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First published online 3 February 2004
doi: 10.1242/jcs.00935

Journal of Cell Science 117, 827-836 (2004)
Published by The Company of Biologists 2004
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Research Article

Endocytic trafficking in actively resorbing osteoclasts

Gudrun Stenbeck* and Michael A. Horton

Bone and Mineral Centre, University College London, 5 University Street, London WC1E 6JJ, UK

* Author for correspondence (e-mail: g.stenbeck{at}ucl.ac.uk)

Accepted 9 October 2003

Endocytosis and the subsequent intracellular trafficking of the endocytosed material are important determinants of cellular function. Osteoclasts, cells of the monocyte/macrophage family, are specialized for the internalization and processing of bone matrix. Transcytosis of endocytosed material has been observed in osteoclasts but the precise mechanism controlling this process is unclear. Here, we investigate the regulation of these trafficking events. To establish the directionality and kinetics of trafficking events in resorbing osteoclasts, we devised a system using fluorescent low-molecular-weight markers as probes to follow the route taken by the digested bone matrix. We demonstrate that this route is largely distinct from the pathway followed by proteins taken up by receptor-mediated endocytosis at the basolateral plasma membrane. Endocytosis and transcytosis from the ruffled border are fast processes, with a half-life of the endocytosed material inside the cells of 22 minutes. We demonstrate the crucial role of the microtubule network in transport from the ruffled-border area and provide evidence for a role of the cytoskeleton in the overall efficacy of trafficking. Moreover, we analyse the effect of the V-ATPase inhibitor bafilomycin A1 on endocytic uptake, which gives insight into the pH-dependent regulation of membrane trafficking and resorption in osteoclasts.

Key words: Transcytosis, Microtubules, Actin, Endocytosis, V-ATPase




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© The Company of Biologists Ltd 2004