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Quantitative analysis of APP axonal transport in neurons: role of JIP1 in enhanced APP anterograde transport.

Chiba K, Araseki M, Nozawa K, Furukori K, Araki Y, Matsushima T, Nakaya T, Hata S, Saito Y, Uchida S, Okada Y, Nairn AC, Davis RJ, Yamamoto T, Kinjo M, Taru H, Suzuki T - Mol. Biol. Cell (2014)

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.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Neuroscience, Graduate School of Pharmaceutical Sciences, Hokkaido University, Sapporo 060-0812, Japan.

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JIP1-dependent anterograde transport of APP cargo with enhanced fast velocity and efficient high frequency. APP-EGFP was expressed in primary cultured neurons from wild-type (A) or JIP1-KO (B, C) mice. JIP1b was exogenously expressed in neurons from JIP1-KO mice (C). The transport of APP cargo is shown with movies (left, top; see also supplemental movies), kymographs showing all vesicle movement (left, middle), and the traces of vesicles undergoing only anterograde transport (left, bottom). The cumulative frequencies of velocities (right) of anterograde transport of APP cargo are shown (data are normalized as percentages, and the position of the average velocity is indicated with an arrow). The ratios of anterograde (A), retrograde (R), and stationary (S) vesicles are indicated in the insets at the right. See Supplemental Movie S1A for A, S1B for B, and S1C for C. Scale bar, 5 μm. Statistical analysis for direction of movement vesicles is summarized in Table 1.
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Figure 1: JIP1-dependent anterograde transport of APP cargo with enhanced fast velocity and efficient high frequency. APP-EGFP was expressed in primary cultured neurons from wild-type (A) or JIP1-KO (B, C) mice. JIP1b was exogenously expressed in neurons from JIP1-KO mice (C). The transport of APP cargo is shown with movies (left, top; see also supplemental movies), kymographs showing all vesicle movement (left, middle), and the traces of vesicles undergoing only anterograde transport (left, bottom). The cumulative frequencies of velocities (right) of anterograde transport of APP cargo are shown (data are normalized as percentages, and the position of the average velocity is indicated with an arrow). The ratios of anterograde (A), retrograde (R), and stationary (S) vesicles are indicated in the insets at the right. See Supplemental Movie S1A for A, S1B for B, and S1C for C. Scale bar, 5 μm. Statistical analysis for direction of movement vesicles is summarized in Table 1.

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).


Quantitative analysis of APP axonal transport in neurons: role of JIP1 in enhanced APP anterograde transport.

Chiba K, Araseki M, Nozawa K, Furukori K, Araki Y, Matsushima T, Nakaya T, Hata S, Saito Y, Uchida S, Okada Y, Nairn AC, Davis RJ, Yamamoto T, Kinjo M, Taru H, Suzuki T - Mol. Biol. Cell (2014)

JIP1-dependent anterograde transport of APP cargo with enhanced fast velocity and efficient high frequency. APP-EGFP was expressed in primary cultured neurons from wild-type (A) or JIP1-KO (B, C) mice. JIP1b was exogenously expressed in neurons from JIP1-KO mice (C). The transport of APP cargo is shown with movies (left, top; see also supplemental movies), kymographs showing all vesicle movement (left, middle), and the traces of vesicles undergoing only anterograde transport (left, bottom). The cumulative frequencies of velocities (right) of anterograde transport of APP cargo are shown (data are normalized as percentages, and the position of the average velocity is indicated with an arrow). The ratios of anterograde (A), retrograde (R), and stationary (S) vesicles are indicated in the insets at the right. See Supplemental Movie S1A for A, S1B for B, and S1C for C. Scale bar, 5 μm. Statistical analysis for direction of movement vesicles is summarized in Table 1.
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Figure 1: JIP1-dependent anterograde transport of APP cargo with enhanced fast velocity and efficient high frequency. APP-EGFP was expressed in primary cultured neurons from wild-type (A) or JIP1-KO (B, C) mice. JIP1b was exogenously expressed in neurons from JIP1-KO mice (C). The transport of APP cargo is shown with movies (left, top; see also supplemental movies), kymographs showing all vesicle movement (left, middle), and the traces of vesicles undergoing only anterograde transport (left, bottom). The cumulative frequencies of velocities (right) of anterograde transport of APP cargo are shown (data are normalized as percentages, and the position of the average velocity is indicated with an arrow). The ratios of anterograde (A), retrograde (R), and stationary (S) vesicles are indicated in the insets at the right. See Supplemental Movie S1A for A, S1B for B, and S1C for C. Scale bar, 5 μm. Statistical analysis for direction of movement vesicles is summarized in Table 1.
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).

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.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Neuroscience, Graduate School of Pharmaceutical Sciences, Hokkaido University, Sapporo 060-0812, Japan.

Show MeSH
Related in: MedlinePlus