beta III spectrin binds to the Arp1 subunit of dynactin.
Cytoplasmic dynein is an intracellular motor responsible for endoplasmic reticulum-to-Golgi vesicle trafficking and retrograde axonal transport. The accessory protein dynactin has been proposed to mediate the association of dynein with vesicular cargo. Dynactin contains a 37-nm filament made up of the actin-related protein, Arp1, which may interact with a vesicle-associated spectrin network. Here, we demonstrate that Arp1 binds directly to the Golgi-associated betaIII spectrin isoform. We identify two Arp1-binding sites in betaIII spectrin, one of which overlaps with the actin-binding site conserved among spectrins. Although conventional actin binds weakly to betaIII spectrin, Arp1 binds robustly in the presence of excess F-actin. Dynein, dynactin, and betaIII spectrin co-purify on vesicles isolated from rat brain, and betaIII spectrin co-immunoprecipitates with dynactin from rat brain cytosol. In interphase cells, betaIII spectrin and dynactin both localize to cytoplasmic vesicles, co-localizing most significantly in the perinuclear region of the cell. In dividing cells, betaIII spectrin and dynactin co-localize to the developing cleavage furrow and mitotic spindle, a novel localization for betaIII spectrin. We hypothesize that the interaction between betaIII spectrin and Arp1 recruits dynein and dynactin to intracellular membranes and provides a direct link between the microtubule motor complex and its membrane-bounded cargo.
Holleran
E A
EA
Department of Cell and Molecular Pharmacology, University of California, San Francisco, California 94143, USA.
Ligon
L A
LA
Tokito
M
M
Stankewich
M C
MC
Morrow
J S
JS
Holzbaur
E L
EL
D000199:Actins / Q000737:chemistry* / Q000378:metabolism; D000818:Animals; D001665:Binding Sites; D001921:Brain / Q000378:metabolism; D019556:COS Cells; D002462:Cell Membrane / Q000378:metabolism; D003593:Cytoplasm / Q000378:metabolism; D003600:Cytosol / Q000378:metabolism; D000072159:Dynactin Complex; D004591:Electrophoresis, Polyacrylamide Gel; D005982:Glutathione Transferase / Q000378:metabolism; D015151:Immunoblotting; D007150:Immunohistochemistry; D008840:Microfilament Proteins; D008869:Microtubule-Associated Proteins / Q000737:chemistry*; D011233:Precipitin Tests; D011485:Protein Binding; D020033:Protein Isoforms; D017434:Protein Structure, Tertiary; D051381:Rats; D013049:Spectrin / Q000737:chemistry* / Q000378:metabolism*; D020798:Two-Hybrid System Techniques; D018909:src Homology Domains
D000199:Actins; C485513:Dbnl protein, rat; D000072159:Dynactin Complex; D008840:Microfilament Proteins; D008869:Microtubule-Associated Proteins; D020033:Protein Isoforms; C491475:actin-related protein 1, rat; D013049:Spectrin; D005982:Glutathione Transferase