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subunit NR2B was much slower at the AIS compared with vesicles carrying
the synaptic vesicle protein VAMP2. Song et al. then investigated whether
this selectivity was dictated by motor proteins or cargo. Using chimeric
kinesins, they found that the observed selective axonal entry was not con-
trolled by the motor domain or the cargo alone, but by the transport efficacy
of the motor-cargo complex as a whole (
Song et al., 2009
).
Both the membrane and cytoplasmic diffusion barriers are abolished
when the actin cytoskeleton is disrupted by Latrunculin A or Cytochalasin
D, indicating a dependence on the anchorage of transmembrane proteins to
the actin cytoskeleton (
Nakada et al., 2003; Song et al., 2009; Winckler
et al., 1999
). Loss of AnkG also removes the barrier for diffusional entry
of large molecules into the axon (
Song et al., 2009
). The AIS cytoskeleton
may therefore be central to regulating axonal entry, but it has also been pro-
posed that the actin cytoskeleton is involved in preventing axonal transport
of dendritic cargo by directing it away from the axon via myosin motors on
polarized actin filaments (
Arnold, 2009
). This is based on the finding that
interfering with myosin Va prevents dendritic accumulation of the AMPA
GluR1 receptor, while fusion of Channelrhodopsin-2 to the myosin-Va-
binding domain results in dendritic targeting (
Lewis, Mao, Svoboda, &
Arnold, 2009
).
The role of the microtubule cytoskeleton in regulating polarized trans-
port has been intensively researched, and it is now understood that differ-
ences in the organization of the microtubule cytoskeleton between
neuronal compartments are involved in regulating axonal entry of cargo.
These include differences in microtubule orientation, types of associated
proteins, and posttranslational modifications (
Hirokawa & Takemura,
2005; Kapitein & Hoogenraad, 2011
). The AIS is characterized by a unique
network of fasciculated microtubules with a polarized orientation and a
lower tyrosinated/detyrosinated ratio than the somatodendritic domain
(
Konishi & Setou, 2009; Palay, Sotelo, Peters, &Orkand, 1968
). These bun-
dles of microtubules are linked to AnkG by the end-binding proteins EB1
and EB3 (
Leterrier et al., 2011; Sobotzik et al., 2009
), which is suggested to
increase microtubule stability at the AIS. Stable microtubules with fast-
growing plus ends oriented away from the cell body are thought to provide
directional information for axonal entry of cargo. The motor domain of
kinesin-1, for example, preferentially binds to microtubules in the AIS. This
preference is abolished when microtubule dynamics are suppressed with
paclitaxel (
Kapitein et al., 2010; Nakata & Hirokawa, 2003
). During devel-
opment, microtubule stabilization precedes axon formation and targeting of
