Abstract
Calreticulin (CRT) is a well-known “eat-me” signal harbored by dying cells participating in their recognition by phagocytes. CRT is also recognized to deeply impact the immune response to altered self-cells. In this study, we focus on the role of the newly exposed CRT following cell death induction. We show that if CRT increases at the outer face of the plasma membrane and is well recognized by C1q even when phosphatidylserine is not yet detected, CRT is also released in the surrounding milieu and is able to interact with phagocytes. We observed that exogenous CRT is endocytosed by THP1 macrophages through macropinocytosis and that internalization is associated with a particular phenotype characterized by an increase of cell spreading and migration, an upregulation of CD14, an increase of interleukin-8 release, and a decrease of early apoptotic cell uptake. Importantly, CRT-induced pro-inflammatory phenotype was confirmed on human monocytes-derived macrophages by the overexpression of CD40 and CD274, and we found that monocyte-derived macrophages exposed to CRT display a peculiar polarization notably associated with a downregulation of the histocompatibility complex of class II molecules hampering its description through the classical M1/M2 dichotomy. Altogether our results highlight the role of soluble CRT with strong possible consequences on the macrophage-mediated immune response to dying cell.
Highlights
It is unambiguously demonstrated that the multifunctional protein calreticulin (CRT) plays an important role in the phagocytic removal of apoptotic cells [1] when exposed at their surface and in the immune response driven by the phagocyte [2]: first acting as an engulfment signal stimulating the recognition of dying cells by phagocytes [3] and second as an element of the antigen presentation machinery of macrophages and dendritic cells (DCs)
We were able to show, by Fluorescence Resonance Energy Transfer (FRET) measurements, that CRT and C1q bind together. This argues in the favor of a physiological function of CRT-dependent C1q opsonization at the very beginning of the programmed cell death process, even though its real effect, either on apoptotic cell uptake or on phagocyte signaling events, remains to be studied in detail
To understand the potential effect of soluble CRT on macrophages, we showed that on THP1 macrophages, the recombinant CRT is efficiently endocytosed by a macropinocytosis-like process
Summary
It is unambiguously demonstrated that the multifunctional protein calreticulin (CRT) plays an important role in the phagocytic removal of apoptotic cells [1] when exposed at their surface and in the immune response driven by the phagocyte [2]: first acting as an engulfment signal stimulating the recognition of dying cells by phagocytes [3] and second as an element of the antigen presentation machinery of macrophages and dendritic cells (DCs). Some of these molecules have been shown to interact together, suggesting that the CRTmediated immune response could be regulated by complex multimolecular interactions
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