHCK A, MHCK B, and MHCK C, and at both the biochemical level and in vivo all three seem to be involved in the control of myosin II filament assembly and localization within the cell (Steimle et al, 2001b;Liang et al, 2002;Nagasaki et al, 2002;Rico and Egelhoff, 2003). We have identified a fourth member of this family in Dictyostelium with similar domain organization that we refer to as MHCK D (our unpublished data).…”

TOR Complex 2 Integrates Cell Movement during Chemotaxis and Signal Relay in Dictyostelium

“…All three mutant strains exhibit moderately altered patterns of chemoattractant-mediated myosin II assembly. In wild-type cells, the levels of myosin II associated with the cytoskeleton undergo an initial slight decrease 5-10 s after uniform chemoattrac- S. Lee et al tant stimulation and then increase about twofold, peaking at ϳ30 s, before decreasing to basal levels (Steimle et al, 2001;Park et al, 2004). The rip3 Ϫ cells exhibit the strongest phenotype, with a significantly reduced peak of myosin II assembly upon chemoattractant stimulation ( Figure 5C).…”