JIP1 Sustains Long Distance Anterograde Transport via Direct Regulation of Kinesin Autoinhibition and Coordination of Retrograde Motor Association

Abstract

Analysis of the highly processive vesicular cargo APP (amyloid precursor protein) in primary mammalian neurons shows axonal transport characterized by long run lengths with few pauses or directional switches. In order to elucidate the molecular mechanism for sustaining long-distance processive motility, we focused on the adaptor protein JIP1, which binds directly to APP and KLC (kinesin light chain). Upon siRNA depletion of JIP1 from primary neurons, we observed significant defects in both anterograde and retrograde APP transport. We further investigated the association of JIP1 with kinesin-1 and identified novel and distinct interactions between JIP1 and both kinesin heavy chain (KHC) stalk and tail. Using in vitro TIRF motility assays with single-molecule resolution, we found that addition of JIP1 relieves autoinhibition of KHC, leading to activation of motility and increased run frequency, run length and speed. Interestingly, truncated constructs of JIP1 that only bind KHC stalk or KHC tail were sufficient to initiate processive runs, but could not fully recapitulate the enhancement of KHC run length or speed observed with full-length JIP1. In addition, JIP1 can coordinate both kinesin- and dynein-driven motility, as we identify a novel interaction between JIP1 and the p150Glued subunit of dynactin. Both in vitro and in primary neurons, addition of the JIP1-binding region of p150Glued competitively inhibits JIP1-mediated enhancement of KHC processivity. In contrast to a stochastic tug-of-war model, our data suggest that JIP1 sustains long-distance transport by coordinating the formation of two distinct motile complexes - an anterograde complex that binds directly to KHC and a retrograde complex via its interaction with dynactin.