The Journal of Experimental Medicine
StemCell Technologies
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Published 19 August 2002. doi:10.1084/jem.20020327
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© Rockefeller University Press, 0022-1007/2002/8/529/ $5.00
The Journal of Experimental Medicine, Volume 196, Number 4, August 19, 2002 529-540
Analysis of Protease Activity in Live Antigen-presenting Cells Shows Regulation of the Phagosomal Proteolytic Contents During Dendritic Cell Activation

Ana-Maria Lennon-Duménil1, Arnold H. Bakker1, René Maehr1, Edda Fiebiger1, Herman S. Overkleeft1, Mario Rosemblatt2, Hidde L. Ploegh1 and Cécile Lagaudrière-Gesbert1

1 Department of Pathology, Harvard Medical School, Boston, MA 02115
2 Departamento de Biologia, Facultad de Ciencias, Universidad de Chile, 6842301 Santiago, Chile

Address correspondence to Hidde L. Ploegh, Department of Pathology, 200 Longwood Avenue, Building 2, Room 137, Boston, MA 02115. Phone: 617-432-4777; Fax: 617-432-4775; E-mail: ploegh{at}hms.harvard.edu

Here, we describe a new approach designed to monitor the proteolytic activity of maturing phagosomes in live antigen-presenting cells. We find that an ingested particle sequentially encounters distinct protease activities during phagosomal maturation. Incorporation of active proteases into the phagosome of the macrophage cell line J774 indicates that phagosome maturation involves progressive fusion with early and late endocytic compartments. In contrast, phagosome biogenesis in bone marrow–derived dendritic cells (DCs) and macrophages preferentially involves endocytic compartments enriched in cathepsin S. Kinetics of phagosomal maturation is faster in macrophages than in DCs. Furthermore, the delivery of active proteases to the phagosome is significantly reduced after the activation of DCs with lipopolysaccharide. This observation is in agreement with the notion that DCs prevent the premature destruction of antigenic determinants to optimize T cell activation. Phagosomal maturation is therefore a tightly regulated process that varies according to the type and differentiation stage of the phagocyte.

Key Words: antigen processing • cathepsin • active site–directed probe • phagocytosis • phagosomal maturation


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