Dynamics of cytoplasmic dynein in living cells and the effect of a mutation in the dynactin complex actin-related protein Arp1

Curr Biol. 2000 May 18;10(10):603-6. doi: 10.1016/s0960-9822(00)00488-7.

Abstract

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.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Aspergillus nidulans / genetics
  • Aspergillus nidulans / physiology*
  • COUP Transcription Factors
  • Cytoplasm / metabolism*
  • DNA-Binding Proteins / genetics*
  • DNA-Binding Proteins / metabolism
  • Dynactin Complex
  • Dyneins / genetics
  • Dyneins / metabolism*
  • Fungal Proteins / genetics
  • Fungal Proteins / metabolism
  • Green Fluorescent Proteins
  • Luminescent Proteins / genetics
  • Luminescent Proteins / metabolism
  • Microtubule-Associated Proteins / genetics
  • Microtubule-Associated Proteins / metabolism
  • Mutation*
  • Receptors, Steroid*
  • Recombinant Fusion Proteins / genetics
  • Recombinant Fusion Proteins / metabolism
  • Transcription Factors / genetics*
  • Transcription Factors / metabolism

Substances

  • COUP Transcription Factors
  • DNA-Binding Proteins
  • Dynactin Complex
  • Fungal Proteins
  • Luminescent Proteins
  • Microtubule-Associated Proteins
  • Receptors, Steroid
  • Recombinant Fusion Proteins
  • Transcription Factors
  • Green Fluorescent Proteins
  • Dyneins
  • NUDA protein, Emericella nidulans