Ubiquitination regulates MHC class II-peptide complex retention and degradation in dendritic cells

Abstract

The expression and turnover of MHC class II-peptide complexes (pMHC-II) on the surface of dendritic cells (DCs) is essential for their ability to activate CD4 T cells efficiently. The half-life of surface pMHC-II is significantly greater in activated (mature) DCs than in resting (immature) DCs, but the molecular mechanism leading to this difference remains unknown. We now show that ubiquitination of pMHC-II by the E3 ubiquitin ligase membrane-associated RING-CH 1 (March-I) regulates surface expression, intracellular distribution, and survival of pMHC-II in DCs. DCs isolated from March-I-KO mice express very high levels of pMHC-II on the plasma membrane even before DC activation. Although ubiquitination does not affect the kinetics of pMHC-II endocytosis from the surface of DCs, the survival of pMHC-II is enhanced in DCs obtained from March-Ideficient and MHC-II ubiquitination-mutant mice. Using pMHC-IIspecific mAb, we show that immature DCs generate large amounts of pMHC-II that are remarkably stable under conditions in which pMHC-II ubiquitination is blocked. Thus, the cellular distribution and stability of surface pMHC-II in DCs is regulated by ubiquitindependent degradation of internalized pMHC-II.T he initiation of an acquired immune response requires coordinated activation of antigen-specific T cells to provide both immune cell help (in the form of cytokines secreted from CD4 T cells) and immune cell effector function (in the form of cytotoxic CD8 T-cell responses and antigen-specific antibody secretion). This cascade of events is regulated primarily by antigen-presenting cells (APCs) in peripheral tissues that take up foreign antigens, process these antigens into immunogenic peptides, and display these antigenic peptides bound to MHC class II molecules on the APC surface (1). Dendritic cells (DCs) are professional APCs that function to prime naïve T cells. In their resting (or immature) state, DCs are relatively poor stimulators of naïve T cells; however, DC activation by a variety of signals induces a DC maturation cascade that up-regulates expression of peptide-loaded MHC-II complexes (pMHC-II), costimulatory molecules, and chemokine receptors that promote DC migration to secondary lymphoid organs and efficient T-cell activation.Given the central role that pMHC-II expressed on the surface of DCs play in the initiation of immune responses, there is intense interest in understanding the mechanisms leading to immunogenic peptide loading onto MHC-II, pMHC-II transport to the cell surface, and turnover of pMHC-II in DCs. Immature DCs express relatively small amounts of specific pMHC-II on their surface after exposure to antigen (2, 3), and large amounts of MHC-II are retained in intracellular antigen-processing compartments (4). Upon activation of these cells with inflammatory cytokines or Toll-like receptor (TLR) ligands (such as LPS or dsRNA), additional pMHC-II are generated (2, 5), and these pMHC-II are "released" from intracellular stores and traffic to the plasma membrane (6). Curiously, pMHC-II that ar...

MHC Class II Antigen Presentation by Dendritic Cells Regulated through Endosomal Sorting

“…It should be noted, however, that the assays used in these and other studies Watts 1990, 1992) on MHCII endocytosis did not discriminate between MHCII -Ii and pMHCII, nor did they correct for recycling of endocytosed MHCII. Other studies could not confirm a role for ubiquitination in endocytosis of pMHCII (Matsuki et al 2007;Walseng et al 2010a). Interference with ubiquitination-dependent endosomal sorting may increase recycling, and this can also impact on the readout of endocytosis assays.…”

Autoregulation of MARCH1 Expression by Dimerization and Autoubiquitination

“…In mouse and human immature conventional DCs, MARCH1 is responsible for the reduced display of MHC II molecules. Upon maturation following TLR stimulation, MARCH1 expression is shut down to favor Ag presentation (14,18,50). Alternatively, the immunosuppressive cytokine IL-10 upregulates MARCH1 gene expression to minimize the display of MHC II molecules (18,51).…”

Differential Kinetics of Antigen Dependency of CD4+ and CD8+ T Cells

“…In view of our finding that CD4 + and CD8 + T cells differ in their temporal requirements of TCR signals, we asked whether the kinetics of Ag presentation differs for their respective MHC ligands on DCs, the main APCs for priming. Because MHC class II molecules are stabilized on the cell surface upon DC activation by downregulated oligo-ubiquitination of class II b-chains (59, 60) by membrane-associated RING-CHlike E3 ligases (61,62) in vitro (63,64) and in vivo (42,65), we tested whether DC activation affects molecules of both MHC classes similarly. We activated dtg-M and -O DCs in vivo by treatment with a stimulatory CD40 mAb and fed the animals with dox for 24 h subsequently.…”