Abraham Tulp

Mannose receptor mediated uptake of antigens strongly enhances HLA-class II restricted antigen presentation by cultured dendritic cells.

Dendritic cells (DCs) use macropinocytosis and mannose receptor mediated endocytosis for the uptake of exogenous antigens. Here we show that the endocytosis of the mannose receptor and mannosylated antigen is distinct from that of a non-mannosylated antigen. Shortly after internalization, however, both mannosylated and non-mannosylated antigen are found in an MIIC like compartment. The mannose receptor itself does not reach this compartment, and probably releases its ligand in an earlier endosomal structure. Finally, we found that mannosylation of peptides strongly enhanced their potency to stimulate HLA class II-restricted peptide-specific T cell clones. Our results indicate that mannosylation of antigen leads to selective targeting and subsequent superior presentation by DCs which may be useful for vaccine design.

Synthesis and assembly of MHC-peptide complexes

The assembly of major histocompatibility complex (MHC) class I and class II molecules, and their association with antigenic peptides, is analogous to a production line. Each molecule in the pathway has a specific job to do before it hands over its partially processed product to the next cog in the cellular machine. This tireless molecular workforce has been quietly generating functional MHC class I and class II complexes for millennia, yet it has only been in the past two decades that we have begun to understand how some of the many components work. MHC class I and class II molecules use different intracellular pathways and, consequently, present different peptides (with some exceptions).

Lectin-induced retardation of subcellular organelles during preparative density gradient electrophoresis: Selective purification of plasma membranes

Plasma membranes (PM) are difficult to separate by conventional means from other cellular compartments. Using a density gradient electrophoresis (DGE) apparatus (7 cm, ∅︁ 2.2 cm), mammalian subcellular organelles were separated from a total postnuclear supernatant. The sialic acid‐binding lectin wheat germ agglutinin (WGA) permitted 1.5‐fold electrophoretic retardation of plasma membranes lagging far behind endoplasmic reticulum, endosomes, Golgi and lysosomes (in order of increasing electrophoretic mobility). Mobilities of the latter organelles were not affected by wheat germ agglutinin. The retarded plasma membrane was monitored by surface iodination, the presence of Ca++‐ and Na+/K+‐ATPases and by the presence of clathrin‐coated pits using Western immunoblotting. In the presence of WGA two clathrin‐containing compartments were detected; in the absence of WGA three clathrin populations were seen in the electropherogram: clathrin‐coated vesicles, clathrin‐coated pits (on the PM) and clathrin‐coated structures on the trans‐Golgi network (TGN). Both in the presence and absence of WGA, plasma membrane domains of different electrophoretic mobilities were detected.