Myosin II functions in actin-bundle turnover in neuronal growth cones

Abstract

Retrograde actin flow works in concert with cell adhesion to generate traction forces that are involved in axon guidance in neuronal growth cones. Myosins have been implicated in retrograde flow, but identification of the specific myosin subtype(s) involved has been controversial. Using fluorescent speckle microscopy (FSM) to assess actin dynamics, we report that inhibition of myosin II alone decreases retrograde flow by 51% and the remaining flow can be almost fully accounted for by the 'push' of plus-end actin assembly at the leading edge of the growth cone. Interestingly, actin bundles that are associated with filopodium roots elongated by approximately 83% after inhibition of myosin II. This unexpected result was due to decreased rates of actin-bundle severing near their proximal (minus or pointed) ends which are located in the transition zone of the growth cone. Our study reveals a mechanism for the regulation of actin-bundle length by myosin II that is dependent on actin-bundle severing, and demonstrate that retrograde flow is a steady state that depends on both myosin II contractility and actin-network treadmilling.

Retrograde Fluxes of Focal Adhesion Proteins in Response to Cell Migration and Mechanical Signals

“…The process may be regulated by the rates of actin nucleation, elongation, dissociation, and possibly myosin-dependent forces that pull on actin filaments (Medeiros et al, 2006). The dependence of the flux on cell migration and mechanical signals explains why prominent stress fibers are present only in relatively immotile cells (Herman et al, 1981;Tomasek et al, 1982), and in cells on stiff substrates (Pelham and Wang, 1997).…”

Cell biology of embryonic migration

“…Their release at the cell's trailing edge is accompanied by stretching, driven by myosin II contractility and regulated by FAK and Src signaling (Henson et al, 1999;Medeiros et al, 2006;Vallotton et al, 2004;Verkhovsky et al, 1999;Zhang et al, 2003). The stretching force created during the focal adhesion release is believed to be sufficient to open the stretch-activated Ca 2+ channels that trigger activation of the calcium-dependent protease calpain that participates in focal adhesion disassembly by cleaving a number of focal adhesion proteins, including talin, vinculin, and FAK (Franco and Huttenlocher, 2005;Franco et al, 2004).…”

Conventional myosins – unconventional functions

“…At the leading edge of the growth cone is a broad, relatively thin, veil-like lamellipodium, actin polymerisa-tion pushing the plasma membrane forwards (Pantaloni et al 2001;Pollard and Borisy 2003); actin filaments, thus formed, are translocated rearwards, initiating a phenomenon known as retrograde actin flow. The forces required to move the bulk of the growth cone forwards are much greater than can be achieved through actin polymerisation alone, hence a requirement for myosin 2 molecular motors, which act vectorially and power forward outgrowth (Wylie et al 1998;Bridgman et al 2001) and rearward retraction (Amano et al 1998;Wylie and Chantler 2003), as well as retrograde actin flow (Lin et al 1996;Cai et al 2006;Medeiros et al 2006). Similar actions underpin the generation of protrusions in non-neuronal cells.…”

“…There is an inverse relationship between neuronal growth cone advance and retrograde actin flow (Lin and Forscher 1995), which is driven by one or more myosin 2 isoforms (Lin et al 1996;Medeiros et al 2006). Although it is clear that retrograde actin flow is integral to forward propulsion of the central domain of the growth cone, the exact identities of the isoforms responsible remain unknown.…”