Doublecortin association with actin filaments is regulated by neurabin II.
Mutations in the human Doublecortin (DCX) gene cause X-linked lissencephaly, a neuronal migration disorder affecting the neocortex and characterized by mental retardation and epilepsy. Because dynamic cellular asymmetries such as those seen in cell migration critically depend on a cooperation between the microtubule and actin cytoskeletal filament systems, we investigated whether Dcx, a microtubule-associated protein, is engaged in cytoskeletal cross-talk. We now demonstrate that Dcx co-sediments with actin filaments (F-actin), and using light and electron microscopy and spin down assays, we show that Dcx induces bundling and cross-linking of microtubules and F-actin in vitro. It has recently been shown that binding of Dcx to microtubules is negatively regulated by phosphorylation of the Dcx at Ser-47 or Ser-297. Although the phosphomimetic green fluorescent protein (GFP)-Dcx(S47E) transfected into COS-7 cells had a reduced affinity for microtubules, we found that pseudophosphorylation was not sufficient to cause Dcx to bind to F-actin. When cells were co-transfected with neurabin II, a protein that binds F-actin as well as Dcx, GFP-Dcx and to an even greater extent GFP-Dcx(S47E) became predominantly associated with filamentous actin. Thus Dcx phosphorylation and neurabin II combinatorially enhance Dcx binding to F-actin. Our findings raise the possibility that Dcx acts as a molecular link between microtubule and actin cytoskeletal filaments that is regulated by phosphorylation and neurabin II.
Tsukada
Miki
M
Max Planck Institute for Experimental Endocrinology, Feodor-Lynen-Strasse 7, D-30625 Hannover, Germany. miki.tsukada@mpihan.mpg.de
Prokscha
Alexander
A
Ungewickell
Ernst
E
Eichele
Gregor
G
D000199:Actins / Q000378:metabolism*; D000818:Animals; D019556:COS Cells; D000089622:Doublecortin Domain Proteins; D000089662:Doublecortin Protein; D051379:Mice; D008840:Microfilament Proteins / Q000502:physiology*; D008869:Microtubule-Associated Proteins / Q000378:metabolism*; D008870:Microtubules / Q000378:metabolism; D009419:Nerve Tissue Proteins / Q000502:physiology*; D009479:Neuropeptides / Q000378:metabolism*; D010766:Phosphorylation; D000099273:Neurabins
D000199:Actins; D000089622:Doublecortin Domain Proteins; D000089662:Doublecortin Protein; D008840:Microfilament Proteins; D008869:Microtubule-Associated Proteins; D009419:Nerve Tissue Proteins; D009479:Neuropeptides; D000099273:Neurabins; C000714328:DCX protein, human; C000714847:Dcx protein, mouse; C079240:Mtap2 protein, mouse