A Role for ARF6 and ARNO in the regulation of endosomal dynamics in neurons.
During development, neuronal processes extend, branch and navigate to ultimately synapse with target tissue. We have shown a regulatory role for ARNO and ARF6 in dendritic branching and axonal elongation and branching during neuritogenesis, particularly with respect to cytoskeletal dynamics. Here, we have examined the role of ARF6 and the ARF GEF ARNO in endosomal dynamics during neurite elongation in hippocampal neurons. Axonal and dendritic endosomes were labeled by expression of the endosomal marker, endotubin. Expression of endotubin-green fluorescent protein resulted in targeting to tubular-vesicular structures throughout the somatodendritic and axonal domains. These endosomal structures did not colocalize with conventional early or late endosomal markers or with the synaptic vesicle marker, SV2. However, they did label with internalized lectin, indicating that they are endosomal structures. Expression of catalytically inactive ARNO (ARNO-E156K) or inactive ARF6 (ARF6-T27N) caused a redistribution of endotubin to the cell surface of the axons and dendrites. In contrast, expression of these constructs had no effect upon the distribution of SV2-positive structures. Furthermore, expression of inactive ARF1 (ARF1-T31N) did not change endotubin distribution. These results suggest that endotubin labels a distinct endosomal structure in neurons and that ARNO and ARF6 mediate neurite extension through the regulation of this compartment.
Hernández-Deviez
Delia
D
Department of Cell Biology and Anatomy, University of Arizona, Arizona Health Sciences Center, PO Box 245044, Tucson, AZ 85724, USA.|Current address: Institute for Molecular Bioscience, University of Queensland, Brisbane, QLD 4072, Australia.
Mackay-Sim
Alan
A
National Centre for Adult Stem Cell Research, Griffith University, Nathan Campus, Brisbane, QLD 4111, Australia.
Wilson
Jean M
JM
Department of Cell Biology and Anatomy, University of Arizona, Arizona Health Sciences Center, PO Box 245044, Tucson, AZ 85724, USA.
D000089982:ADP-Ribosylation Factor 6; D020727:ADP-Ribosylation Factors / Q000502:physiology*; D000818:Animals; D001369:Axons / Q000378:metabolism; D002462:Cell Membrane / Q000378:metabolism; D002478:Cells, Cultured; D004247:DNA / Q000378:metabolism; D011992:Endosomes / Q000378:metabolism*; D020690:GTPase-Activating Proteins / Q000502:physiology*; D006624:Hippocampus / Q000378:metabolism; D008856:Microscopy, Fluorescence; D008954:Models, Biological; D009474:Neurons / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley; D013499:Surface Properties
D000089982:ADP-Ribosylation Factor 6; D020690:GTPase-Activating Proteins; C102474:cytohesin-2; D004247:DNA; D020727:ADP-Ribosylation Factors; C000715349:Arf6 protein, rat