Quantitative analysis of APP axonal transport in neurons: role of JIP1 in enhanced APP anterograde transport.
Bottom Line: In JIP1-deficient neurons, we find that both the fast velocity (∼2.7 μm/s) and high frequency (66%) of anterograde transport of APP cargo are impaired to a reduced velocity (∼1.83 μm/s) and a lower frequency (45%).Furthermore, efficient APP axonal transport is not influenced by phosphorylation of APP at Thr-668, a site known to be phosphorylated by JNK.Our quantitative analysis indicates that enhanced fast-velocity and efficient high-frequency APP anterograde transport observed in neurons are mediated by novel roles of JIP1b.
Affiliation: Laboratory of Neuroscience, Graduate School of Pharmaceutical Sciences, Hokkaido University, Sapporo 060-0812, Japan.Show MeSH
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Mentions: To reveal the role of JIP1 in the enhanced and efficient fast anterograde transport of APP cargo, we analyzed quantitatively the axonal transport of APP cargo in primary cultured mixed cortical/hippocampal neurons from JIP1-KO mice (Whitmarsh et al., 2001). Wild-type (JIP1+/+) and JIP1-deficient (JIP1−/−) neurons expressing APP-enhanced green fluorescent protein (EGFP) were assayed for APP cargo transport using a total internal reflectance fluorescence (TIRF) microscopy system (Araki et al., 2007). APP cargo was subject to fast anterograde transport (2.73 ± 0.69 μm/s, n = 533) in wild-type neurons (Figure 1A and Supplemental Movie S1A) but was transported anterogradely with a reduced average velocity of 1.83 ± 0.49 μm/s (n = 538) in neurons lacking JIP1 (Figure 1B and Supplemental Movie S1B; compare B with A; p < 0.001). Of note, this reduced rate is almost identical to the speed of alcadein cargoes transported by kinesin-1 (Araki et al., 2007), indicating that JIP1 enhances fast anterograde transport. We excluded stationary vesicles moving at <0.4 μm/s because such vesicles are indistinguishable from those exhibiting Brownian motion. The essential role of JIP1 in the enhanced high-speed velocity of APP cargo was confirmed by expressing FLAG-JIP1b in JIP1−/− neurons, which restored the enhanced fast transport of APP-EGFP (2.68 ± 0.65 μm/s, n = 96; Figure 1C and Supplemental Movie S1C; compare C with B; p < 0.001).
Affiliation: Laboratory of Neuroscience, Graduate School of Pharmaceutical Sciences, Hokkaido University, Sapporo 060-0812, Japan.