Dynamics of cytoplasmic dynein in living cells and the effect of a mutation in the dynactin complex actin-related protein Arp1.
Cytoplasmic dynein is a minus-end-directed microtubule motor that participates in multiple cellular activities such as organelle transport and mitotic spindle assembly [1]. To study the dynamic behavior of cytoplasmic dynein in the filamentous fungus Aspergillus nidulans, we replaced the gene for the cytoplasmic dynein heavy chain, nudA, with a gene encoding a green fluorescent protein (GFP)-tagged chimera, GFP-nudA. The GFP-NUDA fusion protein is fully functional in vivo: strains expressing only the GFP-tagged nudA grow as well as wild-type strains. Fluorescence microscopy showed GFP-NUDA to be in comet-like structures that moved in the hyphae toward the growing tip. Retrograde movement of some GFP-NUDA comets after they arrived at the tip was also observed. These dynamics of GFP-NUDA were not observed in cells treated with a microtubule-destabilizing drug, benomyl, suggesting they are microtubule-dependent. The rate of GFP-NUDA tip-ward movement is similar to the rate of cytoplasmic microtubule polymerization toward the hyphal tip, suggesting that GFP-NUDA is associated and moving with the polymerizing ends of microtubules. A mutation in actin-related protein Arp1 of the dynactin complex abolishes the presence of these dynamic GFP-NUDA structures near the hyphal tip, suggesting a targeting role of the dynactin complex.
Xiang
X
X
Department of Biochemistry and Molecular Biology, Uniformed Services University of the Health Sciences, Bethesda, MD 20814, USA. xxiang@usuhs.mil.
Han
G
G
Winkelmann
D A
DA
Zuo
W
W
Morris
N R
NR
D001233:Aspergillus nidulans / Q000235:genetics / Q000502:physiology*; D051836:COUP Transcription Factors; D003593:Cytoplasm / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000235:genetics* / Q000378:metabolism; D000072159:Dynactin Complex; D004398:Dyneins / Q000235:genetics / Q000378:metabolism*; D005656:Fungal Proteins / Q000235:genetics / Q000378:metabolism; D049452:Green Fluorescent Proteins; D008164:Luminescent Proteins / Q000235:genetics / Q000378:metabolism; D008869:Microtubule-Associated Proteins / Q000235:genetics / Q000378:metabolism; D009154:Mutation; D011987:Receptors, Steroid; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D014157:Transcription Factors / Q000235:genetics* / Q000378:metabolism
D051836:COUP Transcription Factors; D004268:DNA-Binding Proteins; D000072159:Dynactin Complex; D005656:Fungal Proteins; D008164:Luminescent Proteins; D008869:Microtubule-Associated Proteins; D011987:Receptors, Steroid; D011993:Recombinant Fusion Proteins; D014157:Transcription Factors; D049452:Green Fluorescent Proteins; D004398:Dyneins; C096113:NUDA protein, Emericella nidulans