Contrasting requirements for ubiquitylation during Fc receptor-mediated endocytosis and phagocytosis.
Fc receptors on leukocytes mediate internalization of antibody-containing complexes. Soluble immune complexes are taken up by endocytosis, while large antibody-opsonized particles are internalized by phagocytosis. We investigated the role of ubiquitylation in internalization of the human FcgammaRIIA receptor by endocytosis and phagocytosis. A fusion of FcgammaRIIA to green fluorescent protein (GFP) was expressed in ts20 cells, which bear a temperature-sensitive mutation in the E1 ubiquitin-activating enzyme. Uptake of soluble IgG complexes mediated by FcgammaRIIA-GFP was blocked by incubation at the restrictive temperature, indicating that endocytosis requires ubiquitylation. In contrast, phagocytosis and phagosomal maturation were largely unaffected when ubiquitylation was impaired. FcgammaRIIA-GFP was ubiquitylated in response to receptor cross-linking. Elimination of the lysine residues present in the cytoplasmic domain of FcgammaRIIA impaired endocytosis, but not phagocytosis. The proteasomal inhibitor clasto-lactacystin beta-lactone strongly inhibited endocytosis, but did not affect phagocytosis. These studies demonstrate a role for ubiquitylation in the endocytosis of immune receptors, and reveal fundamental differences in the mechanisms underlying internalization of a single receptor depending on the size or multiplicity of the ligand complex.
Booth
James W
JW
Cell Biology Program, Hospital for Sick Children and Department of Biochemistry, University of Toronto, Ontario M5G 1X8, Canada.
Kim
Moo-Kyung
MK
Jankowski
Andrzej
A
Schreiber
Alan D
AD
Grinstein
Sergio
S
D017951:Antigen Presentation / Q000502:physiology; D015703:Antigens, CD / Q000502:physiology*; D002460:Cell Line; D004705:Endocytosis / Q000502:physiology*; D049452:Green Fluorescent Proteins; D006801:Humans; D008164:Luminescent Proteins; D010586:Phagocytes / Q000502:physiology*; D010587:Phagocytosis / Q000502:physiology*; D020239:Receptor Cross-Talk; D017452:Receptors, IgG / Q000502:physiology*; D011993:Recombinant Fusion Proteins / Q000502:physiology; D014452:Ubiquitins / Q000378:metabolism*
D015703:Antigens, CD; C080568:Fc gamma receptor IIA; D008164:Luminescent Proteins; D017452:Receptors, IgG; D011993:Recombinant Fusion Proteins; D014452:Ubiquitins; D049452:Green Fluorescent Proteins