Mammalian myosin Iα is concentrated near the plasma membrane in nerve growth cones

Abstract

To determine if unconventional myosins play a role in nerve outgrowth, antibodies specific for rat brain derived mammalian myosin I alpha (MMI alpha) were used to label cultured rat superior cervical ganglion nerve cells. Observations were made at both the light and electron microscopic level of resolution using preparative procedures designed to enhance the ability to precisely determine the relationship between antibody label and cellular structures in order to map the distribution and structural association of this myosin. Immunofluorescence showed that MMI alpha has a punctate distribution throughout the nerve cell body, neurites, and growth cones. In growth cones, MMI alpha staining is sometimes elevated in thin peripheral regions of high actin content at the leading edge. Immunoelectron microscopy using colloidal gold conjugated antibodies showed that in growth cones MMI alpha is absent from membranous organelles and is concentrated primarily in the cell cortex adjacent to the cell membrane. The cortical label is equally distributed between upper and lower membranes. The plasma membrane association of the MMI alpha label persists under conditions in which the actin cytoskeleton is perturbed or removed, suggesting a direct association between a fraction of MMI alpha and the plasma membrane. MMI alpha label is also associated with the non-cortical actin cytoskeleton. Partial disruption of the actin cytoskeleton using cytochalasin B causes redistribution of only a subset of MMI alpha label. These data suggest a complex relationship between MMI alpha, the actin cytoskeleton, and the plasma membrane in the growth cone. In contrast to its localization in the growth cone, in neuronal cell bodies MMI alpha is also associated with tubulovesicular structures. This suggests that at this location MMI alpha may either act as an organelle motor or is passively transported to the plasma membrane on vesicles.

Localization of unconventional myosins V and VI in neuronal growth cones

“…When myosin V and VI were detected with the same secondary antibody on different cultures, the stainings had similar intensities, myosin V appearing somewhat more abundant (data not shown). The punctate distribution of myosin V and myosin VI suggests that these myosins might be associated with organelle structures in the growth cone, although this may not be true for all puncta, as found in other myosin localization studies (Lewis and Bridgman, 1996;Evans et al, 1997). However, in some growth cones (Fig.…”

“…Myosin V and VI are present, but not enriched in growth cone particles. may be associated with vesicular structures; however, this may not be true for all puncta, as recently demonstrated in other myosin localization studies (Lewis and Bridgman, 1996;Evans et al, 1997). Our findings on myosin V distribution in growth cones of cultured chick DRG neurons is in complete agreement with the results of a recent study on myosin V localization in growth cones of rodent superior cervical ganglia neurons (Evans et al, 1997).…”

Diatom gliding is the result of an actin‐myosin motility system

“…These models suggest that actin filaments localized beneath the plasma membrane act as a scaffold for myosins coupled to transmembrane proteins that interact via adhesion molecules with a substratum to transfer the mechanical force generated by the actinmyosin complex. Myosin has been localized beneath the plasma membrane in these cells, and it has been suggested that a mechanical continuum exists between the substrate and the underlying actin cytoskeleton, although no other components of this complex have been identified [Lewis and Bridgman, 1996]. Recently myosin has been co-localized with actin beneath the plasma membrane, and it was suggested that this myosin interacts with phospholipids or a membrane protein during gliding [Dobrowolski et al, 1997;Pinder et al, 1998].…”

Myosin IIB Is Required for Growth Cone Motility

“…Mutations in the myosin II heavy chain A produce congenital defects that result in thrombocytopenia, giant platelets, and leukocyte inclusions but no obvious neurologic defects (Kelley et al, 2000). We cannot rule out the possibility that other myosins enriched in growth cones, such as myosin Ia, myosin Va, and myosin VI, also play a role in motility (Lewis and Bridgman, 1996;Wang et al, 1996;Evans et al, 1997;. Occasionally, filopodia-associated beads moved independently of detectable filopodia shortening.…”