Direct visualization of the movement of the monomeric axonal transport motor UNC-104 along neuronal processes in living Caenorhabditis elegans.
The formation and function of axons depends on the microtubule-based transport of cellular components from their sites of synthesis in the neuronal cell body to their sites of utilization at the axon terminus. To directly visualize this axonal transport in a living organism, we constructed transgenic lines of Caenorhabditis elegans that express green fluorescent protein fused to the monomeric synaptic vesicle transport motor, UNC-104. This UNC-104:: GFP construct rescued the Unc-104 mutant phenotype and was expressed throughout the nervous system. Using time-lapse confocal fluorescence microscopy, we were able to visualize fluorescent motor proteins moving in both directions along neuronal processes, some of which were identified definitely as axons and others as dendrites. Using kymograph analysis, we followed the movement of >900 particles. Most of them moved in one direction, but not necessarily at the same velocity. Ten percent of the observed particles reversed direction of movement during the period of observation, and 10% exhibited periods of movement interspersed with pauses. During episodes of persistent movement, particles moved at an average velocity of 1.02 microm/sec, which is close to the in vitro velocity of microtubule gliding driven by purified monomeric kinesin at high motor density. To our knowledge, this is the first direct visualization and analysis of the movement of specifically labeled microtubule motor proteins along axons in vivo.
Zhou
H M
HM
Section of Molecular and Cellular Biology, University of California at Davis, Davis, California 95616, USA.
Brust-Mascher
I
I
Scholey
J M
JM
D000818:Animals; D001370:Axonal Transport / Q000502:physiology*; D001369:Axons / Q000378:metabolism / Q000648:ultrastructure; D017173:Caenorhabditis elegans; D029742:Caenorhabditis elegans Proteins; D003712:Dendrites / Q000378:metabolism / Q000648:ultrastructure; D015870:Gene Expression; D018014:Gene Transfer Techniques; D049452:Green Fluorescent Proteins; D007734:Kymography; D008164:Luminescent Proteins / Q000235:genetics; D008856:Microscopy, Fluorescence; D020409:Molecular Motor Proteins / Q000378:metabolism* / Q000648:ultrastructure*; D009043:Motor Activity / Q000235:genetics; D009419:Nerve Tissue Proteins / Q000235:genetics / Q000378:metabolism*; D009474:Neurons / Q000378:metabolism / Q000648:ultrastructure; D009928:Organ Specificity; D010641:Phenotype; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D013997:Time Factors
D029742:Caenorhabditis elegans Proteins; D008164:Luminescent Proteins; D020409:Molecular Motor Proteins; D009419:Nerve Tissue Proteins; D011993:Recombinant Fusion Proteins; C423186:UNC-104 protein, C elegans; D049452:Green Fluorescent Proteins