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Published online 24 July 2000. doi:10.1083/jcb.150.2.377
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© The Rockefeller University Press, 0021-9525/2000/7/377/ $5.00
The Journal of Cell Biology, Volume 150, Number 2, July 24, 2000 377-390


Original Article

A Functional Link between Dynamin and the Actin Cytoskeleton at Podosomes

Gian-Carlo Ochoaa,b, Vladimir I. Slepneva,b, Lynn Neffb,c, Niels Ringstada,b, Kohji Takeia,b, Laurie Daniella,b, Warren Kima,b, Hong Caod, Mark McNivend, Roland Baronb,c, and Pietro De Camillia,b
a Howard Hughes Medical Institute, Yale University School of Medicine, New Haven, Connecticut 06510
b Department of Cell Biology, Yale University School of Medicine, New Haven, Connecticut 06510
c Department of Orthopaedic Surgery, Yale University School of Medicine, New Haven, Connecticut 06510
d Mayo Clinic, Rochester, Minnesota 55905

Correspondence to: Pietro De Camilli, Department of Cell Biology, Yale University School of Medicine, 295 Congress Avenue, New Haven, CT 06510. Tel:(203) 737-4465 Fax:(203) 737-4436 E-mail:pietro.decamilli{at}yale.edu.

Cell transformation by Rous sarcoma virus results in a dramatic change of adhesion structures with the substratum. Adhesion plaques are replaced by dot-like attachment sites called podosomes. Podosomes are also found constitutively in motile nontransformed cells such as leukocytes, macrophages, and osteoclasts. They are represented by columnar arrays of actin which are perpendicular to the substratum and contain tubular invaginations of the plasma membrane. Given the similarity of these tubules to those generated by dynamin around a variety of membrane templates, we investigated whether dynamin is present at podosomes. Immunoreactivities for dynamin 2 and for the dynamin 2–binding protein endophilin 2 (SH3P8) were detected at podosomes of transformed cells and osteoclasts. Furthermore, GFP wild-type dynamin 2aa was targeted to podosomes. As shown by fluorescence recovery after photobleaching, GFP-dynamin 2aa and GFP-actin had a very rapid and similar turnover at podosomes. Expression of the GFP-dynamin 2aaG273D abolished podosomes while GFP-dynaminK44A was targeted to podosomes but delayed actin turnover. These data demonstrate a functional link between a member of the dynamin family and actin at attachment sites between cells and the substratum.

Key Words: clathrin, endocytosis, actin patches, Src, osteoclast


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