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Science 19 April 2002:
Vol. 296. no. 5567, pp. 503 - 507
DOI: 10.1126/science.1068793
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Research Articles

Multicolor and Electron Microscopic Imaging of Connexin Trafficking

Guido Gaietta,1 Thomas J. Deerinck,1 Stephen R. Adams,2 James Bouwer,1 Oded Tour,2* Dale W. Laird,3 Gina E. Sosinsky,1 Roger Y. Tsien,2* Mark H. Ellisman1dagger

Recombinant proteins containing tetracysteine tags can be successively labeled in living cells with different colors of biarsenical fluorophores so that older and younger protein molecules can be sharply distinguished by both fluorescence and electron microscopy. Here we used this approach to show that newly synthesized connexin43 was transported predominantly in 100- to 150-nanometer vesicles to the plasma membrane and incorporated at the periphery of existing gap junctions, whereas older connexins were removed from the center of the plaques into pleiomorphic vesicles of widely varying sizes. Selective imaging by correlated optical and electron microscopy of protein molecules of known ages will clarify fundamental processes of protein trafficking in situ.

1 National Center for Microscopy and Imaging Research, Department of Neurosciences, 0608,
2 Department of Pharmacology, University of California, San Diego, 9500 Gilman Drive, La Jolla, CA 92093, USA.
3 Department of Anatomy and Cell Biology, University of Western Ontario, London, Ontario Canada, N6A 5C1
*   Howard Hughes Medical Institute, University of California, San Diego, 9500 Gilman Drive, La Jolla, CA 92093-0648 USA.

dagger    To whom correspondence should be addressed. E-mail: mhellisman{at}ucsd.edu

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