Characterization of the binding mode of JNK-interacting protein 1 (JIP1) to kinesin-light chain 1 (KLC1)

Abstract

JIP1 was first identified as scaffold protein for the MAP kinase JNK and is a cargo protein for the kinesin1 molecular motor. JIP1 plays significant and broad roles in neurons, mainly as a regulator of kinesin1-dependent transport, and is associated with human pathologies such as cancer and Alzheimer disease. JIP1 is specifically recruited by the kinesin-light chain 1 (KLC1) of kinesin1, but the details of this interaction are not yet fully elucidated. Here, using calorimetry, we extensively biochemically characterized the interaction between KLC1 and JIP1. Using various truncated fragments of the tetratricopeptide repeat (TPR) domain of KLC1, we narrowed down its JIP1-binding region and identified seven KLC1 residues critical for JIP1 binding. These isothermal titration calorimetry (ITC)-based binding data enabled us to footprint the JIP1-binding site on KLC1-TPR. This footprint was used to uncover the structural basis for the marginal inhibition of JIP1 binding by the autoinhibitory LFP-acidic motif of KLC1, as well as for the competition between JIP1 and another cargo protein of kinesin1, the W-acidic motif-containing alcadein-α. Also, we examined the role of each of these critical residues of KLC1 for JIP1 binding in light of the previously reported crystal structure of the KLC1-TPR:JIP1 complex. Finally, sequence search in eukaryotic genomes identified several proteins, among which is SH2D6, that exhibit a motif similar to the KLC1-binding motif of JIP1. Overall, our extensive biochemical characterization of the KLC:JIP1 interaction, as well as identification of potential KLC1-binding partners, improves the understanding of how this growing family of cargos is recruited to kinesin1 by KLC1.

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“…MAPK8IP1/JIP1 is a critical regulator of autophagosome transport in neurons, which ensures the fidelity of retrograde autophagosome transport in the axon and is highly sensitive to defects in autophagy [ 27 ]. JIP1 also plays an important role in neurons as a regulator of kinesin-1-dependent transport [ 28 ]. RAPSN , which is considered to be related to brain/cognition in DDG2P, shows low expression in HPA.…”

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“…The tryptophan and tyrosine residues in W-acidic and Y-acidic motifs, respectively, are essential for binding (14,(17)(18)(19). The X-ray crystal structures of cargo-adaptor peptides from SKIP, JIP1 and Torsin A complexed with KLC TPR domains highlight how these residues interact with their receptor.…”

“…These natural motor-cargo adaptor interactions have µM affinities in vitro (14,17,19,20). Previously, we have proposed that tighter interactions are achieved in the cell through avidity, as multiple copies of the adaptor peptides are expressed on the surface of organelles, they may be presented as a pair of motifs in adaptor proteins, and the kinesin-1 heterotetramer presents a pair of KLCs (Figure 1a) (7,11,21).…”

“…Our own work and that of others has identified identified two distinct classes of peptide sequences that are recognised by KLC TPR : tryptophan-acidic (W-acidic) motifs found in the lysosomal cargo adaptor SKIP as well as many others feature a tryptophan residue flanked by aspartic or glutamic acids in the consensus L/M-D/E-W-D/E (7,(11)(12)(13)(14)(15)(16); and Y-acidic motifs at the C termini of the axonal transport cargo adaptor JIP1 and Torsin A display a tyrosine residue flanked by acidic residues (17)(18)(19)(20). In mammals there are four closely related KLC isoforms (KLC1 -4).…”

“…In mammals there are four closely related KLC isoforms (KLC1 -4). Whilst Wacidic motifs typically bind both KLC1 TPR and KLC2 TPR , C-terminal Y-acidic motifs bind KLC1 TPR with a much higher affinity compared to KLC2 TPR (17,19,20). These motifs bind at distinct yet overlapping locations on the concave surface of the KLC TPR .…”