Synaptic scaffolding protein SYD-2 clusters and activates kinesin-3 UNC-104 in C. elegans.
Kinesin-3 motor UNC-104/KIF1A is essential for transporting synaptic precursors to synapses. Although the mechanism of cargo binding is well understood, little is known how motor activity is regulated. We mapped functional interaction domains between SYD-2 and UNC-104 by using yeast 2-hybrid and pull-down assays and by using FRET/fluorescence lifetime imaging microscopy to image the binding of SYD-2 to UNC-104 in living Caenorhabditis elegans. We found that UNC-104 forms SYD-2-dependent axonal clusters (appearing during the transition from L2 to L3 larval stages), which behave in FRAP experiments as dynamic aggregates. High-resolution microscopy reveals that these clusters contain UNC-104 and synaptic precursors (synaptobrevin-1). Analysis of motor motility indicates bi-directional movement of UNC-104, whereas in syd-2 mutants, loss of SYD-2 binding reduces net anterograde movement and velocity (similar after deleting UNC-104's liprin-binding domain), switching to retrograde transport characteristics when no role of SYD-2 on dynein and conventional kinesin UNC-116 motility was found. These data present a kinesin scaffolding protein that controls both motor clustering along axons and motor motility, resulting in reduced cargo transport efficiency upon loss of interaction.
Wagner
Oliver I
OI
DFG Research Center for Molecular Physiology of the Brain (CMPB), Georg August University, 37073 Göttingen, Germany. owagner@life.nthu.edu.tw
Esposito
Alessandro
A
Köhler
Barbara
B
Chen
Chih-Wei
CW
Shen
Che-Piao
CP
Wu
Gong-Her
GH
Butkevich
Eugenia
E
Mandalapu
Sailaja
S
Wenzel
Dirk
D
Wouters
Fred S
FS
Klopfenstein
Dieter R
DR
D000818:Animals; D001369:Axons / Q000378:metabolism; D017173:Caenorhabditis elegans / Q000378:metabolism*; D029742:Caenorhabditis elegans Proteins / Q000235:genetics / Q000378:metabolism*; D036681:Fluorescence Recovery After Photobleaching; D031541:Fluorescence Resonance Energy Transfer; D036341:Intercellular Signaling Peptides and Proteins; D009419:Nerve Tissue Proteins / Q000378:metabolism*; D010750:Phosphoproteins / Q000235:genetics / Q000378:metabolism*; D054730:Protein Interaction Domains and Motifs; D025941:Protein Interaction Mapping; D013572:Synaptic Vesicles / Q000378:metabolism*
D029742:Caenorhabditis elegans Proteins; D036341:Intercellular Signaling Peptides and Proteins; D009419:Nerve Tissue Proteins; D010750:Phosphoproteins; C490598:SYD-2 protein, C elegans; C423186:UNC-104 protein, C elegans