Synthetic virus-like particles target dendritic cell lipid rafts for rapid endocytosis primarily but not exclusively by macropinocytosis.
DC employ several endocytic routes for processing antigens, driving forward adaptive immunity. Recent advances in synthetic biology have created small (20-30 nm) virus-like particles based on lipopeptides containing a virus-derived coiled coil sequence coupled to synthetic B- and T-cell epitope mimetics. These self-assembling SVLP efficiently induce adaptive immunity without requirement for adjuvant. We hypothesized that the characteristics of DC interaction with SVLP would elaborate on the roles of cell membrane and intracellular compartments in the handling of a virus-like entity known for its efficacy as a vaccine. DC rapidly bind SVLP within min, co-localised with CTB and CD9, but not caveolin-1. In contrast, internalisation is a relatively slow process, delivering SVLP into the cell periphery where they are maintained for a number of hrs in association with microtubules. Although there is early association with clathrin, this is no longer seen after 10 min. Association with EEA-1(+) early endosomes is also early, but proteolytic processing appears slow, the SVLP-vesicles remaining peripheral. Association with transferrin occurs rarely, and only in the periphery, possibly signifying translocation of some SVLP for delivery to B-lymphocytes. Most SVLP co-localise with high molecular weight dextran. Uptake of both is impaired with mature DC, but there remains a residual uptake of SVLP. These results imply that DC use multiple endocytic routes for SVLP uptake, dominated by caveolin-independent, lipid raft-mediated macropinocytosis. With most SVLP-containing vesicles being retained in the periphery, not always interacting with early endosomes, this relates to slow proteolytic degradation and antigen retention by DC. The present characterization allows for a definition of how DC handle virus-like particles showing efficacious immunogenicity, elements valuable for novel vaccine design in the future.
Institute of Virology and Immunoprophylaxis, Mittelhäusern, Switzerland.
Rajni
R
Sharma
Arin
A
Ghasparian
John A
JA
Robinson
Kenneth C
KC
McCullough
D000818:Animals; D000941:Antigens / Q000737:chemistry; D051242:Caveolin 1 / Q000096:biosynthesis; D002784:Cholesterol / Q000737:chemistry; D002966:Clathrin / Q000378:metabolism; D003713:Dendritic Cells / Q000166:cytology*; D004305:Dose-Response Relationship, Drug; D004705:Endocytosis; D011992:Endosomes / Q000378:metabolism*; D000949:Histocompatibility Antigens Class II / Q000378:metabolism; D006801:Humans; D007477:Ions; D007700:Kinetics; D021962:Membrane Microdomains / Q000737:chemistry* / Q000378:metabolism; D013552:Swine; D060245:Tetraspanin 29 / Q000096:biosynthesis; D014168:Transferrin / Q000378:metabolism; D014771:Virion / Q000378:metabolism*
D000941:Antigens; D051242:Caveolin 1; D002966:Clathrin; D000949:Histocompatibility Antigens Class II; D007477:Ions; D060245:Tetraspanin 29; D014168:Transferrin; D002784:Cholesterol