Modulation of Rac localization and function by dynamin.
The GTPase dynamin controls a variety of endocytic pathways, participates in the formation of phagosomes, podosomal adhesions, and invadopodia, and in regulation of the cytoskeleton and apoptosis. Rac, a member of the Rho family of small GTPases, controls formation of lamellipodia and focal complexes, which are critical in cell migration and phagocytosis. We now show that disruption of dynamin(-2) function alters Rac localization and inhibits cell spreading and lamellipodia formation even though Rac is activated. Dominant-negative K44A dynamin(-2) inhibited cell spreading and lamellipodia formation on fibronectin without blocking cell adhesion; dynamin(-2) depletion by specific small interfering RNA inhibited lamellipodia in a similar manner. Dyn2(K44A) induced Rac mislocalization away from cell edges, into abnormal dorsal ruffles, and led to increased total Rac activity. Fluorescence resonance energy transfer imaging of Rac activity confirmed its predominant localization to aberrant dorsal ruffles in the presence of dominant-negative dyn2(K44A). Dyn2(K44A) induced the accumulation of tubulated structures bearing membrane-bound Rac-GFP. Constitutively active but not wild-type GFP-Rac was found on macropinosomes and Rac-dependent, platelet-derived growth factor-induced macropinocytosis was abolished by Dyn2(K44A) expression. These data suggest an indispensable role of dynamin in Rac trafficking to allow for lamellipodia formation and cell spreading.
Schlunck
Günther
G
Division of Vascular Biology, The Scripps Research Institute, La Jolla, California 92037, USA.
Damke
Hanna
H
Kiosses
William B
WB
Rusk
Nicole
N
Symons
Marc H
MH
Waterman-Storer
Clare M
CM
Schmid
Sandra L
SL
Schwartz
Martin Alexander
MA
D000818:Animals; D002448:Cell Adhesion / Q000502:physiology; D002465:Cell Movement / Q000502:physiology*; D002478:Cells, Cultured; D034285:Dynamin II / Q000187:drug effects / Q000378:metabolism* / Q000502:physiology; D005353:Fibronectins / Q000378:metabolism; D031541:Fluorescence Resonance Energy Transfer; D051379:Mice; D009154:Mutation; D041681:NIH 3T3 Cells; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism* / Q000502:physiology; D051057:Proto-Oncogene Proteins c-akt; D011554:Pseudopodia / Q000378:metabolism*; D034741:RNA, Small Interfering / Q000494:pharmacology; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000187:drug effects / Q000378:metabolism / Q000502:physiology
D005353:Fibronectins; D034741:RNA, Small Interfering; D011993:Recombinant Fusion Proteins; D017346:Protein Serine-Threonine Kinases; D051057:Proto-Oncogene Proteins c-akt; D034285:Dynamin II