Interleukin‐10‐induced MARCH1 mediates intracellular sequestration of MHC class II in monocytes

Abstract

IL-10 is a potent anti-inflammatory cytokine interfering with antigen presentation by inducing the intracellular sequestration of MHC class II (MHC-II) molecules. Here we studied the contribution of membrane-associated RING-CH (MARCH) ubiquitin ligase family members to the IL-10-induced down-regulation of MHC-II molecules. We found that MARCH1 and MARCH8 proteins are the most potent family members for the downregulation of MHC-II surface expression in transfected cells, but only MARCH1 mRNA expression is strongly induced by IL-10 in human primary monocytes. We detected monoand poly-ubiquitinated forms of MHC-II molecules both in IL-10-treated monocytes and in cells transfected with MARCH1. We also show direct interaction between MHC-II and MARCH1 molecules in co-immunoprecipitation assays. Finally, we found that siRNAmediated knockdown of MARCH1 reverses IL-10-induced MHC-II down-regulation in primary monocytes. Thus, the immunosuppressive effect of IL-10 on antigen presentation is mediated through induced expression of MARCH1.

Tolerogenicity is not an absolute property of a dendritic cell

“…Transcription of MARCH1 and MARCH8 was, however, essentially unchanged in all of the populations (B6 data not shown). This contrasts with previous studies using human monocyte-derived DC where downregulation of MARCH1 followed maturation, and a pronounced increase in MARCH1 was seen in response to IL-10 [22,23].…”

“…The E3 ubiquitin ligases MARCH1 and MARCH8 are known to be involved in regulating MHC class II and CD86, with maturationinduced loss of ubiquitination promoting cell surface expression [19][20][21]. Pronounced changes previously reported in the level of MARCH1 mRNA in human monocyte-derived DC, 20-fold on maturation and 40-fold in response to 23], were not evident in our data (Supporting Information Fig. 2A).…”

Autoregulation of MARCH1 Expression by Dimerization and Autoubiquitination

“…For ubiquitination experiments, lysis buffer was supplemented with MG132 and N-ethylmaleimide (18). Cells were lysed on ice and centrifuged.…”

“…Recently, we showed that MARCH1 is induced by IL-10 in human primary monocytes and ubiquitinates MHC II molecules, leading to their intracellular retention and reduced Ag presentation (18). Several studies reported that the endogenous MARCH1 protein is hardly detectable even after an upregulation in mRNA expression (12,18,19).…”

“…Several studies reported that the endogenous MARCH1 protein is hardly detectable even after an upregulation in mRNA expression (12,18,19). Jabbour et al (19) recently estimated that the half-life of MARCH1 is ∼30 min.…”